Opposite effects of adrenalectomy on eicosanoid release in rat peritoneal macrophages and spleen

The effect of adrenalectomy on the formation of cyclo-oxygenase and lipoxygenase products by activated peritoneal rat macrophages was determined and compared with that of the spleen. After isolation, the cells and tissues were incubated with [1-14C] arachidonic acid and the Ca-ionophore A23187 and the metabolites isolated by HPLC chromatography. The main components formed in the macrophages of the controls are 6-keto-PGF1 alpha, TxB2 and 12-HETE. One peak represents 5, 12 di HETE. Smaller amounts of PGF2 alpha, PGE2, PGD2, LTB4 and 15-HETE are also present. After adrenalectomy, a considerable increase occurs in the amounts of LTB4, 15-HETE and 12-HETE. The increase in the PG is smaller. The compounds formed from endogenous arachidonic acid are also determined. In the cells of the controls, the formation of LTB4 is considerably increased after adrenalectomy. In the spleen, PGD2 and 12-HETE are decreased after adrenalectomy. The effect of the macrophages is most probably related to a diminished amount or inactivation of lipocortin, a glucocorticosteroid induced peptide with PlA2 inhibitory activity in adrenalectomized animals. In the decrease in formation in the spleen, the absence of the permissive effect of glucocorticosteroids on the hormone-induced lipolysis may play a role.     

Ethanol inhibits zymosan-stimulated eicosanoid production in mouse peritoneal macrophages

Resident peritoneal macrophages synthesized and released eicosanoids when challenged by zymosan, a phagocytosable particle. Incubation of these cells with ethanol resulted in dose-dependent inhibition of arachidonic acid release and eicosanoid generation in response to zymosan. Ethanol affected the extent but not the ratio of eicosanoids released. When assayed in a cell-free system, endogenous phospholipase A₂ activity was neither affected by the presence of ethanol in the incubation medium nor by preincubation of the cells with ethanol. Ethanol also inhibited arachidonic acid release in response to phorbol myristate acetate, a compound that, like zymosan, triggered a pertussis-toxin-sensitive response. When cells that had been previously treated with pertussis toxin were used, no further inhibitory effect of ethanol was seen in response to both zymosan and phorbol myristate acetate. On the other hand, ethanol had no effect on arachidonic acid release stimulated by ionophore A23187 or lipopolysaccharide, two compounds that triggered a pertussis-toxin-insensitive response. Moreover, ethanol was able to nearly abolish arachidonic acid release in response to fluoroaluminate, a direct activator of G-proteins. Altogether, the results of this study suggest that ethanol inhibits zymosan-stimulated eicosanoid production by interacting with a G-protein — or a G-protein-mediated process — that is critically involved in arachidonic acid mobilization.     

Effect of adrenalectomy on rat peritoneal macrophage response

Glucocorticoid hormones are important for vital functions and act to modulate inflammatory and immune responses. In contrast to other hormonal systems no endogenous mediators have been identified that can directly counter-regulate their potent anti-inflammatory and immunosuppressive properties. Glucocorticoids are known to interfere with the ability of the macrophage not only to induce and amplify an immune response but also to inhibit macrophage inflammatory effector functions. Although the actual immunocompetence of animals undergoing endocrine gland ectomy has never been directly studied, there is no doubt that adrenal hormones are deeply involved in the development and maintenance of the immunitory functions and this may in turn influence the inflammatory reaction. To study the effect of endogenous glucocorticoids on the functions of rat peritoneal macrophages and induction of humoral immune response we observed some of the rat peritoneal macrophage effector functions, provided that endogenous glucocorticoids are depicted by adrenalectomy. The mean phagocytic index (PI) of control macrophage (Mphi) is increased from 23,825 +/- 427 to 31,895 +/- 83 after adrenalectomy (P < or = 0.001). Intracellular killing capacity in control cell is 82% which is found to be 73% in case of adrenalectomised cell (p < 0.05). The amount of nitric oxide released from control Mphi 20.25 +/- 1 microM following adrenalectomy shows the amount of nitric oxide release was 18.25 microM (p < or = 0.01 ). The percentage of DNA fragmentation in control Mphi was 68.82 +/- 4 which was reduced to 56.76 +/- 1 after adrenalectomy (p < or = 0.01). In sheep red blood cell (SRBC) immunised and adrenalectomised animal, agglutination titre was obtained at lowest antibody concentration (1 : 128) whereas serum from SRBC immunised normal rats showed early agglutination (1: 32). Endogenous glucocorticoid depleted rats show enhanced phagocytic capacity, antibody raising capacity as well as on the other hand adrenal hormone insufficiency reduces the intracellular killing capacity, nitric oxide (NO) release, improper cell maturation and heightens the probability of infection. These observations demonstrate a counter-regulatory system via glucocorticoid that functions to control inflammatory and immune responses.     

