Suppression of T lymphocyte chemotactic factor production by the opioid peptides beta-endorphin and met-enkephalin

The opioid peptides beta-endorphin and met-enkephalin have been shown to modulate human lymphocyte proliferation, mononuclear cell locomotion, natural killer cell activity, and neutrophil locomotion. This study demonstrates that beta-endorphin and met-enkephalin inhibit the production of a T lymphocyte chemotactic factor (LCF) by concanavalin A (Con A)-stimulated peripheral blood mononuclear cells. Inhibition of LCF production was observed by using concentrations of 10(-11) to 10(-6) M beta-endorphin or met-enkephalin but not alpha-endorphin. A bimodal pattern of suppression of LCF production was observed with both met-enkephalin and beta-endorphin when titrated from 10(-12) to 10(-6) M concentrations, with the peaks of suppressive activity occurring at concentrations of 10(-11) M and 10(-6) M. Timed studies of the production of LCF over a 54-hr period showed that there was an appreciable lag in the onset of measurable LCF activity in mononuclear supernatants produced in the presence of beta-endorphin and met-enkephalin. The suppression of LCF production mediated by opioid peptides in mononuclear supernatants was abrogated by depletion of glass-adherent mononuclear cells before culturing with opioids and Con A. The inhibitory effect of opioid peptides on LCF production was prevented by the addition of indomethacin to cell cultures. Additional experiments showed that exogenous prostaglandin E2 (PGE2) suppressed Con A-stimulated LCF production when added at concentrations ranging from 10(-6) to 10(-8) M. Other studies suggested that the mechanism of opioid peptide-mediated suppression of LCF production was due to an enhanced sensitivity of mononuclear cells to the inhibitory action of PGE2. These data provide further evidence for modulation of the immune response in humans by the neuroendocrine hormones beta-endorphin and met-enkephalin and further suggest a link between this modulation and arachidonic acid metabolism.     

Enhancement of natural cytotoxicity by beta-endorphin

The role of enkephalins, beta-endorphin, or other neuropeptides produced by the nervous system in the alteration of immune responsiveness is generally unknown. The present studies were undertaken to investigate the role of these neuropeptides in the modulation of human spontaneous cytotoxicity induced by natural killer (NK) cells. Natural cytotoxicity was measured by using a standard 51Cr release assay with radiolabeled K562 cells. NK activity was significantly enhanced by both beta-endorphin (30.5 +/- 11.5%, M +/- SE, relative enhancement at 50:1, effector:target (E:T) ratio, 10(-14)M beta-endorphin) and methionine-enkephalin (met-enkephalin) (27.4 +/- 9.7% relative enhancement at 10(-9)M). The magnitude of relative enhancement significantly correlated with increasing concentrations of beta-endorphin. Leucine-enkephalin, alpha-endorphin, and morphine did not augment NK activity. The enhancement of NK activity with beta-endorphin increased at all E:T ratios tested. Naloxone inhibited the augmentation of NK activity produced by beta-endorphin and met-enkephalin. By using a combination of a standard 51Cr release and soft agarose single cell analysis assays, beta-endorphin increased both the number of E:T cell conjugates and the number of active killer cells among target-binding cells. The maximal effector cell recycling capacity was increased by 170%. These studies provide new insight into the mechanisms by which neuropeptides produced by the nervous system can alter immune responsiveness.     

Effects of oxytocin alone and in combination with selected hypothalamic hormones on ACTH, beta-endorphin, LH and PRL secretion by anterior pituitary cells of cyclic pigs

