Glucocorticoid sensitivity of interleukin-1 agonist and antagonist secretion: the effects of age and gender

Interleukin-1 (IL-1) is a primary mediator of inflammation that is regulated, in part, by the hypothalamic-pituitary-adrenal axis. The purpose of this study was to determine if gender- or age-related differences exist in the sensitivity of IL-1-producing cells to hydrocortisone. Peripheral blood mononuclear cells (PBMC) isolated from men and women (21-77 yr old) were incubated with hydrocortisone (0, 50, 100, 500, or 1,000 ng/ml) with or without lipopolysaccharide (LPS). Secretion of IL-1beta and IL-1 receptor antagonist was inhibited in a dose-dependent manner (P = 0.001) without age- or gender-related differences. Hydrocortisone decreased soluble IL-1 receptor type II (sIL-1RII) secretion by unstimulated cells (P = 0. 0001), but it increased secretion by LPS-stimulated cells (P = 0. 0001) in all groups. Unstimulated cell supernatants from men contained greater concentrations of sIL-1RII than the supernatants from women (P = 0.011). Compared with men, PBMCs from women were less responsive to hydrocortisone inhibition of sIL-1RII secretion, regardless of age (P = 0.001), and compared with the follicular phase, sIL-1RII secretion was lower in the luteal phase of the menstrual cycle (P < 0.05). These data indicate that basal secretion and glucocorticoid modulation of sIL-1RII secretion by cultured PBMCs are gender dependent. Moreover, glucocorticoid influences on sIL-1RII secretion depend on the presence or absence of gram-negative bacterial toxins.     

Antibiotics and peptides with agonist and antagonist chemotactic activity.

It has been found that the polypeptide antibiotics gramicidin S, tyrocidin and bacitracin, containing Leu-Phe or Ile-Phe sequences, are chemoattractants for neutrophils. Related synthetic pentapeptides containing the sequence Leu-Phe have stronger biological activities, provided the N-terminal residue is acylated. The formylated peptide f-L-Phe-D-Leu-L-Phe-D-Leu-L-Phe is a potent agonist (4 × 10^?9 M) whereas the t-butyloxycarbonyl analog is a good antagonist (8 × 10^?7 M).     

Brassinolide-2,3-acetonide: A brassinolide-induced rice lamina joint inclination antagonist

A novel chemical tool compound that is an antagonist of brassinolide (BL, 1)-induced rice lamina joint inclination was developed. Although 2-O-, 3-O-, 22-O-, or 23-O-methylation of BL causes a critical decrease in biological activity,⁵ a crystal structure of the extracellular leucine-rich repeat (LRR) domain of BRASSINOSTEROID-INSENSITIVE I (BRI1) bound to BL3, 4 indicates that the loss of activity of the O-methylated BL may result from not only the low affinity to BRI1, but also from blocking the interaction with another BR signaling factor, a partner protein of BRI1 (e.g., BRI1-ASSOCIATED KINASE 1, BAK1). On the basis of this hypothesis we synthesized the BL 2,3-acetonide 2, the 22,23-acetonide 3, and the 2,3:22,23-diacetonide 4 to assess the possibility of 2-O- and 3-O- or/and 22-O- and 23-O-alkylated BL as an antagonist in BR signaling evoked by exogenously applied BL. The 2,3-acetonide 2 more strongly inhibited the lamina inclination caused by BL relative to the 22,23-acetonide 3, whereas the diacetonide 4 had no effect most likely due to its increased hydrophobicity. This suggested that the 2,3-hydroxyl groups of BL play a more significant role in the interaction with a BRI1 partner protein rather than BRI1 itself in rice lamina joint inclination. Taken together it was demonstrated that BL, the most potent agonist of BRI1, is transformed into an antagonist by functionalization of the 2,3-dihydroxyl groups as the acetonide. This finding opens the door to the potential development of a chemical tool that modulates protein–protein interactions in the BR signaling pathway to dissect the BR-dependent processes.     

