Inter-site variation in allometry and wood density of Goupia glabra Aubl. in Amazonia.

The present study aims to compare the allometry and wood density of Goupia glabra Aubl. (Goupiaceae) in two different terra-firme sites in Amazonian forest. A total of 65 trees ≥ 10 cm DBH was sampled in both sites, with 39 trees in Nova Olinda do Norte (NOlinda, near the Amazon River) and 29 trees in Apuí (near the southern edge of the Amazon forest). Except for the relationship between DBH (diameter at breast height) and Ht (total height), allometric relationships for G.glabra differed significantly between sites. Apuí had lower intercept and greater slope for log10 (DBH) versus log10 (Hs - stem height), and, conversely, greater intercept and lower slope for log10 (DBH) versus log10 (Ch - crown height). The slope differed significantly between the sites for DBH versus Cd (crown diameter), with greater slope found for NOlinda. Mean basic wood density in Apuí was 8.8% lower than in NOlinda. Our findings highlight the variation in adaptive strategy of G. glabra due to environmental differences between sites. This is probably because of different canopy-understory light gradients, which result in differentiation of resource allocation between vertical and horizontal growth, which, in turn, affects mechanical support related to wood density. We also hypothesize that differences in soil fertility and disturbance regimes between sites may act concomitantly with light.

Inter-site variation in allometry and wood density of Goupia glabra Aubl. in Amazonia / Variação entre-sítios na alometria e densidade da madeira de Goupia glabra Aubl. na Amazônia

Abstract The present study aims to compare the allometry and wood density of Goupia glabra Aubl. (Goupiaceae) in two different terra-firme sites in Amazonian forest. A total of 65 trees ≥ 10 cm DBH was sampled in both sites, with 39 trees in Nova Olinda do Norte (NOlinda, near the Amazon River) and 29 trees in Apuí (near the southern edge of the Amazon forest). Except for the relationship between DBH (diameter at breast height) and Ht (total height), allometric relationships for G.glabra differed significantly between sites. Apuí had lower intercept and greater slope for log10 (DBH) versus log10 (Hs – stem height), and, conversely, greater intercept and lower slope for log10 (DBH) versus log10 (Ch – crown height). The slope differed significantly between the sites for DBH versus Cd (crown diameter), with greater slope found for NOlinda. Mean basic wood density in Apuí was 8.8% lower than in NOlinda. Our findings highlight the variation in adaptive strategy of G. glabra due to environmental differences between sites. This is probably because of different canopy-understory light gradients, which result in differentiation of resource allocation between vertical and horizontal growth, which, in turn, affects mechanical support related to wood density. We also hypothesize that differences in soil fertility and disturbance regimes between sites may act concomitantly with light.

Resumo O presente estudo tem como objetivo comparar a alometria e a densidade da madeira de Goupia glabra em dois diferentes sítios de floresta de terra firme na Amazonia. Um total de 65 árvores com DAP ≥ 10 cm foi amostrado em ambos os sítios, sendo 39 árvores em Nova Olinda do Norte (NOlinda, próximo ao rio Amazonas) e 29 em Apuí (próximo à borda sul da Amazônia). Exceto para a relação entre o DBH (diâmetro a altura do peito) e a Ht (altura total), as relações alométricas para G. glabra diferiu significativamente entre os sítios. Apuí apresentou menor intercepto e maior inclinação para a relação log10 (DBH) versus log10 (Hs – altura do fuste) e, ao contrário, maior intercepto e menor inclinação para log10 (DBH) versus log10 (Ch – altura da copa). A inclinação diferiu significativamente entre os sítios para DBH versus Cd (diâmetro da copa), com maior inclinação encontrada para NOlinda. A densidade básica média da madeira in Apuí foi 8.8% menor do que em NOlinda. Os resultados deste estudo destacam a variação na estratégia adaptativa de G. glabra devido às diferenças ambientais entre os sítios. Isto é provavelmente consequência dos diferentes gradientes de luz o que resulta na diferenciação na alocação de recursos entre o crescimento vertical e horizontal o que, por sua vez, afeta o suporte mecânico relacionado à densidade da madeira. Nós também levantamos a hipótese de que as diferenças em termos de fertilidade e regimes de distúrbios entre os sítios podem agir concomitantemente com o regime de luz.

Carnitine-related alterations in patients with intermittent claudication: indication for a focused carnitine therapy.