Oxytocin (OT) is involved in the stimulation of secretion of anterior pituitary hormones in females during the periovulatory and periparturient periods. In the present study we examined the role of OT in control of ACTH, beta-endorphin, LH and PRL secretion in vitro from dispersed anterior pituitary cells collected from gilts during the luteal (Days 10-12; n=6) and follicular (Days 18-20; n=5) phases of the estrous cycle. Isolated anterior pituitary cells (1 x 10(6)/ml) were transferred into 24-well plates, separately for each animal, and were pre-incubated for three days at 37 degrees C in atmosphere of 5% CO(2) and 95% air. The cells which attached to the dishes were incubated (3.5 h, 37 degrees C) in McCoy's medium in the absence (control) or in the presence of the following factors: CRH alone (10(-10), 10(-9), 10(-8), 10(-7) M), OT alone (10(-8), 10(-7), 10(-6) M), LVP alone (10(-7) M), OT (10(-7) M) plus CRH (10(-9) M) and LVP (10(-7) M) plus CRH (10(-9) M) for studying ACTH and beta-endorphin secretion; OT alone (10(-8), 10(-7), 10(-6) M), GnRH alone (100 ng/ml), CRH alone (10(-9) M), OT (10(-7) M) plus GnRH (100 ng/ml) and OT (10(-7) M) plus CRH (10(-9) M) for studying LH and PRL secretion. Concentrations of the studied hormones in media were analyzed by RIA. Oxytocin alone increased ACTH (at doses 10(-7), 10(-6) M), beta-endorphin (at dose 10(-8) M), LH (at dose 10(-8) M) and PRL (at doses 10(-7), 10(-6) M) secretion by pituitary cells isolated only from luteal-phase gilts. None of the studied hormone concentrations in the medium was increased in response to OT when pituitary cells of follicular-phase gilts were examined. Oxytocin in combination with CRH exerted an additive effect on beta-endorphin secretion during the luteal phase. Summarizing, in the present study the stimulatory effect of oxytocin on ACTH, beta-endorphin, LH and PRL secretion by pituitary cells isolated from gilts during the luteal phase was demonstrated. However, the cells collected from follicular-phase gilts appeared to be unresponsive to OT. Moreover, interaction between OT and CRH in affecting beta-endorphin secretion was shown. These results suggest that OT may be transiently involved in the modulation of anterior pituitary hormone secretion in cyclic pigs.     

Opioid peptides modulate production of interferon gamma by human mononuclear cells

The opioid neuropeptides have previously been shown to bind to and affect leukocyte function including lymphocyte proliferation, NK-cell activity, mononuclear cell chemotaxis, immunoglobulin synthesis, and lymphokine production. The effect of the opioid peptides beta-endorphin and Met-enkephalin on interferon gamma (IFN) production by concanavalin A-stimulated human mononuclear cells was examined. Both beta-endorphin and Met-enkephalin enhanced IFN production by the majority of donor mononuclear cells tested and did so at concentrations between 10(-14) and 10(-10) M. When 10(-12) M beta-endorphin or Met-enkephalin were included in concanavalin A-stimulated mononuclear cell cultures, IFN concentrations were significantly enhanced to 205 +/- 45 and 252 +/- 67% of control, respectively. Although the majority of cell preparations tested exhibited an enhanced production of IFN in response to these opioid peptides, some did not. When beta-endorphin or Met-enkephalin were utilized at 10(-11) M, 10 of 15 and 7 of 11 responded with IFN production greater than 20% above the control (untreated) level. There was not an absolute correlation between an enhanced response to beta-endorphin and Met-enkephalin, suggesting the presence of multiple receptor types on these cells for opioids. The opioid receptor antagonist, naloxone, did not significantly prevent the opiate effect. When 10(-8) M naloxone was included in cultures containing 10(-12) M beta-endorphin or Met-enkephalin no significant inhibition of the effect of either opioid on IFN production was observed.     

Combinative ligand-receptor interactions: epinephrine depresses RAW264 macrophage antibody-dependent phagocytosis in the absence and presence of met-enkephalin

Macrophages have been shown to possess cell surface receptors for opiates and catecholamines. The abilities of these ligands to affect RAW264 macrophage antibody-dependent effector activity directed against sheep red blood cells were tested. Phagocytosis was measured by the uptake of 51Cr labeled erythrocytes and optical microscopy. Cytolysis was measured by 51Cr-release assays. Met-enkephalin increased specific antibody-dependent phagocytosis in a dose-dependent fashion; the optimal dose was found to be 10(-8) M. Epinephrine diminished phagocytosis in a dose-dependent manner exhibiting a maximal inhibition at 10(-4)-10(-5) M. This inhibition can be blocked by propranolol. The combined effects of simultaneous treatment with met-enkephalin and epinephrine were measured. At the several doses tested, the combined effects of these two ligands on the amount of phagocytosis were equivalent to or more inhibitory than epinephrine alone. Thioglycolate-elicited murine peritoneal macrophages demonstrated similar responses to epinephrine, met-enkephalin, and their combination. Therefore, in vitro models more closely approximating in vivo neuroregulation of macrophage function demonstrate phagocytic inhibition.     