Non-core region modulates interleukin-11 signaling activity: generation of agonist and antagonist variants

Human interleukin-11 (hIL-11) is a pleiotropic cytokine administered to patients with low platelet counts. From a structural point of view hIL-11 belongs to the long-helix cytokine superfamily, which is characterized by a conserved core motif consisting of four α-helices. We have investigated the region of hIL-11 that does not belong to the α-helical bundle motif, and that for the purpose of brevity we have termed "non-core region." The primary sequence of the interleukin was altered at various locations within the non-core region by introducing glycosylation sites. Functional consequences of these modifications were examined in cell-based as well as biophysical assays. Overall, the data indicated that the non-core region modulates the function of hIL-11 in two ways. First, the majority of muteins displayed enhanced cell-stimulatory properties (superagonist behavior) in a glycosylation-dependent manner, suggesting that the non-core region is biologically designed to limit the full potential of hIL-11. Second, specific modification of a predicted mini α-helix led to cytokine inactivation, demonstrating that this putative structural element belongs to site III engaging a second copy of cell-receptor gp130. These findings have unveiled new and unexpected elements modulating the biological activity of hIL-11, which may be exploited to develop more versatile medications based on this important cytokine.     

New agonist and antagonist analogues of bradykinin

Three new analogues of bradykinin (BK) have been tested for their agonistic and antagonistic actions on the rabbit jugular vein and the guinea pig ileum (B2 receptors), and six were studied on rabbit aorta strips (B1 receptors). Substitution of Gly4, Phe5, and Phe8 in BK with D-Trp gives analogues with a relative affinity lower than 1.0% as compared with BK. These analogues have no antagonistic properties on the rabbit jugular vein and on guinea pig ileum (B2 receptors). Substitution of Pro7 in des-Arg9-BK by Gly and by D-Ala give compounds that antagonise the effects of kinins on the rabbit aorta strips (B1-receptor system). These new antagonists are fairly potent with a pA2 value of 6.03 to 7.29 and seem competitive because the pA2--pA10 values approximate 0.95. These results suggest that the orientation of Phe8 is critical for the activation of B1 receptors by kinins.     

Muscle pain induces task-dependent changes in cervical agonist/antagonist activity.

This study examined the effect of experimental neck muscle pain on the EMG-force relationship of cervical agonist and antagonist muscles. Surface EMG signals were detected from the sternomastoid, splenius capitis, and upper trapezius muscles bilaterally from 14 healthy subjects during cervical flexion and extension contractions of linearly increasing force from 0 to 60% of the maximum voluntary contraction (MVC). Measurements were performed before and after injection of 0.5 ml hypertonic and isotonic saline into either the sternomastoid or splenius capitis in two experimental sessions. EMG average rectified value (ARV) of the sternomastoid, splenius capitis, and upper trapezius muscles and the muscle fiber conduction velocity (CV) of the sternomastoid muscle were estimated at 5% MVC force increments. During cervical flexion with injection of hypertonic saline in sternomastoid, ARV of sternomastoid was lower on the side of pain in the force range 25-60% MVC (P < 0.05) and was associated with a bilateral reduction of splenius capitis and upper trapezius ARV (P < 0.01). During cervical extension, injection of hypertonic saline in splenius capitis resulted in lower estimates of splenius capitis ARV on the painful side from 45 to 60% MVC (P < 0.05), which was associated with a bilateral increase in upper trapezius ARV estimates from 50 to 60% MVC (P < 0.001). However, no significant change was identified for estimates of sternomastoid ARV. Experimentally induced neck muscle pain resulted in task-dependent changes in cervical agonist/antagonist activity without modifications in muscle fiber CV.     