BACKGROUND: Carnitine metabolism is altered in peripheral arterial disease. L-carnitine supplementation may correct these alterations and improve walking performance. METHODS AND RESULTS: Plasma levels of carnitine and its esters were measured at rest and after maximally tolerated exercise in 22 claudicant patients and 8 normal subjects. One week later, this protocol was repeated in patients after random administration of placebo or L-carnitine (500 mg IV as a single bolus). Two groups of patients emerged. In 10 patients (group IC1), the plasma level of acetylcarnitine at rest was 3.7 +/- 0.2 micromol/L and increased significantly (P<.01) at maximally tolerated exercise. In 12 patients (group IC2), the resting level of plasma acetylcarnitine was elevated (7.9 +/- 0.7 micromol/L, P<.01) and decreased with exercise. Furthermore, group IC2 patients had a significantly lower walking capacity than group IC1 patients. In both groups, placebo did not affect the metabolic profile, nor did it improve exercise performance. Conversely, after L-carnitine administration, all but one patient in group IC2 (n=7) showed an increase in plasma acetylcarnitine concentration during exercise versus the decrease observed without L-carnitine. This metabolic effect was accompanied by a significant increase (P<.01) in walking capacity. Interestingly, in group IC1 patients (n=5), L-carnitine neither improved walking capacity nor modified the metabolic profile. Statistical analysis showed that changes in walking capacity with L-carnitine treatment were influenced exclusively by exercise-induced changes in plasma acetylcarnitine. CONCLUSIONS: In patients with intermittent claudication, assessment of plasma acetylcarnitine at rest and after exercise may be a means to select a target population for L-carnitine therapy.

Mitochondrial alterations induced by aspirin in rat hepatocytes expressing mitochondrially targeted green fluorescent protein (mtGFP).

Mitochondria in primary living hepatocytes were visualized in cells transfected with a chimeric plasmid encoding for the green fluorescent protein (GFP) of Aequorea victoria engineered to be specifically targeted to mitochondria, as described recently (Rizutto et al. (1995) Curr. Biol. 5, 635-642). The identification of the fluorescent organelles as authentic mitochondria was confirmed by double labeling with rhodamine 123. Acetylsalicylate treatment of hepatocytes induced in mitochondria typical morphological alterations closely analogous to the swelling promoted by acetylsalicylate in isolated mitochondria. Cyclosporin A, which in isolated mitochondria prevents the changes induced by acetylsalicylate, had no protective action but induced per se specific alterations in the morphology of mitochondria. Moreover, exposure of hepatocytes to cyclosporin A followed by acetylsalicylate caused the same mitochondrial changes induced by each of the two compounds separately. The structural alterations caused by acetylsalicylate were constantly associated with a decrease in mitochondrial urea synthesis and cell viability.

The alterations in the energy linked properties induced in rat liver mitochondria by acetylsalicylate are prevented by cyclosporin A or Mg2+.

The alterations in rat liver mitochondria induced by acetylsalicylate in the presence of low concentrations of Ca2+ (large amplitude swelling, permeability to 14C]sucrose, collapse of transmembrane potential and effluxes of endogenous Mg2+ and accumulated Ca2+) were fully prevented by either cyclosporin A or Mg2+. Cyclosporin A and Mg2+ were also capable of restoring transmembrane potential upon its decrease induced by acetylsalicylate. The loss of endogenous Mg2+ was the primary effect promoted by acetylsalicylate; the other noxious effects followed. These results indicate that Mg2+ are fundamental components of the mitochondrial permeability barrier and that their loss might be responsible for the membrane transition induced by acetylsalicylate.

Specific degradation of troponin T and I by mu-calpain and its modulation by substrate phosphorylation.

The degradation of troponin (Tn) subunits by calpain was studied by incubating either isolated cardiac Tns or myocardial cryosections with two different calpain isoenzymes isolated from rat skeletal muscle. Western-blot analysis with monoclonal antibodies against TnI and TnT showed that mu-calpain was at least ten times more active than m-calpain in degrading TnI and TnT both in vitro and in situ. TnC was completely resistant to both proteinase forms. Phosphorylation by cyclic AMP-dependent protein kinase (PKA) isolated from rat skeletal muscle reduced the sensitivity of TnI to degradation. This effect in combination with an increased efficiency of the endogenous inhibitor [Salamino, De Tullio, Michetti, Mengotti, Melloni and Pontremoli (1994) Biochem. Biophys. Res. Commun. 199, 1326-1332] probably reduces the proteolytic activity of calpain in cells on PKA stimulation. Conversely, phosphorylation by protein kinase C (PKC) resulted in a twofold increase in the degradation of TnI. Degradation by m-calpain was not modified by Tn phosphorylation. The different sensitivity to mu-calpain might be related to changes in TnI oligomeric structure. Indeed, on PKC phosphorylation, the apparent molecular mass of TnI calculated from the distribution coefficient of Tn complex in Sephadex G-100 matrix was reduced from 90 to 30 kDa suggesting dissociation of the Tn complex.