Release of immunoreactive met-enkephalin by intraventricular beta-endorphin in anesthetized rats

The release of immunoreactive met-enkephalin and leu-enkephalin from the spinal cord by intraventricular injection of different doses of beta-endorphin was studied using the intrathecal perfusion technique. The intraventricular beta-endorphin elicited the release of immunoreactive met-enkephalin from the spinal cord in a dose dependent manner. Immunoreactive leu-enkephalin in the spinal perfusate was not increased after intraventricular beta-endorphin injection. Both immunoreactive met-enkephalin and leu-enkephalin in the spinal cord were not changed by low doses (2-6 micrograms) of beta-endorphin but were increased markedly by 60-70% after high doses of beta-endorphin (32-64 micrograms). It is likely that the biosynthesis of enkephalins was also increased after intraventricular beta-endorphin injection. Intraventricular naloxone, 30 micrograms did not induce any release of immunoreactive met-enkephalin from the spinal cord and did not block the release of immunoreactive met-enkephalin induced by intraventricular beta-endorphin, 15 micrograms.     

Human neutrophil migration is enhanced by beta-endorphin

Using a serum-free chemotaxis-under-agarose assay, we measured the effect of beta-endorphin on directed migration of human neutrophils toward 10(-7) M N-formyl-methionyl-leucyl-phenylalanine (FMLP). Neutrophils were pre-incubated with a range of beta-endorphin concentrations. beta-endorphin enhanced migration of neutrophils toward FMLP. This effect was maximal at 10(-9) M beta-endorphin. Naloxone inhibited the beta-endorphin effect, suggesting that enhanced migration is mediated via an opiate receptor.     

Selective regulation of CD8 effector T cell migration by the p110 gamma isoform of phosphatidylinositol 3-kinase

Chemokine-mediated T cell migration is essential to an optimal immune response. The p110gamma isoform of PI3K is activated by G protein-coupled receptors and regulates neutrophil and macrophage chemotaxis. We used p110gamma-deficient mice to examine the role of p110gamma in CD8 T cell migration and activation in response to viral challenge. Naive CD8 T cell migration in response to CCL21 in vitro and trafficking into secondary lymphoid organs in vivo was unaffected by the loss of p110gamma. Furthermore, loss of p110gamma did not affect CD8 T cell proliferation and effector cell differentiation in vitro in response to anti-CD3 stimulation or in vivo in response to vaccinia virus (VV) challenge. However, there was reduced migration of p110gamma knockout (p110gamma(-/-)) CD8 effector T cells into the peritoneum following i.p. challenge with VV. The role of p110gamma in CD8 effector T cell migration was intrinsic to T cells, as p110gamma(-/-) CD8 effector T cells exhibited impaired migration into the inflamed peritoneum following secondary transfer into wild-type recipients. In addition, p110gamma(-/-) CD8 effector T cells exhibited impaired migration in vitro in response to inflammatory chemoattractants. Although wild-type mice efficiently cleared VV at high viral doses, infection of p110gamma knockout mice resulted in visible illness and death less than a week after infection. Thus, p110gamma is dispensable for constitutive migration of naive CD8 T cells and subsequent activation and differentiation into effector CD8 T cells, but plays a central role in the migration of effector CD8 T cells into inflammatory sites.     

A comparative study of met-enkephalin effects on the secretion by murine lymphocytes of antibodies to different antigens]

The influence of met-enkephalin on specific antibody production by lymphocytes from mouse lymph nodes was studied in vitro in productive phase of immune response. It was shown that the peptide did not influence secretion of IgM-antibody to T-independent antigen-trinitrobenzensulfoacidic group, but suppressed secretion of IgG-antibody to T-dependent antigens both during primary and secondary response. The efficiency of superlow concentrations of the peptide (10(-15)-10(-14) M) for the response to ovalbumin, but not for the response to bovine gamma-globulin was shown. All effects of met-enkephalin were naloxone-reversible. The existence of individual distribution in dose-dependences of peptide action on antibody secretion in response to ovalbumin was demonstrated.     