Distributed moment histogram: A neurophysiology based method of agonist and antagonist trunk muscle activity prediction

A neurocortical-based technique of muscle recruitment is presented to solve the muscle indeterminacy problem for lumbar torso modeling. Cortical recordings from behaving primates have established motor cortex cells that respond to a wide range of task directions, but are tuned to a preferred direction. A characteristic activity pattern of these neurons seems to be associated with effort direction. It was hypothesized that a model which recruits muscles based on a similar distribution would predict antagonistic muscle activity with greater realism than a widely referenced optimization formulation. The predictions of the Distributed Moment Histogram (DMH) method were evaluated under common speed (<30^os⁻¹) sagittal plane lifting conditions using five subjects. The predicted forces showed high correspondence with agonist and antagonist myoelectric patterns. The mean coefficient of determination for the erector spinae was r²=0.91, and 0.41 for the latissimus. For the antagonistic muscles, the rectus abdominus was found to be electrically silent (<3% MVC) and no activity was predicted by the method. The external oblique muscle was observed to be minimally active (<16% MVC), and the DMH method predicted its mostly constant activity with a mean standard error of 1.6% MVC. The realistic antagonistic predictions supported the hypothesis and justify this cortical based technique as an alternative for muscle tension estimation in biomechanical torso modeling. A primary advantage of this method is its computational simplicity and direct physiologic analogy     

Microbiological transformation of steroids. 1-Dehydrogenation of 17,21-dihydroxypregn-4-ene-3,20-dione with Glomerella cingulata

The micro-organism Glomerella cingulata dehydrogenates 17,21-dihydroxypregn-4-ene-3,20-dione to 17,21-dihydroxypregna-1,4-diene-3,20-dione in high yield practically without by-products.     

Small molecules with similar structures exhibit agonist, neutral antagonist or inverse agonist activity toward angiotensin II type 1 receptor

Small differences in the chemical structures of ligands can be responsible for agonism, neutral antagonism or inverse agonism toward a G-protein-coupled receptor (GPCR). Although each ligand may stabilize the receptor conformation in a different way, little is known about the precise conformational differences. We synthesized the angiotensin II type 1 receptor blocker (ARB) olmesartan, R239470 and R794847, which induced inverse agonism, antagonism and agonism, respectively, and then investigated the ligand-specific changes in the receptor conformation with respect to stabilization around transmembrane (TM)3. The results of substituted cysteine accessibility mapping studies support the novel concept that ligand-induced changes in the conformation of TM3 play a role in stabilizing GPCR. Although the agonist-, neutral antagonist and inverse agonist-binding sites in the AT(1) receptor are similar, each ligand induced specific conformational changes in TM3. In addition, all of the experimental data were obtained with functional receptors in a native membrane environment (in situ).     

PQ-69, a novel and selective adenosine A1 receptor antagonist with inverse agonist activity

Potent and selective adenosine A₁ receptor (A₁AR) antagonists with favourable pharmacokinetic properties used as novel diuretics and antihypertensives are desirable. Thus, we designed and synthesized a series of novel 4-alkylamino substitution-2-arylpyrazolo[4,3-c]quinolin-3-one derivatives. The aim of the present study is to characterize the biological profiles of the optimized compound, PQ-69. In vitro binding assay revealed a K_i value of 0.96 nM for PQ-69 in cloned hA₁ receptor, which was 217-fold more selective compared with hA_2A receptors and >1,000-fold selectivity for hA₁ over hA₃ receptor. The results obtained from [³⁵S]-GTPγS binding and cAMP concentration assays indicated that PQ-69 might be an A₁AR antagonist with inverse agonist activity. In addition, PQ-69 displayed highly inhibitory activities on isolated guinea pig contraction (pA2 value of 8.99) induced by an A₁AR agonist, 2-chloro-N6-cyclopentyl adenosine. Systemic administration of PQ-69 (0.03, 0.3, 3 mg/kg) increased urine flow and sodium excretion in normal rats. Furthermore, PQ-69 displayed better metabolic stability in vitro and longer terminal elimination half-life (t_1/2) in vivo compared with 1,3-dipropyl-8-cyclopentylxanthine. These findings suggest that PQ-69 exhibits potent antagonist effects on A₁AR in vitro, ex vivo and in vivo, it might be a useful research tool for investigating A₁AR function, and it could be developed as a potential therapeutic agent.