Dual effects of staurosporine on arachidonic acid metabolism in rat peritoneal macrophages

Staurosporine is a microbial anti-fungal alkaloid having a most potent inhibitory activity on protein kinase C and is recently found as a non-12-O-tetradecanoylphorbol-13-acetate (non-TPA)-type tumor promoter of mouse skin, although tumor promotion induced by a TPA-type tumor promoter teleocidin is suppressed by staurosporine. When rat peritoneal macrophages were incubated in the medium containing various concentrations of staurosporine, prostaglandin E2 production and release of radioactivity from [3H]arachidonic acid-labeled macrophages were stimulated at concentrations of 1 and 10 ng/ml. But higher concentrations of staurosporine such as 100 and 1000 ng/ml showed no stimulative effect on prostaglandin E2 production although cytoplasmic free calcium levels were increased in a dose-dependent manner. Staurosporine-induced stimulation of prostaglandin E2 production was inhibited by treatment with cycloheximide, suggesting that a certain protein synthesis is prerequisite for the stimulation of arahcidonic acid metabolism. At higher concentrations (100 and 1000 ng/ml), staurosporine inhibited TPA-type tumor promoter (TPA, teleocidin and aplysiatoxin)-induced stimulation of arachidonic acid metabolism probably due to the inhibition of protein kinases. Tumor promotion activity and anti-tumor promotion activity of staurosporine might be explained by the fact that the lower concentrations of staurosporine stimulate arachidonic acid metabolism and the higher concentrations of staurosporine inhibit the tumor promoter-induced arachidonic acid metabolism, respectively.     

Different effects of concanavalin A and its succinylated derivative on superoxide release in peritoneal macrophages

The effects of tetravalent concanavalin A and its succinylated derivative on the intracellular production of superoxide anion (O₂^?) and its release into cell exterior of peritoneal macrophages were observed. Both tetravalent concanavalin A and its succinylated derivative induced marked enhancement of intracellular reduction of nitroblue tetrazolium, which could be inhibited by α-methyl-D-glucoside. The extent of activation of nitroblue tetrazolium reduction induced by both types of the lectin paralleled the activation ratio of oxygen consumption.There was littele difference in the extent of intracellular O₂^? production induced by two types of the lectin. Nitroblue tetrazolium reduction was not affected significantly by pretreatment with colchicine, rotenone or malonate, inhibitors of the cytoskeletal system and of the electron transport system. In contrast, tetravalent concanavalin A induced a higher rate of superoxide release than did succinylated divalent concanavalin A, which lacks the cross-linking activity of surface glycoproteins.These results indicate that superoxide production following oxygen consumption and superoxide release into cell exterior are controlled independently by a separate membrane mechanism and that superoxide production system is not essentially dependent on the involvement of the cytoskeletal system.     

Different effects of concanavalin A and its succinylated derivative on superoxide release in peritoneal macrophages.

