RESUMO
The effects of a high glucose concentration (HGC) on renal phosphatidylcholine (PtdCho) biosynthesis were studied. In control rats, HGC increased papillary PtdCho biosynthesis. In chronic diabetic rats, an increase above that induced by diabetes was observed. Such glucose-responsive phospholipid pools were shown to be transient in adult control rats, while in acute diabetic and aged control and chronic diabetic rats they seem to be of slow breakdown or permanent. Deoxyglucose evokes the HGC effect only in the presence of 5 mM glucose. Neomycin, which blocks phospholipase C action, corrected the HGC effect in control and chronic diabetic rats, but not the increase due to diabetes. CDP-choline: 1,2-diacylglycerol cholinephosphotransferase activity was increased by both in vivo and simulated diabetes. Therefore, transient high extracellular glucose levels promote a reversible increase in papillary (32)P-PtdCho, while diabetes causes an irreversible increase resulting in PtdCho accumulation, possibly related to papillary necrosis.
Assuntos
Glicemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Medula Renal/metabolismo , Fosfatidilcolinas/biossíntese , Animais , Desoxiglucose/metabolismo , Medula Renal/efeitos dos fármacos , Masculino , Microssomos/metabolismo , Neomicina/farmacologia , Fosfatidilcolinas/metabolismo , Ratos , Ratos WistarRESUMO
Phosphatidylinositol (PtdIns) synthesis and polyphosphoinositide (PPI) formation were measured as the incorporation of [32P]orthophosphate ([32P]Pi) or [3H]inositol into non-stimulated intact human neutrophil membrane phospholipids. The rate of PtdIns "de novo" synthesis appeared to be a slow mechanism when compared to the rapid incorporation of [32P]Pi into PPIs. Of the "de novo" synthesized [3H]PtdIns, 70% was further phosphorylated to PPI. Nevertheless, this PPI pool represented less than 0.01% of the total nmols of PPIs formed evaluated as [32P]Pi labeling, indicating that PPI formation mainly involves a no "de novo" synthesized phosphatidylinositol pool. When evaluated at short incubation times, oscillations in the formation of PPIs were detected. A rapid phase was characterized after 30 s of incubation with [32P]Pi Phosphorylation levels returned to an equilibrium state within a minute, and the second phase peaked at 5 min., returning to equilibrium at 15 min. The fluctuant kinetics though not the equilibrium level of PPI formation, could be abolished by neomycin. On the other hand, a selective inhibition of the rapid phase of PPI synthesis occurred in the presence of the tyrosine kinase inhibitor genistein. When the incorporations of [gamma-32P]-adenosine triphosphate (ATP) or [32P]Pi into human neutrophil particulate fraction membranes were evaluated, PPIs synthesis showed fluctuations independently of the precursor used. Noticeably, [32P]from [32P]Pi was incorporated more efficiently into PPIs than that from [gamma-32P]ATP, when evaluated in parallel using equal specific activities for both radiolabeled precursors and under non-ATP synthesizing conditions. Moreover, the incorporation of [32P]Pi into particulate fraction PPIs was not abolished by high concentrations of non-radiolabeled ATP, and metabolically inhibited PMNs showed high rates of PPI synthesis. These data suggest that PPI formation is not necessarily a futile cycle in PMNs.
Assuntos
Lipídeos de Membrana/biossíntese , Neutrófilos/metabolismo , Fosfatos de Fosfatidilinositol/biossíntese , Trifosfato de Adenosina/metabolismo , Adulto , Antimetabólitos/farmacologia , Desoxiglucose/farmacologia , Difosfatos/metabolismo , Metabolismo Energético/efeitos dos fármacos , Genisteína/farmacologia , Humanos , Cinética , Magnésio/farmacologia , Neomicina/farmacologia , Neutrófilos/efeitos dos fármacos , Oligomicinas/farmacologia , Fosfatidilinositóis/biossíntese , Fosforilação , Proteínas Tirosina Quinases/antagonistas & inibidoresRESUMO
Rat renal papilla is the zone of the kidney enjoying the most active phospholipid metabolism and also the highest prostaglandin production. We studied the phospholipid biosynthesis and the relationship between phospholipid de novo synthesis and prostaglandin biosynthesis in rat renal papilla. Indomethacin inhibited the biosynthesis of phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine and phosphatidic acid. Exogenous PGF2alpha and PGD2 restored biosynthetic activity in the presence of indomethacin and also increased the activity of the enzymes involved in the Kennedy pathway. The decrease in phospholipid biosynthesis maintained a linear relationship with the decrease in prostaglandin biosynthesis. Moreover, esculetin, which stimulates prostaglandin synthesis, brought about a significant increase in 32P incorporation to the three phospholipids studied. The evidence presented in this paper indicates that renal PGF2alpha and PGD2 modulate phospholipid de novo synthesis in rat renal papilla.
