血小板生成刺激因子对巨核血小板系统体外生成的影响

本 文应用血小板生成液体培养体系及纯化的血小板生成刺激因子(TSF)研究了 TSF对巨核细胞成熟及血小板生成的作用。TSF 在0.5—2U/ml 浓度范围内能够刺激巨核细胞DNA 合成,胞浆成熟,胞体直径增加以及血小板直径增加,但对巨核细胞与血小板计数没有影响。实验表明 TSF 作为一种血小板生成素,通过促进巨核细胞分化成熟,以增加血小板体积的方式,促进血板小生成。     

Decreased pulmonary vascular responsiveness in rats raised on an essential fatty acid deficient diet

Leukotriene C₄ is produced during hypoxic pulmonary vasoconstriction and leukotriene inhibitors preferentially inhibit the hypoxic pressor response in rats. If lipoxygenase products are important in hypoxic vasoconstriction, then an animal deficient in arachidonic acid should have a blunted hypoxic pressor response. We investigated if vascular responsiveness was decreased in vascular rings and isolated perfused lungs from rats raised on an essential fatty acid deficient diet (EFAD) compared to rats raised on a normal diet. Rats raised on the EFAD diet had decreased esterified plasma arachidonic acid and increased 5-, 8-, 11- eicosatrieonic acid compared to rats raised on the normal diet (control). Compared to the time matched responses in control isolated perfused lungs the pressor responses to angiotensin II and alveolar hypoxia were blunted in lungs from the arachidonate deficient rats. This decreased pulmonary vascular responsiveness was not affected by the addition of indomethacin or arachidonic acid to the lung perfusate. Similarly, the pulmonary artery rings from arichidonate deficient rats demonstrated decreased reactivity to norepinephrine compared to rings from control rats. In contrast, the tension increases to norepinephrine were greater in aortic rings from the arachidonate deficient rats compared to control. Stimulated lung tissue from the arachidonate deficient animals produced less slow reacting substance and platelet activating factor like material but the same amount of 6-keto-PGF_1α and TXB₂ compared to control lungs. Thus there is an associated between altered vascular responsiveness and impairment of stimulated production of slow reacting substance and platelet activating factor like materiali rat raised on an EFAD diet.     

Effect of the protein kinase inhibitors, staurosporine and K-252a, on PGI2 production by rat liver cells (the C-9 cell line)

Staurosporine and K-252a, known inhibitors of several protein kinases, stimulated PGI₂ production (measured as 6-keto-PGF_1α in rat liver cells (the C-9 cell line). Preincubation of the rat liver cells with staurosporine or K-252a enhanced the PGI₂ production stimulated by 12-O-tetradecanoylphorbol-13-acetate (TPA), platelet activating factor (PAF) and the Ca²⁺-ionophore a-23187, but not the PGI₂ synthesis stimulated by exogeneous arachidonic acid. These results suggest that phosphorylation of some proteins or certain amino acids on a protein can regulate arachidonic acid metabolism probably in the pathway leading to deesterification of phospholipids.     

The Effect of Lipopolysaccharide from <Emphasis Type="Italic">Proteus mirabilis</Emphasis> on the Level of the Stable End Metabolic Products of Nitric Oxide in Blood Platelets

Nitric oxide (^•NO) plays an important role in a number of physiologic processes. Evidence exists that ^•NO, which stimulates soluble guanylate cyclase and enhances cyclic guanosine monophosphate (cGMP) levels, may inhibit platelet activation. In contrast, during platelet activation induced by different agonists, synthesis of ^•NO in platelets occurs. In these studies, production of the stable end-products of ^•NO-nitrite and nitrate (NO_x) in human platelets, stimulated by different doses of lipopolysaccharide from Proteus mirabilis (LPS; endotoxin), has been evaluated. LPS is a weak platelet agonist that may activate various steps of platelet activation with the generation of reactive oxygen species. The mechanism of platelet activation induced by the endotoxin is not known. The aim of the present study was to measure the level of nitrite and NO_x in blood platelets treated with LPS and to examine the level of nitrotyrosine in platelet proteins caused by LPS. Our results show that LPS at a low concentration (6.8 ng/ml) caused a decrease (approximately 80%) in the NO_x level, whereas at higher concentrations (13.6 and 25 ng/ml) it induced an increase in the NO_x level (approximately 210% and 260%, respectively). Our results indicate that LPS, like other agonists (thrombin, platelet-activating factor), can stimulate ^•NO production in platelets. After incubating platelets with LPS, we also observed a distinct increase in platelet protein nitration (3-nitrotyrosine).     