The effects of tetravalent conconavalin A and its succinylated derivative on the intracellular production of superoxide anion (O-2) and its release into cell exterior of peritoneal macrophages were observed. Both tetravalent concanavalin A and its succinylated derivative induced marked enhancement of intracellular reduction of nitroblue tetrazolium, which could be inhibited by alpha-methyl-D-glucoside. The extent of activation of nitroblue tetrazolium reduction induced by both types of the lectin paralleled the activation ratio of oxygen consumption. There was little difference in the extent of intracellular O-2 production induced by two types of the lectin. Nitroblue tetrazolium reduction was not affected significantly by pretreatment with colchicine, rotenone or malonate, inhibitors of the cytoskeletal system and of the electron transport system. In contrast, tetravalent concanavalin A induced a higher rate of superoxide release than did succinylated divalent concanavalin A, which lacks the cross-linking activity of surface glycoproteins. These results indicate that superoxide production following oxygen consumption and superoxide release into cell exterior are controlled independently by a separate membrane mechanism and that superoxide production system is not essentially dependent on the involvement of the cytoskeletal system.     

Effect of aflatoxins on rat peritoneal macrophages

Phagocytosis, intracellular killing of Candida albicans, and superoxide production by rat peritoneal macrophages exposed to aflatoxins B1, B2, G1, G2, B2a, and M1 at several times and concentrations were analyzed to evaluate the intensity of a depressive effect for each mycotoxin. All aflatoxins used at very low concentrations had a depressive effect on the functions of macrophages. The biggest impairment of phagocytosis, intracellular killing, and spontaneous superoxide production was observed in macrophages exposed to aflatoxins B1 and M1.     

Hydrogen peroxide release by rat peritoneal macrophages in the presence and absence of tumor cells

The ability of mineral oil-elicited rat peritoneal macrophages to release hydrogen peroxide (H₂O₂) to the extracellular medium was measured in the presence and absence of rat lymphoma cells grown in tissue culture, and in the presence of phorbol myristate acetate (PMA). Horseradish peroxidase (HRP)-catalyzed oxidation of scopoletin or phenol red was used to measure H₂O₂ release during incubation of cells in monolayer culture for periods up to 24 h. Macrophages appeared to release H₂O₂ with or without PMA, although PMA greatly increased the amount of H₂O₂ released in short (1 to 4 h) incubations. Tumor cells did not replace PMA as a triggering agent for H₂O₂ release. Instead, tumor cells inhibited H₂O₂ release. The probable basis for inhibition was competition between macrophages and tumor cells for the supply of oxygen (O₂). Tumor cells did not inhibit H₂O₂ release when the O₂ concentration was held constant. The rates at which macrophages took up O₂ and released H₂O₂ were proportional to the O₂ concentration, as measured with the O₂ electrode. Rates of H₂O₂ release could be calculated from the difference in the rate constants for O₂ uptake measured in the presence of two different extracellular H₂O₂-consuming systems (HRP-scopoletin vs catalase). PMA-stimulated uptake of O₂ and release of H₂O₂ were highest in a small subpopulation of macrophages, obtained at the lowest-density position on gradients of bovine serum albumin. These cells also released H₂O₂ in the absence of PMA. Tumor cells had no effect on the rate constants for O₂ uptake and H₂O₂ release by the unfractionated macrophages or the macrophage subpopulations.     