Inhibition of macroautophagy and proteolysis in the isolated rat hepatocyte by a nontransportable derivative of the multiple antigen peptide Leu8-Lys4-Lys2-Lys-beta Ala.

The multiple antigen peptide derivative, Leu8-Lys4-Lys2-Lys-beta Ala (Leu8-MAP), was synthesized by attaching the carboxyl of leucine to the NH2 termini of a branched lysine core, termed MAP, creating a molecule of about 1900 Da with 8 leucine residues. On a molar basis (independent of the number of leucine substitutions), Leu8-MAP was as effective as leucine in suppressing macroautophagy and proteolysis; moreover, it exhibited the same apparent Km (about 0.1 mM). The effect was specific for leucine since Ile8-MAP was inactive. It is of interest, though, that Leu8-MAP did not elicit the multiphasic response typical of leucine but instead evoked the single site inhibition normally seen with leucine plus the co-regulator alanine. Some free leucine was produced from Leu8-MAP during hepatocyte incubations, but the amounts were insufficient to account for the inhibition. Although this degradation created species of Leu-MAP that had lost 1-3 residues of leucine, their inhibitory effectiveness was not diminished. Because the extracellular/intracellular distribution ratio of [3H]-Leu8-MAP was 100:1 or greater, the direct transport of Leu8-MAP across the plasma membrane into the cytosolic compartment can be excluded. Hence, cytosolic concentrations of Leu8-MAP will be at least 100-fold smaller than those of leucine under conditions of comparable proteolytic inhibition. For these and related reasons, effects attributable to the recognition of Leu8-MAP cannot be explained by signals generated within the cytosol. They could, however, be mediated from site(s) on the plasma membrane or within associated vesicles.

Involvement of erythrocyte calpain in glycine- and carnitine-treated isovaleric acidemia.

When a 12-y-old girl suffering from isovaleric acidemia was treated with L-carnitine, there was a considerable increase in her blood and urine concentration of isovalerylcarnitine. When later the patient received an infusion of glycine in place of carnitine, isovalerylcarnitine reverted toward the low levels found in a normal subject. At the end of either treatment, erythrocyte calpain was measured and found to be decreased after carnitine therapy (140 versus 96 U/mg Hb with glycine or carnitine, respectively). Because we have previously shown that the activity of calpain isolated from erythrocytes was markedly modified by isovalerylcarnitine, the present results might be seen as the consequence of the chronic exposure of the patient's red blood cells to high levels of isovalerylcarnitine. The lowered calpain activity was also proved by an increase in erythrocyte band 3 phosphorylation together with an increased erythrocyte fragility after calcium loading in the presence of the ionophore A-23187. Calpastatin, the natural inhibitor of calpain, was only slightly modified.

Contrasting effects of propionate and propionyl-L-carnitine on energy-linked processes in ischemic hearts.

Propionyl-L-carnitine, unlike L-carnitine, is known to improve myocardial function and metabolism altered during the course of ischemia-reperfusion. In this study, the effect of propionyl-L-carnitine has been compared with that of propionate and carnitine on the performance of rat hearts perfused with a glucose-containing medium either under normoxia, ischemia, or postischemic reperfusion. In the postischemic phase, contractile parameters were partially restored both in the control and in the propionate plus carnitine-treated hearts, were markedly impaired by propionate, and were fully recovered by propionyl-L-carnitine. In addition, propionyl-L-carnitine, but not propionate, reduced the functional decay of mitochondria prepared from the ischemic hearts. Even in normoxic conditions propionate, unlike propionyl-L-carnitine, caused a drastic reduction of free CoA and L-carnitine. The concomitant increase in lactate production and decrease in ATP content might be explained by the inhibition of pyruvate dehydrogenase caused by the accumulation of propionyl-CoA. Indeed, when pyruvate was the only oxidizable substrate, propionate induced a gradual decrease in developed pressure, which was largely prevented by L-carnitine. The protective effect of propionyl-L-carnitine may be a consequence of the anaplerotic utilization of propionate in the presence of an optimal amount of ATP and free L-carnitine.

[Support for the parents after the death of a newborn infant]. / Supporto ai genitori dopo la morte di un neonato.