Electronic supplementary material

The online version of this article (doi:10.1007/s11302-014-9424-5) contains supplementary material, which is available to authorized users.     

Discovery of 1,1-dioxo-1,2,6-thiadiazine-5-carboxamide derivatives as cannabinoid-like molecules with agonist and antagonist activity

A series of new 2-substituted 1,1-dioxo-1,2,6-thiadiazine-5-carboxylate derivatives have been prepared from monosubstituted sulfamides in order to obtain N-substituted 1,1-dioxo-1,2,6-thiadiazine-5-carboxamides as novel cannabinoid derivatives, analogues of Rimonabant (SR141716A). Their potential functional activity on cannabinoid receptors has been evaluated in vitro and in vivo in mice, showing that two compounds (37 and 39) behave as cannabinoid agonists in vitro. Their potency is lower than that of the reference compound, WIN 55,212-2, but their efficacy is similar to that of this cannabinoid agonist, although no in vivo activity is observed. Another derivative (38) behaves as a cannabinoid antagonist both in vitro and in vivo, being its efficacy and potency similar to that of the well-known antagonist SR141716A.     

Opioid agonist and antagonist bivalent ligands as receptor probes

Bivalent ligands are molecules which contain two pharmacophores linked by a connecting chain (spanner). The present report describes the use of oxymorphamine (Oxy) and naltrexamine (Nal) as the opioid agonist and antagonist pharmacophores separated by a variable length spanner composed of succinyl-bis-oligoglycine. The agonist series, [CH2CO(Gly)nOxy]2, and antagonist series, [CH2CO(Gly)nNal]2, were synthesized (n = 0-4) and tested on the electrically stimulated GPI. All of the antagonist bivalent ligands (Nal) antagonized the effects of morphine, with the greatest potency enhancement (60 x) residing with the succinyl (n = 0) congener. A dramatically different SAR profile was observed in the agonist (Oxy) series where the greatest potency enhancement (17 x) occurs when n = 2. By contrast with the antagonist series the agonist bivalent ligand with n = 0 is equipotent to its monovalent agonist analogue. The significance of these results with respect to the possibility of discrete opioid agonist and antagonist recognition sites are discussed.     

Ecdysteroid agonist and antagonist activities in species of the Solanaceae

Previously, it has been shown that certain withanolides from Iochroma gesnerioides (Solanaceae) possess ecdysteroid antagonistic activity. Phytoecdysteroids (agonists) are widely distributed in the plant world, but solanaceous species have not been extensively examined for their presence. We have now surveyed 128 species of solanaceous plants for the presence of ecdysteroid agonist and antagonist activities using the Drosophila melanogaster B(II) cell line bioassay. Only weak antagonistic activity was associated with a few of the methanolic extracts, including those from species known to contain high levels of withanolides. Therefore, the major withanolides are inactive per se, but they may be activated after ingestion by invertebrate predators. Several extracts possessed ecdysteroid agonist activity as a consequence of the presence of phytoecdysteroids. Phytoecdysteroid-accumulating species are at least as common in the Solanaceae as they are in plants in general. Preliminary characterization of the identities of the phytoecdysteroids present in the most active extracts has been performed by hplc separations on normal- and reversed-phase systems in conjunction with ecdysteroid-specific radioimmunoassay and bioassay. Each of the phytoecdysteroid-accumulating species examined (Browallia speciosa, Nierembergia hippomanica var violacea, N. solanacea and Solanum nigrum) contain a cocktail of ecdysteroids, of which 20-hydroxyecdysone and polypodine B (5beta,20-dihydroxyecdysone) are major components.     