Pertussis toxin and H-7 distinguish mechanisms involved in eicosanoid release from lipopolysaccharide-primed macrophages. Eicosanoid release from lipopolysaccharide-primed macrophages

Release of eicosanoids is an important response of macrophages to inflammation and bacterial infection. At low concentrations, bacterial lipopolysaccharide (1-2 micrograms/ml) fails to stimulate eicosanoid release in resident peritoneal macrophages but primes the macrophages for a greatly enhanced release of eicosanoids on stimulation with the calcium ionophore A23187 (0.1 microM) or with phorbol 12-myristate 13-acetate (50 nM), an activator of protein kinase C. Incubation of macrophages with Bordetella pertussis toxin, prior to priming with lipopolysaccharide, inhibited the release of both cyclooxygenase and lipoxygenase products upon A23187 stimulation. Pertussis toxin treatment of macrophages had no effect on eicosanoid release when the stimulus was phorbol 12-myristate 13-acetate. The presence of 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), an effective inhibitor of protein kinase C, during lipopolysaccharide priming and subsequent stimulation significantly inhibited eicosanoid release when phorbol 12-myristate 13-acetate was the stimulus, but did not affect eicosanoid release stimulated by A23187. Based on these results, at least two mechanisms, distinguished by apparent differences in sensitivity to pertussis-toxin-sensitive, guanine-nucleotide-binding proteins and protein kinase C, are involved in eicosanoid secretion by lipopolysaccharide-activated macrophages in response to A23187 and phorbol 12-myristate 13-acetate.     

Enhancement of eicosanoid synthesis in mouse peritoneal macrophages by the organic mercury compound thimerosal

Availability of the common precursor arachidonic acid represents the fundamental prerequisite of the cellular eicosanoid synthesis. The amount of free arachidonic acid is regulated not only by phospholipases, which liberate this polyunsaturated fatty acid from lipid pools, but also by the reacylating enzyme acylCoA:lysophosphatide acyltransferase. We have previously shown (Goppelt-Strübe, G., C.-F. Körner, G. Hausmann, D. Gemsa, and K. Resch. Control of Prostanoid Synthesis: Role of Reincorporation of Released Precursor Fatty Acids. Prostaglandins : 373. 1986.) that the organic mercury compound thimerosal in murine peritoneal macrophages inhibits arachidonic acid reincorporation into cellular lipids, thereby leading to an enhanced prostanoid synthesis. In this report we show that the production of leukotriene C₄ was also increased after the addition of thimerosal to mouse peritoneal macrophages in a time and dose dependent manner. Concomitantly, thimerosal led to a significant rise of the intracellular calcium concentration as measured by fura-2 fluorescence. Simultaneous addition of thimerosal and indomethacin or exogeneous arachidonic acid to the cells resulted in a synergistic enhancement of leukotriene C₄ synthesis. On the other hand, another sulfhydryl group blocking agent, ethacrynic acid, was found to be ineffective in increasing leukotriene C₄ levels even in combination with exogeneous arachidonic acid. Thimerosal therefore provides a helpful tool in studying the basic regulatory mechanisms of the cellular leukotriene synthesis.     

The effects of peritoneal macrophages on monolayered luteal cell progestin secretion in the rat

Functionally active or regressing luteal cells were obtained from pseudo-pregnant (psp) rats between days 5-8 of psp or on day 15 of psp, respectively. They were monolayer-cultured (10(6)/dish) in the presence of 0.2 micrograms/ml LH 2.0 micrograms/ml PRL and 10 micrograms/ml pregnenolone for 4 days with or without macrophages, although functionally active luteal cells secreted progesterone dominantly during day 1 of culture (Day 1), the amounts of progesterone and 20 alpha-OH-P secreted were inverted on Day 2, and the dominance of 20 alpha-OH-P continued from Day 2 to Day 4. In the functionally regressing luteal cell culture, more 20 alpha-OH-P than progesterone was secreted throughout the culture period. The addition of peritoneal macrophages (2.5 X 10(6] to the active luteal cell monolayer lengthened the dominance of progesterone secretion for an additional day and the inversion occurred on Day 3. The progestin ratio (progesterone/20 alpha-OH-P) on Day 2 was maintained significantly higher. The daily addition of macrophages maintained the progesterone dominance throughout the culture period. On the other hand, macrophages had no effect on luteal cells already functionally regressing. These results indicate that macrophages are effective in maintaining the progesterone secreting activity of luteal cells in vitro.     