Requirements for different Ca2+ pools in the activation of rabbit platelets: I. Release reaction and protein phosphorylation

We examined the role of Ca²⁺, both extracellular and intracellular in origin, in the release reaction and protein phosphorylation in rabbit platelets stimulated with platelet activating factor (acetylglyceryl ether phosphorylcholine), thrombin, or ionophore A23187. In the presence of extracellular Ca²⁺, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8), a putative antagonist of intracellular Ca²⁺ transport, blocked platelet activating factor-initiated serotonin release at a half-maximal inhibitor concentration of 40 μM, compared to 350 μM for thrombin-induced release and greater than 500 μM, for A23187-induced release. Platelet activating factor-induced phosphorylation of two platelet proteins of M_r=41 000 (P7P) and 20 000 (P9P) was inhibited by TMB-8, an effect which was additive to that caused by removing extracellular Ca²⁺. TMB-8 demonstrated only minor to non-existant inhibitory effects on phosphorylation in thrombin- or A23187-stimulated platelets. In contrast to P9P phosphorylation, phosphorylation of P7P caused by platelet activating factor was more dependent on a TMB-8 sensitive step than on the availability of extracellular Ca²⁺. Experiments with buffers containing fixed concentrations of free Ca²⁺ revealed that both processes (release and phosphorylation), when stimulated by platelet activating factor and thrombin, had the same threshold requirement (1–3 μM) for extracellular free Ca²⁺. These studies provide evidence that stimulation of rabbit platelets by platelet activating factor is more dependent on a TMB-8-sensitive intracellular Ca²⁺ source than is stimulation caused by thrombin. Furthermore, our data indicate that activation of different intracellular processes involved in platelet secretion (such as P7P and P9P phosphorylation) may require Ca²⁺ from different pools.     

Acetylsalicylic acid stimulates murine megakaryocyte precursor cells

Depression of platelet function with a single intraperitoneal injection of acetylsalicylic acid was found to produce significant increases in several thrombocytopoietic indicators despite no observed change in platelet counts. There was an increase in the number of megakaryocytic precursor cells (small acetylcholinesterase positive or "SAChE+" cells), platelet size, and 35S incorporation into platelets. The results are qualitatively comparable to data from previous experiments showing that treatment of mice with a thrombocytopoiesis-stimulating factor (TSF or thrombopoietin) and rabbit anti-mouse platelet serum will elevate thrombocytopoiesis. The results presented herein indicate that interruption of platelet function by aspirin results in the production of new platelets, presumably by the action of a feedback system controlling thrombocytopoiesis.     

Requirement for different Ca2+ pools in the activation of rabbit platelets: II. Phospholipase activity

The effects of extracellular Ca²⁺ concentration and the putative antagonist of intracellular Ca²⁺ movement, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8) on platelet phospholipase activity and thromboxane B₂ synthesis were examined in rabbit platelets stimulated by platelet activating factor, thrombin and ionophore A23187. TMB-8 markedly inhibited the platelet activating factor-induced decrease in [¹⁴C]arachidonate content in platelet phsophatidylacholine and phosphatidylinositol, while showing minimal effects on thrombin-induced phospholipase activation. A23187 stimulation of these processes was inhibited to an intermediated degree by TMB-8. In contrast, extracellular Ca²⁺ removal inhibited phospholipase activity to a similar degree with all three stimuli. Moreover, the threshold concentration of extracelullar Ca²⁺ for phospholiphase activation, as measured by thromboxane B₂ synthesis, was similar for platelet activating factor- and thrombin-stimulated platelets. The data provide evidence that, while platelet activating factor and thrombin may, to some extent, have similar requirements for extracellular Ca²⁺, they utilize a TMB-8 sensitive step to different degrees during activation of platelet phospholipase.     