The neonatologist in NICU has many duties, first of all caring the baby and supporting the parents in facing stressing situations. When the baby dies most doctors think their job is over, and only in few hospitals there is the opportunity for the parents to meet the staff again. We report our recent experience to meet parents after baby's death. We offer them this possibility when they are leaving the hospital and, after about one month, we call them by phone to arrange an appointment. We have realized that they need to talk at least once with the staff (doctors and nurses) to examine and solve doubts about cares and to relieve their sufferance. In our experience, even limited, we found that nobody refuses this opportunity and that in most instances parents required more than one meeting and, finally, that talking of the baby with people who took care of him helps them in accepting baby's death. We found this experience very useful both for parents and staff, so we hope this opportunity will extend to other hospitals.

Spermine-mediated casein kinase II-uptake by rat liver mitochondria.

Spermine, ubiquitous intracellular polyamine, is able to promote the transmembrane translocation of casein kinase CKII through the outer membrane of rat liver mitochondria and its binding to more internal mitochondrial structures. These findings suggest that spermine may play a critical role in regulating the subcellular distribution of casein kinase CKII.

Multiphasic control of proteolysis by leucine and alanine in the isolated rat hepatocyte.

Autophagically mediated proteolysis in the perfused rat liver is under complex multiphasic control by a small group of amino acids dominated by leucine. Because there have been no prior reports of such regulation in the isolated hepatocyte, our goal was to determine whether it is a manifestation of interactions between diverse cells in the intact liver or, alternatively, the expression of a unique control mechanism within a single population of cells. Hepatocytes were isolated from livers of ad libitum-fed rats and incubated with cycloheximide at low density (approximately 10(6) cells/ml) for the determination of valine release. As in perfusion experiments with synchronously fed rats, proteolytic responses to leucine in cells from fed rats were mediated through two inhibitory mechanisms that alternated randomly on a day-to-day basis. The first (L) represented a typical multiphasic dose-response with low- and high-concentration inhibition separated by a sharp zonal loss of inhibition that could be abolished by alanine. The second (H) mediated inhibition only at high concentrations. It disappeared after 24 h of starvation, leaving L as the prevailing mode. The findings indicate that both macroautophagy and the multiphasic mechanism for regulating it coexist in a single population of hepatocytes, making the cells suitable for studies aimed at defining the putative plasma membrane site of leucine recognition.

Control of hepatic proteolysis by leucine and isovaleryl-L-carnitine through a common locus. Evidence for a possible mechanism of recognition at the plasma membrane.

Deprivation-induced proteolysis in the perfused rat liver is controlled through the multiphasic action of 7 regulatory amino acids of which L-leucine plays the dominant role. Recently, isovaleryl-L-carnitine (IVC) was shown to mimic the leucine's effects, suggesting that the two molecules share structural features that are recognized at a common site(s). In this study we find that each evokes identical responses consisting of inhibitory effects at 0.08 and 0.8 mM, separated by a sharp zonal loss of inhibition at 0.15 mM. As monitored by density shifts of beta-hexosaminidase in colloidal silica gradients, macroautophagy is suppressed by both. Responses to Leu and IVC at 0.08 and 0.15 mM are stereospecific and require a reactive group at the alpha-carbon (or equivalent) and a high degree of branched chain specificity. In addition, 0.5 mM Ala coregulates with IVC and Leu by decreasing the zonal loss at 0.15 mM. The fact that the multiphasic responses can be duplicated with equimolar mixtures of Leu + IVC indicates that both react at the same site(s). IVC is readily taken up by a saturable process, but owing to its rapid hydrolysis in the cell, the ratio of internal to external IVC remains low over a 4-fold concentration range. These findings, together with a kinetic analysis of concerted responses to regulatory amino acids, suggest that the recognition sites are at a position in the cell, possibly at the plasma membrane, to react reversibly with plasma amino acids.

Effects of palmitoyl CoA and palmitoyl carnitine on the membrane potential and Mg2+ content of rat heart mitochondria.

Palmitoyl CoA and palmitoyl carnitine added to rat heart mitochondria in amounts above 20 and 50 nmoles/mg protein, respectively, induced a fall in transmembrane potential and loss of endogenous Mg2+. The dissipation of membrane potential by low concentrations of palmitoyl CoA in the presence of Ca2+, but not that of high concentrations of palmitoyl CoA alone, was prevented by either ruthenium red, Cyclosporin A or Mg2+, but reversed only by Mg2+. The fall of membrane potential induced by palmitoyl carnitine was not prevented by any of these factors. It is suggested that the action of both palmitoyl CoA and palmitoyl carnitine at high concentrations is due to a non specific disruption of membrane architecture, while that of low concentrations of palmitoyl CoA in the presence of Ca2+ is associated specifically with energy dissipation due to Ca2+ cycling.