Assuntos
Medula Renal/metabolismo , Fosfolipídeos/biossíntese , Prostaglandinas/fisiologia , Animais , Indometacina/farmacologia , Masculino , Ácido Palmítico/metabolismo , Ratos , Ratos Wistar , Umbeliferonas/farmacologiaRESUMO
The biosynthesis of prostaglandins (PGs) from the endogenous and exogenous precursor, arachidonic acid (AA), in renal papilla, medulla and cortex from neonatal to adult rats was investigated. Rat renal papilla and medulla incubated in the presence of [1-14C]AA released radioactive PGE2, PGF2 alpha and PGD2 which increased with age. No radioactive prostaglandins were found in the supernatants of renal cortex at any age studied. The amount of total prostaglandins released from the endogenous precursor also increased from 10 to 70 days of age, PGD2 being the prostaglandin that showed the most important rise. In the cortex, only PGE2 release increased with age. Cyclooxygenase (COX) activity was measured in papillary, medullary and cortical homogenates by using [1-14C]AA as substrate. Papillary and medullary COX activity increased after 10 days of age and continued to rise up to day 30 thereafter remaining unaltered until adulthood. Cortical COX activity was very low and decreased with age. These findings indicate the low capacity of the neonatal rat kidney to synthesize PGs.
Assuntos
Rim/crescimento & desenvolvimento , Rim/metabolismo , Prostaglandinas/biossíntese , Animais , Ácido Araquidônico/metabolismo , Dinoprosta/biossíntese , Dinoprostona/biossíntese , Córtex Renal/metabolismo , Medula Renal/metabolismo , Prostaglandina D2/biossíntese , Prostaglandina-Endoperóxido Sintases/metabolismo , Ratos , Ratos WistarRESUMO
Phospholipid content and metabolism were studied in rat renal papillary, medullary and cortical slices. The highest concentration of phospholipids was found in cortex and the lowest in papilla samples (ratio cortex/medulla, 1.3; cortex/papilla, 3.7). The profile of the various phospholipids was different depending on the zone. The most important difference was the relative concentrations of sphingomyelin (CerPCho) and phosphatidylinositol (PtdIns) with ratios for PtdIns/CerPCho of 5.0, 3.3 and 2.5 in papilla, medulla, and cortex, respectively. In the three zones, PtdIns showed the highest specific activity for [2-14C]glycerol and [1-14C]arachidonic acid incorporation. By contrast, a higher amount of [1-14C]palmitic acid was incorporated into phosphatidylcholine than into any other phospholipid. The various radioactive precursors were only poorly incorporated into phosphatidylethanolamine. No radioactivity was associated with phosphatidylserine. The papilla possesses the most active phospholipid metabolism of all the pathways studied.
Assuntos
Ácidos Graxos/metabolismo , Rim/metabolismo , Fosfolipídeos/metabolismo , Animais , Ácido Araquidônico/metabolismo , Técnicas In Vitro , Rim/química , Córtex Renal/química , Córtex Renal/metabolismo , Medula Renal/química , Medula Renal/metabolismo , Pelve Renal/química , Pelve Renal/metabolismo , Masculino , Ácido Palmítico , Ácidos Palmíticos/metabolismo , Fosfolipídeos/análise , Ratos , Ratos Endogâmicos , Distribuição TecidualRESUMO
In view of the fact that mepacrine (Mp) is usually used as an inhibitor of the endogenous phospholipase A2, and since this enzyme produces the release of arachidonic acid (AA) from membrane phospholipids, we studied the effect of different concentrations of Mp on the mobilization of [1-14C]AA in rat renomedullary phospholipids. During the acylation period, 0.1 mM Mp did not produce any significant change in the incorporation of [1-14C]AA into phosphatidylcholine (PC) and phosphatidylethanolamine (PE), and only a slight increase in phosphatidylinositol (PI). Higher concentrations of Mp (0.5 to 1.0 mM) produced a decrease of radioactivity in PE and PC with an increase in PI. Using prelabeled slices, a dose-dependent decrease in the 14C-radioactivity in PE and PC was observed, with a parallel increase in PI. This effect of Mp persisted even in the presence of a physiological activator of phospholipase A2, bradykinin (BK). No change in the net amount of phospholipids was observed at any of the Mp concentrations used. The results of this study show that Mp, at concentrations generally used to inhibit phospholipase A2, produced a transfer of arachidonic acid from PE and PC to PI, rather than a blockade in the release of AA from membrane phospholipids.
Assuntos
Ácidos Araquidônicos/metabolismo , Fosfatidilinositóis/metabolismo , Quinacrina/farmacologia , Animais , Ácido Araquidônico , Coenzima A/fisiologia , Relação Dose-Resposta a Droga , Técnicas In Vitro , Medula Renal/metabolismo , Fosfolipídeos/análise , Ratos , Ratos EndogâmicosRESUMO
The effect of bradykinin on the mobilization of arachidonic acid was analyzed separately by acylation and degradation. Acylating activity was determined by the incorporation of [14C]arachidonic acid into the phospholipids at different times. Different concentrations of bradykinin had no effect on the phospholipid acylating activities. The degradation of the phospholipids was performed on renal medullary slices prelabeled with [14C]arachidonic acid. Treatment with bradykinin produced an initial degradation of phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol, with a concomitant increase in lysophosphatidylcholine, lysophosphatidylethanolamine and lysophosphatidylinositol within 5 min of incubation. Phosphatidylcholine-, phosphatidylethanolamine- and phosphatidylinositol increased thereafter and reached the control values after 10 min of incubation. After 30 min, incubation of prelabeled slices with bradykinin produced a significant concentration-dependent increase in the phospholipid-labeling by reutilization of [14C]arachidonic acid. The effect of bradykinin on the phospholipid-labeling was blocked by preincubation with increasing concentrations of dBc AMP. Mepacrine also blocked the bradykinin stimulation in phosphatidylcholine and phosphatidylethanolamine, but had no effect on bradykinin-induced changes in the phosphatidylinositol arachidonic acid moiety.