Enhancement of eicosanoid synthesis in mouse peritoneal macrophages by the organic mercury compound thimerosal

Availability of the common precursor arachidonic acid represents the fundamental prerequisite of the cellular eicosanoid synthesis. The amount of free arachidonic acid is regulated not only by phospholipases, which liberate this polyunsaturated fatty acid from lipid pools, but also by the reacylating enzyme acylCoA:lysophosphatide acyltransferase. We have previously shown (Goppelt-Strübe, G., C.-F. K?rner, G. Hausmann, D. Gemsa, and K. Resch. Control of Prostanoid Synthesis: Role of Reincorporation of Released Precursor Fatty Acids. Prostaglandins 32:373. 1986.) that the organic mercury compound thimerosal in murine peritoneal macrophages inhibits arachidonic acid reincorporation into cellular lipids, thereby leading to an enhanced prostanoid synthesis. In this report we show that the production of leukotriene C4 was also increased after the addition of thimerosal to mouse peritoneal macrophages in a time and dose dependent manner. Concomitantly, thimerosal led to a significant rise of the intracellular calcium concentration as measured by fura-2 fluorescence. Simultaneous addition of thimerosal and indomethacin or exogeneous arachidonic acid to the cells resulted in a synergistic enhancement of leukotriene C4 synthesis. On the other hand, another sulfhydryl group blocking agent, ethacrynic acid, was found to be ineffective in increasing leukotriene C4 levels even in combination with exogeneous arachidonic acid. Thimerosal therefore provides a helpful tool in studying the basic regulatory mechanisms of the cellular leukotriene synthesis.     

Effect of aflatoxins on rat peritoneal macrophages.

Phagocytosis, intracellular killing of Candida albicans, and superoxide production by rat peritoneal macrophages exposed to aflatoxins B1, B2, G1, G2, B2a, and M1 at several times and concentrations were analyzed to evaluate the intensity of a depressive effect for each mycotoxin. All aflatoxins used at very low concentrations had a depressive effect on the functions of macrophages. The biggest impairment of phagocytosis, intracellular killing, and spontaneous superoxide production was observed in macrophages exposed to aflatoxins B1 and M1.     

Carbohydrate metabolism in the rat peritoneal macrophages

Rat peritoneal macrophages derive energy differently from other tissues. Resting rat peritoneal macrophages have been taken for the present investigation. Lactate produced by extracellular glycolysis in the peritoneal lavage fluid, is readily converted into pyruvate by resting peritoneal macrophages and is oxidised in mitochondria. Glycolytic enzymes other than phosphoglucoisomerase and lactate dehydrogenase could not be substantially demonstrated. Glucose-6-phosphate dehydrogenase was detected. The presence of glucose-6-phosphate dehydrogenase along with phosphoglucoisomerase indicates the operation of the hexose monophosphate shunt as a pathway supplementary to glycolysis. Resting rat peritoneal macrophages thus appear to utilize extracellular lactate as their main energy source instead of glucose, bypass glycolysis and have active hexose monophosphate shunt.     

The effects of platelet-activating factor on flow rate and eicosanoid release in the isolated perfused rat gastric vascular bed