Some interrelationships between glucose levels thromboxane production and prostacyclin production in normal and diabetic mice

We investigated the relationship between glucose levels and platelet thromboxane production or aortic prostacyclin production, using radioimmunoassay (RIA) to measure thromboxane B₂ and 6-keto-PGF_2α. We found a direct relationship (p<.05) between plasma glucose levels and thromboxane A₂ production by arachidonate stimulated platelets in platelet rich plasma of normal mice. However, when mice were deprived of food overnight, the glucose level fell but the TxB₂ production rose significantly. Moreover, mice with streptozotocin diabetes had significantly elevated glucose levels. but normal TxB₂ production, which also rose significantly after fasting. Thus in our laboratory both fasting and diabetes nullify or reverse the direct relationship between glucose levels and TxB₂ production seen in normal fed mice. This makes it diffisult to ascribe the correlation between glucose and TxB₂ levels in normal fed animals to cause and effect. RIA revealed an inverse correlation between glucose levels and 6-keto-PGF_1α production which was highly significant in aortas taken from fasted mice and stimulated for 10 minutes with 0.1mM arachidonate. This inverse correlation was present with either normal or diabetic aortas. Moreover, fasting increased the production of 6-keto-PGF_1α. However there was a significant elevation of 6-keto production by aortas of mice with diabetes of 5–6 weeks duration, compared to aortas of normal mice. Therefore either diabetes in these mice reversed a normal inhibitory effect of glucose on 6-keto production, or else the inverse correlation between glucose levels and 6-keto production does not represent a cause and effect relationship between the two variables.     

Platelet phosphoinositide turnover in streptozotocin-induced diabetes

Increased platelet aggregation and secretion in response to various agonists has been described in both diabetic humans and animals. Alterations in the platelet membrane fatty acid composition of phospholipids and changes in the prostacyclin and thromboxane formation could only partly explain the altered platelet function in diabetes. In the present study, we have examined the role of phosphoinositide turnover in the diabetic platelet function. We report alterations in 2-[³H] myo-inositol uptake, phosphoinositide turnover, inositol phosphate and diacylglycerol (DAG) formation, phosphoinositide mass, and phospholipase C activity in platelets obtained from streptozotocin (STZ)-induced diabetic rats. There was a significant increase in the 2-[³H) myo-inositol uptake in washed platelets from diabetic rats. Basal incorporation of 2-[³H] myo-inositol into phosphatidylinositol 4,5-bisphosphate (PIP₂), phosphatidylinositol 4-phosphate (PIP) or phosphatidylinositol (PI) in platelets obtained from diabetic rats was, however, not affected. Thrombin stimulation of platelets from diabetic rats induced an increase in the hydrolysis of [³²P]PIP₂ but indicated no change in the hydrolysis of [³²P]PIP and [³²P]PI as compared to their basal levels. Thrombin-induced formation of [³H]inositol phosphates was significantly increased in both diabetic as well as in control platelets as compared to their basal levels. This formation of [³H]inositol phosphates in diabetic platelets was greater than controls at all time intervals studied. Similarly, there was an increase in the release of DAG after thrombin stimulation in the diabetic platelets. Based on these results, we conclude that there is an increase in the transport of myoinositol across the diabetic platelet membrane and this feature, along with alterations in the hydrolysis of PIP₂, inositol phosphates and DAG in the diabetic platelets, may play a role in increased phosphoinositide turnover which could explain the altered platelet function in STZ-induced diabetes.     

Two-chain structure of T-suppressor factor: Antigen-specific T-suppressor factor occurs as a single molecule and as separate antigen-binding and I-J+ parts,both of which are required for biological activity

Antigen-specific T-suppressor factor (TsF), which acts at the expression stage of the contact sensitivity reaction, was produced by culturing the lymphoid cells of mice injected with picryl-sulphonic (trinitrobenzenesulphonic) acid and then painted with picryl chloride. Supernatant activity was found around 50–60 and 90 kDa on Sephadex gel filtration. The activity at 50–60 kDa was due to two separate (or readily separable) molecules, one antigen binding and the other bearing I-J determinants as shown by affinity chromatography on insolubilized antigen and anti-I-J. These two separate molecules were inactive alone but complemented each other and may be designated as the variable chain of TsF (TsF_v) and the I-J⁺ chain. The use of gel filtration and sequential absorption of individual pools on anti-I-J antibody followed by antigen, together with a complementation assay, also showed a TSF_v chain at 30–40 kDa and an I-J⁺ chain at 20–30 kDa. The higher-molecular-weight activity around 90 kDa was due to a single molecule which was both antigen binding and I-J⁺. This molecule dissociated on treatment with the reducing agent dithioerythritol followed by alkylation into two separate chains, one antigen binding and the other I-J⁺, both of which were required for activity. There was a requirement for genetic matching between the antigen-binding chain (TsF_v) and the I-J⁺ chain for biological activity. These data support a two-chain model of TsF in which TsF_v and the I-J⁺ chain occur as a single disulphide-bonded molecule around 90 kDa, or as two separate (or readily separable) chains of lower molecular weight which were inactive alone but complemented each other.     