[Obstetric/neonatological integration and care of healthy newborn infants]. / L'integrazione ostetrico/neonatologica e l'assistenza al neonato sano.

We report how we changed the model of the organization and the assistance in our Department of healthy newborns (2200-2400/years). After we have realized that mothers were not satisfied of the rules of the hospital and personnel was not satisfied of the job, we decided to begin a process of analysis and review of the procedures on full term newborn. During this process we found out that the most important thing was to have clear in mind the problems and the needs of the mother and the baby, and not those of nurses and doctors. A similar process took place in the Department of Obstetrics. In this way we, Obstetrics and Neonatologist together, began to offer a more human approach to birth, and rooming-in began. We stopped to attend every normal delivery, to separate immediately mother and baby, to feed the baby at fixed time, to give him supplementations. We tried to have with the mother a better relationship, visiting the baby in presence of the mother an receiving grom Obstetrics as soon as possible every information about pregnancy. We realized that this was possible only if the Neonatologist and the Obstetric were of the same opinion about a more human approach to birth. We stress this point, well aware that it's impossible to reach this goal unless everybody in any way involved in birth work in great harmony with all the others. A further result of this "new" way of working has been the program of early discharge: if desired, and whenever possible, the mother and the baby go home 48-72 hours after delivery. We report here preliminary data.

Propionyl-L-carnitine: biochemical significance and possible role in cardiac metabolism.

Propionyl-CoA is formed principally during amino acid catabolism. It is then converted chiefly to succinate in a described three-step sequence. Free propionate is formed from propionyl-CoA to a very limited extent, but this anion can participate in a futile cycle of activation and hydrolysis, which can significantly deplete mitochondrial ATP. Free CoA and propionyl-CoA cannot enter or leave mitochondria, but propionyl groups are transferred between separate CoA pools by prior conversion to propionyl-L-carnitine. This reaction requires carnitine and carnitine acetyl transferase, an enzyme abundant in heart tissue. Propionyl-L-carnitine traverses both mitochondrial and cell membranes. Within the cell, this mobility helps to maintain the mitochondrial acyl-CoA/CoA ratio. When this ratio is increased, as in carnitine deficiency states, deleterious consequences ensue, which include deficient metabolism of fatty acids and urea synthesis. From outside the cell (in blood plasma), propionyl-L-carnitine can either be excreted in the urine or redistributed by entering other tissues. This process apparently occurs-without prior hydrolysis and reformation. It is suggested that heart tissue utilizes such exogenous propionyl-L-carnitine to stimulate the tricarboxylic acid cycle (via succinate synthesis) and that this may explain its known protective effect against ischemia.

Transport and action of spermine in rat heart mitochondria.

At concentrations of 0.5-1.0 mM, spermine fully prevents the fall of membrane potential induced in rat heart mitochondria either by aging at room temperature or by the addition of palmitoyl CoA. Spermine also prevents the inhibitory action of palmitoyl CoA on adenylate translocase activity. When added to heart mitochondria de-energized by the same damaging conditions (aging or addition of palmitoyl CoA) spermine restores both membrane potential (provided that ATP is also added) and the activity of adenylate translocase. A part of added spermine is immediately bound to anionic sites on mitochondrial membranes, another part is slowly transported into heart mitochondria. Whereas binding is an energy independent process, transport is driven by the transmembrane potential. Spermine penetrates the mitochondrial matrix at significant rates only at high membrane potential, such as that produced either by phosphate transport or addition of nigericin.

Protein phosphorylation in rat liver mitochondria.

Incubation of rat liver mitochondria in the presence of either [32P] Pi or [y32-P] ATP resulted in a phosphorylation of four proteins with Mr 50, 47, 44 and 36 kDa, respectively. The endogenous phosphorylation of these proteins in the presence of [32P] Pi was markedly influenced by the osmolarity of the incubation medium and differentially affected by various effectors of mitochondrial functions, such as Ca2+, oligomycin, FCCP, arsenite and dichloroacetate. In particular, the 36 kDa protein, unlike the other proteins, appears to be phosphorylated also by direct incorporation of [32P], independently of respiratory chain-linked ATP synthesis. The four proteins, located in the mitoplasts, seem to be phosphorylated by different protein kinases, as suggested by the observation that the endogenous phosphorylation of 36 kDa protein resulted selectively increased by addition of exogenous protein kinases, such as casein kinases S and TS. A tentative identification of these phosphorylatable protein is discussed.