In vivo and in vitro effects of angiotensin II on the release of beta-endorphin-like immunoreactivity

The effect of angiotensin II (A II) on the release of beta-endorphin-like immunoreactivity (beta-END-LI) in rats was studied in vivo and in vitro. Intravenous injection of 1 microgram/100 g body weight of A II resulted in significant increase in the plasma beta-END-LI level after 10 and 20 min. Intraventricular injection of 1 ng/100 g body weight of A II also resulted in significant increase in the plasma beta-END-LI level after 10 min. A II at concentrations of 10(-12) M-10(-10) caused dose-dependent stimulation of beta-END-LI release from dispersed cells of rat anterior pituitary. On gel chromatography, the beta-END-LI released by incubation of the cells with 10(-10) M A II separated into two components which eluted in the same positions as human beta-lipotropin and human beta-endorphin, respectively. The ratio of beta-LPH to beta-END in these fractions was 5:1 on a molar basis. A II did not stimulate beta-END-LI release in Ca++-free-medium. [Sar1, Ala8]-A II at concentrations of 10(-9) M - 10(-7) M did not stimulate beta-END-LI release from the cells. Addition of [Sar1, Ala8]-A II to the incubation medium inhibited A II-induced beta-END-LI release from the cells. These results indicate that A II acts, at least in part, directly on anterior pituitary cells to stimulate beta-END-LI release and that calcium ion is involved in the mechanism of this effect.     

Combinative ligand-receptor interactions: effects of cAMP, epinephrine, and met-enkephalin on RAW264 macrophage morphology, spreading, adherence, and microfilaments

Cell surface ligand-receptor interactions play a central role in the regulation and expression of macrophage function. Included among these macrophage membrane receptors are the beta-adrenergic and opioid receptors. We studied the abilities of epinephrine, met-enkephalin, forskolin, and adenosine 3':5' cyclic monophosphate (cAMP) analogues to affect macrophage morphology, spreading, and adherence. Cell spreading was quantitated by measuring the perimeters of adherent cell images recorded by videomicroscopy. Epinephrine induced a dose-dependent decrease in macrophage spreading; at 10(-5) M epinephrine the mean perimeter was 10.4 +/- 0.3 microns in comparison to 15.0 +/- 1.0 microns for controls. The inhibition of spreading can be blocked by the antagonist propranolol. On the other hand, met-enkephalin induced a dose-dependent increase in macrophage spreading, with a perimeter of 18.5 +/- 1.0 microns at 10(-8) M. Since catecholamines and opioids are simultaneously released from chromaffin cells of the adrenal, we examined the combinative effects due to treatment with both ligands. When macrophages were exposed to 10(-5) M epinephrine and 10(-8) M met-enkephalin, cell morphology and spreading were indistinguishable from that due to 10(-5) M epinephrine alone. The epinephrine dose-response curve in the presence of 10(-8) M met-enkephalin was similar to that of epinephrine alone. The beta-adrenergic receptor is apparently capable of diminishing or abrogating the opioid receptor signal(s). These combinative and epinephrine-mediated effects may be at least partially accounted for by the action of cAMP. Forskolin and the cAMP analogues N6-2'-O-dibutyryladenosine 3':5' cyclic monophosphate (dbcAMP) and 8-bromoadenosine 3':5' cyclic monophosphate (Br-cAMP) affected cell morphology and spreading in the same fashion as epinephrine. These differences in morphology and spreading behavior were accompanied by changes in the distribution of F-actin, as judged by phalladicin staining and fluorescence microscopy. We suggest that cAMP and microfilaments play important roles in receptor-mediated neuroregulation of macrophage function.     

Release of prostaglandin E2 and thromboxane B2 by mixed isolated human lung cells

Human lung specimens were minced and treated for 30 min with collagenase (1 mg ml-1) and DNase (0.1 mg ml-1) to obtain a suspension of viable (approximately 80%) and metabolically active lung cells (5 x 10(6) cells per gram of tissue). Treatment of these mixed lung cells with bradykinin (1.25 x 10(-6) to 1 x 10(-5) M) and f-Met-Leu-Phe (f-MLP; 1 x 10(-8) to 5 x 10(-6) M) did not stimulate to a substantial extent the release of prostaglandins and thromboxanes (measured with novel Enzyme Immunoassays). The only concentration of PAF that stimulated significantly the release of icosanoids from lung cells was 5 x 10(-7) M. Phorbol myristate (PMA; 5 x 10(-8) to 2 x 10(-6) M) and ionophore a-21387 (2.5 x 10(-6) to 2 x 10(-5) M) strongly stimulated the release of prostaglandins and thromboxanes by dispersed human lung cells. These findings support previous observations showing that human lungs have the enzymes necessary for the synthesis and release of prostaglandins and thromboxanes but stimulation of the release of these mediators is not obtained with the hormonal stimuli that are active in guinea pigs. Studies in progress will purify the cell populations and characterize the cells responsible for the release of these icosanoids.     