Activated Platelets from Diabetic Rats Cause Endothelial Dysfunction by Decreasing Akt/Endothelial NO Synthase Signaling Pathway

Diabetes is associated with endothelial dysfunction and platelet activation, both of which may contribute to increased cardiovascular risk. The purpose of this study was to characterize circulating platelets in diabetes and clarify their effects on endothelial function. Male Wistar rats were injected with streptozotocin (STZ) to induce diabetes. Each experiment was performed by incubating carotid arterial rings with platelets (1.65×10⁷ cells/mL; 30 min) isolated from STZ or control rats. Thereafter, the vascular function was characterized in isolated carotid arterial rings in organ bath chambers, and each expression and activation of enzymes involved in nitric oxide and oxidative stress levels were analyzed. Endothelium-dependent relaxation induced by acetylcholine was significantly attenuated in carotid arteries treated with platelets isolated from STZ rats. Similarly, treatment with platelets isolated from STZ rats significantly reduced ACh-induced Akt/endothelial NO synthase signaling/NO production and enhanced TXB₂ (metabolite of TXA₂), while CD61 (platelet marker) and CD62P (activated platelet marker) were increased in carotid arteries treated with platelets isolated from STZ rats. Furthermore, the platelets isolated from STZ rats decreased total eNOS protein and eNOS dimerization, and increased oxidative stress. These data provide direct evidence that circulating platelets isolated from diabetic rats cause dysfunction of the endothelium by decreasing NO production (via Akt/endothelial NO synthase signaling pathway) and increasing TXA₂. Moreover, activated platelets disrupt the carotid artery by increasing oxidative stress.     

Coronary vasoconstrictor extracted from blood plasma stimulates platelet lipoxidase activity

The effects of a coronary vasoconstrictor, obtained from human blood plasma, on aggregation and arachidonate metabolism by human platelets was determined. At low concentrations, the vasoactive factor stimulated formation of prostaglandins, thromboxane B₂, and 12-L-hydroxyeicosatetraenoic acid in both intact platelets and in platelet microsomal enzyme preparations. As factor concentration was increased, thromboxane B₂ formation decreased, but production of the other products continued to rise. Low concentrations of factor initiated platelet aggregation, whereas high concentrations prevented arachidonate-induced aggregation. Low levels of factor could induce aggregation via stimulation of thromboxane A₂ production. Increases in formation of 12-hydroperoxyeicosatetraenoic acid at high factor concentrations could inhibit formation of thromboxane A₂ and thus prevent aggregation.     

Reduced PGE1 stimulated 3H-cAMP accumulation in platelets from schizophrenics.

Accumulation of ³H-cAMP was measured in platelet rich plasma (PRP) pulse labelled with ³H-adenine and incubated with prostaglandin E₁ (PGE₁). Less PGE₁ stimulated ³H-cAMP accumulation was seen in PRP from schizophrenics than in PRP from controls at all PGE₁ concentrations studied. Levels of ³H-cAMP in PRP incubated without PGE₁ and platelet incorporation of total tritium did not differ between these groups. The possibility that reduced functional prostaglandin activity may be an etiopathologic factor in schizophrenia, and the compatibility of this postulate with diverse theoretical and empirical observations related to schizophrenia are discussed.     

2-(6-carboxyhexyl)cyclopentanone hexylhydrazone: A potent inhibitor of the blood platelet cyclo-oxygenase

Previous studies have demonstrated that 13-azaprostanoic acid (13-APA) is a potent and specific antagonist of thromboxane A₂/prostaglandin H₂ (TXA₂/PGH₂) at the platelet receptor level. In the present study we evaluated the effects of a new azaprostanoid, 2-(6-carboxyhexyl) cyclopentanone hexylhydrazone (CPH), on human platelet function. This hydrazone was found to completely inhibit arachidonic acid (AA)-induced platelet aggregation at 1 uM CPH. On the other hand, CPH was not an effective inhibitor of PGH₂-induced aggregation. Furthermore, 100 uM CPH was completely ineffective in blocking platelet aggregation stimulated by adenosine diphosphate (ADP) or the stable prostaglandin endoperoxide analog U46619 (which presumably acts at the TXA₂/PGH₂ receptor). Measurement of platelet thromboxane B₂ (TXB₂) production demonstrated that the primary site-of-action of CPH is at the cyclo-oxygenase level. Thus, CPH inhibited TXB₂ formation from AA in a dose-dependent manner (0.1 uM–100 uM CPH)². In contrast, CPH blocked TXB₂ production from PGH₂ only at the highest CPH concentration tested, i.e., 100 uM. These results indicate that relative to 13-APA, addition of a second nitrogen at C₁₄ and a double bond between the 12- and 13- positions results in a loss of receptor activity but produces a high affinity for the platelet cyclo-oxygenase.     