In the isolated rat stomach perfused via the vasculature in situ under constant pressure bolus injections of platelet-activating factor (PAF, 3, 16, or 50 ng) induced dose-dependent, long-lasting reductions of flow rates and simultaneously significant increases in the release of cysteinyl-leukotrienes (cys-LT), thromboxane (TX) B₂ and 6-keto-prostaglandin (PG) F_1α. Reversed phase high pressure liquid chromatography demonstrated the release of a mixture of comparable amounts of LTC₄, LTD₄ and LTE₄ by PAF. Inhibition of cys-LT sythesis by the lipoxygenase inhibitors nordihydroguaiaretic acid (NDGA) and L-651, 896 did not significantly affect PAF-induced flow reduction indicating that endogenous cys-LT are of minor importance for the PAF effect on gastric vascular flow. This conclusion is supported by the fact that the cys-LT receptor antagonist FPL 55712 in a concentration (1 × 10⁻⁶ M) that completely antagonized gastric flow reduction by exogenous LTC₄ (1 × 10⁻⁷ M) had no effect on the PAF-induced reduction of flow. The cyclooxygenase inhibitor indomethacin aggravated the PAF-induced flow reduction suggesting that the endogenous vasodilator PGI₂ might act as a functional PAF antagonist in the rat gastric vascular bed. In contrast to FPL 55712 the PAF antagonist BN 52021 significantly and concentration-dependently antagonized the PAF effect on gastric vascular flow. The results demonstrate that PAF and LTC₄ induce flow reductions in the rat gastric vascular bed by activating different receptors and that endogenous eicosanoids released by PAF do not contribute significantly to the PAF effect on gastric vascular flow.     

Downstream effects of splanchnic ischemia-reperfusion injury on renal function and eicosanoid release

Rothenbach, Patricia, Richard H. Turnage, Jose Iglesias,Angela Riva, Lori Bartula, and Stuart I. Myers. Downstream effectsof splanchnic ischemia-reperfusion injury on renal function andeicosanoid release. J. Appl. Physiol.82(2): 530-536, 1997.This study examines the hypothesis thatintestinal ischemia-reperfusion (I/R) injury contributes to renaldysfunction by altered renal eicosanoid release. AnesthetizedSprague-Dawley rats underwent 60 min of sham or superior mesentericartery (SMA) occlusion with 60 min of reperfusion. The I/R groupsreceived either allopurinol, pentoxifylline, 1-benzylimidazole, orcarrier before SMA occlusion. In vivo renal artery blood flow wasmeasured by Transonic flow probes, the kidneys were then perfused invitro for 30 min, and the effluent was analyzed for eicosanoid releaseand renal function. Intestinal I/R caused a twofold increase in theratio of renal release of thromboxaneB₂ to prostaglandinE₂ and to 6-ketoprostaglandin F₁ compared with the shamlevel, with a corresponding 25% decrease in renal sodium and inulinclearance and renal blood flow. Pentoxifylline or allopurinolpretreatment restored renal eicosanoid release and renal sodium andinulin clearance to the sham level but did not alter renal blood flow.Pretreatment with 1-benzylimidazole restored renal function, eicosanoidrelease, and renal blood flow to sham levels. These data suggest thatsevere intestinal I/R contributes to the downregulation of renalfunction. The decrease in renal function is due in part to toxic oxygen metabolites, which occur in the milieu of altered renal eicosanoid release, reflecting a decrease in vasodilator and an increase invasoconstrictor eicosanoids.

     

The effects of platelet-activating factor on flow rate and eicosanoid release in the isolated perfused rat gastric vascular bed

In the isolated rat stomach perfused via the vasculature in situ under constant pressure bolus injections of platelet-activating factor (PAF, 3, 16, or 50 ng) induced dose-dependent, long-lasting reductions of flow rates and simultaneously significant increases in the release of cysteinyl-leukotrienes (cys-LT), thromboxane (TX) B2 and 6-keto-prostaglandin (PG) F1 alpha. Reversed phase high pressure liquid chromatography demonstrated the release of a mixture of comparable amounts of LTC4, LTD4 and LTE4 by PAF. Inhibition of cys-LT synthesis by the lipoxygenase inhibitors nordihydroguaiaretic acid (NDGA) and L-651,896 did not significantly affect PAF-induced flow reduction indicating that endogenous cys-LT are of minor importance for the PAF effect on gastric vascular flow. This conclusion is supported by the fact that the cys-LT receptor antagonist FPL 55712 in a concentration (1 x 10(-6) M) that completely antagonized gastric flow reduction by exogenous LTC4 (1 x 10(-7) M) had no effect on the PAF-induced reduction of flow. The cyclooxygenase inhibitor indomethacin aggravated the PAF-induced flow reduction suggesting that the endogenous vasodilator PGI2 might act as a functional PAF antagonist in the rat gastric vascular bed. In contrast to FPL 55712 the PAF antagonist BN 52021 significantly and concentration-dependently antagonized the PAF effect on gastric vascular flow. The results demonstrate that PAF and LTC4 induce flow reductions in the rat gastric vascular bed by activating different receptors and that endogenous eicosanoids released by PAF do not contribute significantly to the PAF effect on gastric vascular flow.     