Effects of proopiomelanocortin-derived peptides, methionine-enkephalin and forskolin on the maturation of ovine fetal adrenal cells in culture

The present study examined the effects of peptides derived from the non-ACTH part of proopiomelanocortin (POMC), of met-enkephalin and of forskolin, alone or in combination with ACTH-(1-24), on the development of the ability of ovine fetal adrenal cells to produce both cAMP and corticosteroids in culture. N-POMC-(1-61) amide, gamma 2-melanocyte-stimulating hormone (MSH) and gamma 3-MSH behaved similarly in our system: 1) they increased slightly corticosterone (B) and cortisol (F) production without modification of cAMP output when added alone to the incubation medium, but this effect was observed only after 3 days in culture; 2) they potentiated the steroidogenic response to 10(-11) M ACTH-(1-24) but again only from Day 3 onwards; and 3) a 5-day treatment with the peptides induced fetal adrenal cell maturation resulting in the same enhancement of the B + F production stimulated by 10(-8) M ACTH-(1-24) without modification of the response in both cAMP and pregnenolone (P5). N-POMC-(1-80) and beta-lipotropic hormone (LPH) shared several common features in that, 1) the stimulation of B + F production by each of them alone was always significant and higher than that obtained with the other POMC-derived peptides [except ACTH-(1-24]; 2) they did not potentiate the steroidogenic action of 10(-11) M ACTH-(1-24); and 3) when cells were cultured in their presence for 5 days it resulted in an enhancement of the response to ACTH-(1-24), not only in B + F production but also in cAMP and P5 outputs. No effect of met-enkephalin was observed. The development of cAMP and B + F responses to ACTH-(1-24) provoked by forskolin was very close to that induced by the hormone itself, but forskolin, as opposed to ACTH-(1-24), was unable to induce a desensitization of the cAMP response. These data show that N-POMC-derived peptides can potentiate the acute steroidogenic activity of ACTH-(1-24) on ovine fetal adrenal cells after several days in culture.     

Influence of opioid peptides on human neutrophil apoptosis and activation in vitro

BACKGROUND: It has been shown that cells of the immune system release opioid peptides and possess receptors for them. The concentrations of opioid peptides in the peripheral circulation rapidly increase during inflammation and acute stress response. AIMS: The effect of opioid peptides Met-enkephalin (M-ENK) and beta-endorphin (beta-END) on the oxidative metabolism of normal human neutrophils and their death by apoptosis in vitro was investigated. METHODS: Isolated from peripheral blood, neutrophils were incubated in the presence or absence of 10(-6) to 10(-10) M of M-ENK and beta-END for 12 and 18 h. Apoptosis of neutrophils was determined in vitro by flow cytometric analysis of cellular DNA content and Annexin V-FITC protein binding to the cell surface. The MTT-reduction assay was employed to estimate the oxidative metabolism of neutrophils. RESULTS: Treatment with M-ENK caused a significant increase in apoptotic cells after 18 h of culture: *0 M (control) versus 10(-10) M, p < or = 0.02; **10(-10) M versus 10(-10) M, p < or = 0.02. Treatment with beta-END caused a significant increase in apoptotic cells after 12 h of culture: 0 M versus 10(-8) M, p < or = 0.03; **0 M versus 10(-10) M, p < or = 0.04. We found the significant increase in MTT reduction by neutrophils in the presence of M-ENK and beta-END both before and after the culture. However, the ability of neutrophils to reduce the MTT salt to formazan decreased significantly after the culture. CONCLUSIONS: We observed that the in vitro effect of opioid peptides on the neutrophil survival and their functional state was time and dose dependent. The presence of antioxidants in the culture medium modifies neutrophil survival.     

Effects of beta-endorphin on spontaneous uterine contractions. Prostaglandins production and 45Ca2+ uptake in uterine strips from ovariectomized rats.