Actions of vanadate of arachidonic acid metabolism by cells in culture

Sodium vanadate (11 μM) amplified the PGI₂ production of rat liver cells (the C-9 cell line) incubated with thrombin, platelet activating factor, lysine-vasopressin, the Ca²⁺-ionophore A-23187, interleukin-1ß, 12-tetradecanoylphorbol-13-acetate, teleocidin, epidermal growth factor, palytoxin, thapsigargin and colchicine but not that stimulated by exogenous arachidonic acid. Sodium vanadate (2.2 μM) also amplified PGF_2α production of dog kidney cells (the MDCK cell line) incubated with norepinephrine and, at 0.4 μM, PGI₂ production of bovine aorta smooth muscle cells stimulated by serotonin. Sodium vanadate (55 μM) did not affect production of PGE₂ and PGF_2α in rat basophil leukemia cells (the RBL-1 cell line) stimulated by the Ca²⁺-ionophore A-23187, but did inhibit synthesis of peptide-containing leukotrienes and 12-hydroxyeicosatetraenoic acid. When used with cultured cells at micromolar concentrations, vanadate is known to inhibit protein tyrosine-phosphate phosphatases. These results suggest that in some cells deesterification of lipids is positively regulated, at least in part, by phosphorylation of tyrosine whereas in leukocytes, lipoxygenase activities are negatively regulated, at least in part, by phosphorylation of tyrosine.     

Low ethanol intake prevents salt-induced hypertension in WKY rats

Low alcohol intake in humans lowers the risk of coronary heart disease and may lower blood pressure. In hypertension, insulin resistance with altered glucose metabolism leads to increased formation of aldehydes. We have shown that chronic low alcohol intake decreased systolic blood pressure (SBP) and tissue aldehyde conjugates in spontaneously hypertensive rats and demonstrated a strong link between elevated tissue aldehyde conjugates and hypertension in salt-induced hypertensive Wistar-Kyoto (WKY) rats. This study investigated the antihypertensive effect of chronic low alcohol consumption in high salt-treated WKY rats and its effect on tissue aldehyde conjugates, platelet cytosolic free calcium ([Ca^{2 +}] _i )_,and renal vascular changes. Animals, aged 7 weeks, were divided into three groups of six animals each. The control group was given normal salt diet (0.7% NaCl) and regular drinking water; the high salt group was given a high salt diet (8% NaCl) and regular drinking water; the high salt + ethanol group was given a high salt diet and 0.25% ethanol in drinking water. After 10 weeks, SBP, platelet [Ca^{2 +}] _i , and tissue aldehyde conjugates were significantly higher in rats in the high salt group as compared with controls. Animals on high salt diets also showed smooth muscle cell hyperplasia in the small arteries and arterioles of the kidney. Ethanol supplementation prevented the increase in SBP and platelet [Ca^{2 +}] _i and aldehyde conjugates in liver and aorta. Kidney aldehyde conjugates and renal vascular changes were attenuated. These results suggest that chronic low ethanol intake prevents salt-induced hypertension and attenuates renal vascular changes in WKY rats by preventing an increase in tissue aldehyde conjugates and cytosolic [Ca^{2 +}] _i .     

Hydrogen peroxide and peroxynitrite enhance Ca2+ mobilization and aggregation in platelets from type 2 diabetic patients

Cytosolic Ca²⁺ mobilization, especially Ca²⁺ entry, is enhanced in platelets from type 2 diabetic individuals, which might result in platelet hyperaggregability. In the present study, we report an increased oxidant production in resting and stimulated platelets from diabetic donors. Pretreatment of platelets with catalase or trolox, an analog of vitamin E, reversed the enhanced Ca²⁺ entry, evoked by thapsigargin plus ionomycin or thrombin, observed in platelets from diabetic subjects, so that in the presence of these scavengers Ca²⁺ entry was similar in platelets from healthy and diabetic subjects. In contrast, mannitol was without effect on Ca²⁺ mobilization. Catalase and trolox reduced thrombin-induced aggregation in platelets from type 2 diabetic subjects, while mannitol did not modify thrombin-induced platelet hyperaggregability. We conclude that H₂O₂ and ONOO⁻ are likely involved in the enhanced Ca²⁺ mobilization observed in platelets from type 2 diabetic patients, which might lead to platelet hyperactivity and hyperaggregability.     