Hydrolysis of cyclic GMP in rat peritoneal macrophages

Intact rat peritoneal macrophages (rPM) treated with 3-isobutyl-1-methylxanthine (IBMX), an inhibitor of phosphodiesterases (PDEs), accumulated more cGMP than untreated cells. A PDE activity toward [(3)H]cGMP was detected in the soluble and particulate fractions of rPM. The hydrolysis of cGMP was Ca(2+)/calmodulin-independent but increased in the presence of cGMP excess. Similar results were obtained when [(3)H]cAMP was used as a substrate. The hydrolytic activity towards both nucleotides was inhibited in the presence of IBMX. Therefore, the PDEs of families 2, 5, 10 and 11 are potential candidates for cGMP hydrolysis in the rPM. They may not only regulate the cGMP level in a feedback-controlled way but also link cGMP-dependent pathways with those regulated by cAMP.     

Caveolin isoforms in resident and elicited rat peritoneal macrophages

Caveolin--an integral membrane protein--is the principal component of caveolae membranes in vivo. Multiple forms of caveolin have been identified: caveolin-1alpha, caveolin-1beta, caveolin-2 and caveolin-3. They differ in their specific properties and tissue distribution. When we studied the lysate of resident and elicited macrophages isolated from rat peritoneal cavity by Western blot analysis, we identified two different proteins (approximately 29 kDa and approximately 20 kDa) which were labelled with anti-caveolin antibodies. The approximately 20-kDa protein was labelled specifically only by anti-VIP21/caveolin-1, while the approximately 29-kDa protein was labelled by anti-VIP21/caveolin-1 and anti-caveolin-2. The presence of the approximately 29-kDa protein was characteristic of resident macrophages, and only a small amount of the approximately 20-kDa protein was detected in these cells. Elicitation resulted in a significant increase in the amount of the approximately 20-kDa protein labelled by anti-VIP21/caveolin-1 only. According to its molecular mass and antibody-specificity, this protein might be identical with the caveolin-1beta isoform. Our morphological (confocal and electron microscopical) studies have shown that in resident cells caveolin was present in the cytoplasm, in smaller vesicles and multivesicular bodies around the Golgi area. Only a very small amount of caveolae was found on the surface of these cells. In elicited macrophages, caveolae (labelled with the anti-VIP21/caveolin-1 antibody) appeared in large numbers on the cell surface, but caveolin detected by anti-caveolin-2 was also found in small vesicles and multivesicular bodies in the cytoplasm. According to these results, the absence of caveolae in resident cells can be explained by the absence of caveolin-1. The expression of the approximately 29-kDa (caveolin-related) protein in resident macrophages seems to be insufficient for caveolae formation. Elicitation significantly increased the expression of caveolin-1, and the increased amount of caveolin-1 resulted in caveolae formation on the cell surface.     

The glucose-induced insulin release after adrenalectomy in the rat

Plasma glucose and insulin levels following glucose loading were investigated in adrenalectomized rats. Both oral and intravenous administration of glucose induced an elevation in plasma glucose and insulin level. The increases of plasma glucose and insulin concentrations were significantly higher in the adrenalectomized rats compared with the controls. We conclude, that corticoid hormones are capable of inhibiting glucose-induced insulin release in the rat.