The effects of beta-endorphin, Met-enkephalin, dynorphin and SKF 10047 on the constancy of the isometric developed tension (IDT) of the spontaneous contractions of uterine strips isolated from ovariectomized rats were explored. beta-endorphin (10(-6) M) was the only opioid that depressed significantly uterine constancy of IDT in a concentration dependent fashion. Naloxone, neither at 10(-8) M nor at 10(-6) M, altered the negative inotropic influence of beta-endorphin. Moreover, the basal synthesis and outputs of some prostaglandins (PGE1, PGE2 and PGF2 alpha) from rat uteri and the effect of beta-endorphin (10(-6) M), were determined. It was found that the basal synthesis and release of PGs in uteri were significantly inhibited by this endogenous opioid. The effects of beta-endorphin (10(-8), 10(-6) and 10(-5) M) on the basal; and oxytocin or A23187, induced 45Ca2+ uptake, as well as the influence of naloxone were also studied. beta-endorphin at three of the concentrations tested decreased basal uterine 45Ca2+ uptake and this action was not prevented by naloxone (10(-8) M). The presence of oxytocin and of A23187 augmented significantly 45Ca2+ uptake, an effect that was antagonized by beta-endorphin (10(-6) M). The possible role of beta-endorphin in uterine functioning via the modulation of uterine PG synthesis and Ca2+ uptake is discussed.     

Chlorpropamide alcohol flush and circulating met-enkephalin: a positive link.

Chlorpropamide-alcohol flushing may be due to sensitivity to endogenous opiates. To investigate this possibility the plasma met-enkephalin and beta-endorphin responses to sherry with and without chlorpropamide were studied in six patients with non-insulin dependent diabetes and in six normal subjects. After chlorpropamide all patients showed a rise in met-enkephalin concentrations from a basal level of 50 +/- 7.2 ng/l to a peak of 75 +/- 8.1 ng/l (p less than 0.001). In contrast, before chlorpropamide treatment was started met-enkephalin values did not change after alcohol. No significant changes in beta-endorphin values were observed. In six normal subjects pretreated with chlorpropamide the met-enkephalin concentration also rose from a basal level of 72 +/- 15 ng/l to a peak of 103 +/- 9.4 ng/l (p less than 0.002). Again, the met-enkephalin rise was not observed after placebo. Neither beta-endorphin concentrations nor facial temperature changed significantly. These data suggest that endogenous opiates may be implicated in CPAF. Furthermore, this is the first study in which a significant change in circulating met-enkephalin values has occurred.     

Chemotaxis of large granular lymphocytes

The hypothesis that large granular lymphocytes (LGL) are capable of directed locomotion (chemotaxis) was tested. A population of LGL isolated from discontinuous Percoll gradients migrated along concentration gradients of N-formyl-methionyl-leucyl-phenylalanine (f-MLP), casein, and C5a, well known chemoattractants for polymorphonuclear leukocytes and monocytes, as well as interferon-beta and colony-stimulating factor. Interleukin 2, tuftsin, platelet-derived growth factor, and fibronectin were inactive. Migratory responses were greater in Percoll fractions with the highest lytic activity and HNK-1+ cells. The chemotactic response to f-MLP, casein, and C5a was always greater when the chemoattractant was present in greater concentration in the lower compartment of the Boyden chamber. Optimum chemotaxis was observed after a 1 hr incubation that made use of 12 micron nitrocellulose filters. LGL exhibited a high degree of nondirected locomotion when allowed to migrate for longer periods (greater than 2 hr), and when cultured in vitro for 24 to 72 hr in the presence or absence of IL 2 containing phytohemagluttinin-conditioned medium. The chemotactic LGL was HNK-1+, OKT11+ or HNK-1+, OKT11- on the basis of monoclonal antibody and complement depletion. They did not bear either T cell or monocyte cell surface markers, exhibiting an OKT3-, OKT4-, OKT8-, OKM1-, and MO2- phenotype, and did not form E rosettes at 29 degrees C, which is characteristic of lytic NK cells in contrast to T cells. Furthermore, a rat LGL leukemia (RNK) exhibited a chemotactic response to both f-MLP and casein. LGL chemotaxis to f-MLP could be inhibited in a dose-dependent manner by the inactive structural analog CBZ-phe-met, and the RNK tumor line specifically bound f-ML[3H]P, suggesting that LGL bear receptors for the chemotactic peptide.     