Sources of Variability in Platelet Accumulation on Type 1 Fibrillar Collagen in Microfluidic Flow Assays

Microfluidic flow assays (MFA) that measure shear dependent platelet function have potential clinical applications in the diagnosis and treatment of bleeding and thrombotic disorders. As a step towards clinical application, the objective of this study was to measure how phenotypic and genetic factors, as well as experimental conditions, affect the variability of platelet accumulation on type 1 collagen within a MFA. Whole blood was perfused over type 1 fibrillar collagen at wall shear rates of 150, 300, 750 and 1500 s⁻¹ through four independent channels with a height of 50 µm and a width of 500 µm. The accumulation of platelets was characterized by the lag time to 1% platelet surface coverage (Lag_T), the rate of platelet accumulation (V_PLT), and platelet surface coverage (SC). A cohort of normal donors was tested and the results were correlated to plasma von Willebrand factor (VWF) levels, platelet count, hematocrit, sex, and collagen receptors genotypes. VWF levels were the strongest determinant of platelet accumulation. VWF levels were positively correlated to V_PLT and SC at all wall shear rates. A longer Lag_T for platelet accumulation at arterial shear rates compared to venous shear rates was attributed to the time required for plasma proteins to adsorb to collagen. There was no association between platelet accumulation and hematocrit or platelet count. Individuals with the AG genotype of the GP6 gene had lower platelet accumulation than individuals with the AA genotype at 150 s⁻¹ and 300 s⁻¹. Recalcified blood collected into sodium citrate and corn trypsin inhibitor (CTI) resulted in diminished platelet accumulation compared to CTI alone, suggesting that citrate irreversibly diminishes platelet function. This study the largest association study of MFA in healthy donors (n = 104) and will likely set up the basis for the determination of the normal range of platelet responses in this type of assay.     

Role of lipoxygenase metabolism in platelet function: Effect of aspirin and salicylate

Aspirin inhibits thromboxane A₂ (TxA₂) production whereas its salicylate moiety inhibits 12-hydroxy-eiosatetraenoic acid (12-HETE) production in the platelet. The significance of the latter effect on platelet function is unclear. We examined the effects of aspirin and salicylate on (i) platelet/ collagen adhesion using ³H-adenine-labelled human platelets and collagen- coated discs, (ii) platelet aggregation induced by thrombin, collagen, ADP and arachidonic acid, and (iii) platelet TxA₂ and 12-HETE synthesis as measured by radioimmunoassay and high pressure liquid chromatography respectively. Aspirin (50 μM) decreased platelet aggregation and increased platelet adhesion. The decrease in aggregation was associated with inhibition of TxA₂ production and the increase in adhesion was associated with enhanced 12-HETE production. Salicylate had the opposite effects. Platelet aggregation was increased and platelet adhesion decreased. The increased aggregation was associated with enhanced TxA₂ production and the decrease in aggregation was associated with inhibition of 12-HETE production. These observations suggest that 12-HETE facilitates platelet adhesion which can be altered by salicylate treatment.     

Effect of PGE1 on gluconeogenesis and glycerol esterification in perfused liver of fasted rats

Using perfused livers of rats fasted for 48 hours, glucose production and incorporation of 2-¹⁴C pyruvate (trace dose) into perfusate glucose were studied. Both were found to be inhibited by PGE₁ (infused at a concentration of 0.5 μg/min) by about 60 %. The incorporation of 1-¹⁴C glycerol into perfusate glucose and into glycerol-glyceride part of the liver glycerides were also studied, using the same test conditions. The former incorporation was significantly inhibited (56%) and the latter strongly stimulated (360 %) by PGE₁. PGE₁ had no effect on glucose production in a perfusate overloaded with sodium pyruvate, nor on pyruvate carboxylase and phospho-enolpyruvate carboxykinase activity. This was in contrast with the results obtained in perfusions with a trace dose of 2-¹⁴C pyruvate. The results showed that PGE₁, at the physiological concentration used, stimulated the incorporation of 1-¹⁴C glycerol into glycerol-glyceride part of liver glycerides and, when there was no overload of pyruvate present in the perfusion medium, inhibited gluconeogenesis at some point, possibly, but perhaps not exclusively, between the glycerol and glucose steps.