Human endothelial cells are chemotactic to endothelial cell growth factor and heparin

The response of human endothelial cell migration to various extracellular matrix components and growth factors has been assessed. Human endothelial cells demonstrate increased chemotaxis and chemokinesis when placed in a modified Boyden chamber with endothelial cell growth factor (ECGF) used at a concentration of 10(-9) M. Anti-ECGF antibody inhibits the chemotactic response. Heparin (10(-8) to 10(-10) M) was also chemotactic and was shown to potentiate the chemotactic activity of ECGF. Although laminin, fibronectin, the polypeptide (epidermal, fibroblast, and nerve) growth factors, and collagen types I, II, III, IV, and V demonstrate a chemotactic response, these activities were one third to one half less than observed with ECGF. These data suggest that ECGF and heparin may play a significant role as response modifiers of human endothelial cell migration which may be relevant to tumor metastasis, wound healing, and atherogenesis.     

Glucocorticoid inhibition of immunoreactive beta-endorphin release from the anterior lobe of the rat pituitary: in vitro and in vivo studies

Glucocorticoid control of pituitary beta-endorphin (beta-END) release was investigated in vitro and in vivo. Cultured cells of both rat anterior (AL) and neurointermediate (NIL) lobe released beta-END-like immunoreactivity (beta-END-LI) in response to epinephrine (10(-7) M); however, only the response of AL cells was prevented by corticosterone (10(-8)-10(-6) M) or dexamethasone (10(-9)-10(-7) M). Gel chromatographic analysis (Sephadex G-50) revealed that the major forms of beta-END-LI released by AL cells corresponded to beta-END and beta-lipotropin (beta-LPH) in molecular size, whereas virtually all of the immunoreactivity released by NIL cells resembled beta-END. In vivo administration of dexamethasone attenuated the stress-induced release of beta-END-LI in a dose- and time-related fashion, having a more pronounced effect on plasma levels of beta-END-LI corresponding to beta-LPH in molecular size. Metyrapone (100 mg/kg), an inhibitor of glucocorticoid synthesis, evoked a rapid (20-40 min) four- to sixfold increase in total plasma beta-END-LI and 75% of this rise was due to immunoreactivity resembling beta-LPH in size. This response was diminished by coadministration of either dexamethasone (0.05-1.25 mg/kg) or corticosterone (0.05-1.25 mg/kg) and completely prevented by 4-hr pretreatment with dexamethasone (50 micrograms/kg). The briskness of the plasma beta-END-LI response to acute changes in glucocorticoid status suggests that a "rapid" feedback mechanism operates in the physiologic control of pituitary beta-END-LI secretion. Moreover, the ability of glucocorticoids to selectively inhibit AL release of beta-END-LI in vitro and their pronounced effect on plasma levels of beta-END-LI resembling beta-LPH, a marker of AL secretion, together indicate that glucocorticoids exert a selective influence over the secretion of AL corticotrophs in vivo. This demonstration of differential regulation of the AL versus IL secretion of beta-END-LI in vivo most likely reflects a phenomena having biologic importance related to the different physiologic actions of the several molecular forms of beta-END-LI secreted by the two tissues.     

Regulation of IL-1beta and IL-8 production by mu-, delta-opiate receptors agonists in vitro

The beta-endorphin 10(-7-)-10(-11) M in LPS (lypopolisaccharide) presence and in spontaneous cultures promoted the IL-1beta production in mixed leukocyte fraction. LPS-induced IL-8 production in leukocyte fraction was inhibited by beta-endorphin 10(-7), 10(-11) M. The enchasing effect of beta-endorphin on IL-1beta production was not blocked by naloxone and naltrindole. The inhibitory effect of beta-endorphin on IL-8 production was blocked by naloxone and naltrindole. In mononuclear and neutrophile fractions beta-endorphin and delta-agonist DADLE enchased IL-1beta production in spontaneous and LPS-stimulating cultures, when IL-8 production inhibited beta-endorphin and delta-agonist DADLE only in LPS presence. No effect of mu-agonist DAGO were observed on IL-1beta production, whereas LPS-induced IL-8 secretion in neutrophile fraction inhibited by DAGO.