The effects of antagonist prefatigue on agonist torque and electromyography

This study assessed the effects of hamstring prefatigue on peak torque, peak power, time to peak torque, knee angle of peak torque, and electromyography (EMG) activity of the hamstrings and quadriceps group during knee extensions at angular velocities of 60 degrees, 180 degrees, and 300 degrees.s(-1). Twenty Division I wrestlers performed 5 maximal knee extensions in prefatigued and nonfatigued conditions of the hamstring group. This study demonstrated that when the hamstrings were prefatigued, the quadriceps produced significant decreases in peak torque of 1.7% (p < 0.05), peak power of 11% (p < 0.05), and rate to peak torque of 6.4% (p < 0.01) as compared with the nonfatigued state. When the hamstrings were prefatigued, they produced a 25% greater amount of EMG activity during knee extension (p < 0.01) than when not prefatigued. There was no significant difference in quadriceps EMG activity whether the hamstring group was prefatigued or not (p > 0.05). The decrease in quadriceps peak torque during the prefatigued condition was more pronounced (p < 0.01) at an angular velocity of 60 degrees.s(-1) than at 180 degrees or 300 degrees.s(-1). In other words, prefatiguing the antagonist appears to be most detrimental to torque output of the quadriceps in the condition that most closely replicates the speed at which "isotonic" weight training occurs (60 degrees.s(-1)) and suggests a limitation to agonist-antagonist superset training.     

Biased signaling: potential agonist and antagonist of PAR2

Protease activated receptor 2 (PAR2) has emerged as one of the promising therapeutic targets to inhibit rapidly metastasizing breast cancer cells. However, its elusive molecular mechanism of activation and signaling has made it a difficult target for drug development. In this study, in silico methods were used to unfold PAR2 molecular mechanism of signaling based on the concept of GPCR receptor plasticity. Although, there are no conclusive evidences of the presence of specific endogenous ligands for PAR2, the efficacy of synthetic agonist and antagonist in PAR2 signaling has opened up the possibilities of ligand-mediated signaling. Furthermore, it has been proved that ligands specific for one GPCR can induce signaling in GPCRs belonging to other subfamilies. Therefore, the aim of this study was to identify potential agonists and antagonists from the GPCR ligand library (GLL), which may induce biased signaling in PAR2 using the concept of existence of multiple ligand-stabilized receptor conformations. The results of our in silico study suggest that PAR2 may show biased signaling mainly with agonists of serotonin type 1, β-adrenergic type 1,3 and antagonists of substance K (NK1), serotonin type 2, dopamine type 4, and thromboxane receptors. Further, this study also throws light on the putative ligand-specific conformations of PAR2. Thus, the results of this study provide structural insights to putative conformations of PAR2 and also gives initial clues to medicinal chemists for rational drug design targeting this challenging receptor.     

Antiglucocorticoid steroids have increased agonist activity in those hepatoma cell lines that are more sensitive to glucocorticoids

FU5-5 rat hepatoma (Reuber H35) cells are hypersensitive in that the same percentages of full induction of tyrosine aminotransferase (TAT) occur at much lower concentrations of glucocorticoids than in the related HTC rat hepatoma (Morris) cells. Unexpectedly, these hypersensitive FU5-5 cells also exhibited more agonist activity with the affinity labeling antiglucocorticoids cortisol 21-mesylate and dexamethasone 21-mesylate than did HTC cells (Mercier et al., Endocrinology 112, 601-609 [1983]). In the present study, several other antiglucocorticoids (11-desoxycortisone, progesterone, dexamethasone oxetanone, and RU 486 in addition to dexamethasone 21-mesylate) and the antiandrogen cyproterone acetate were examined to see if chemically unreactive, reversible antisteroids also would exhibit an altered activity (i.e. increased agonist activity) in FU5-5 cells. Each antiglucocorticoid examined did display a 2-fold increased amount of agonist activity in FU5-5 cells, as compared to HTC cells; only RU 486 was predominantly an antagonist in FU5-5 cells but the potency of RU 486 was about 9-fold less than in HTC cells. Dexamethasone, and especially progesterone, was metabolized in FU5-5 and HTC cells. However, differential metabolism in FU5-5 vs HTC cells cannot account for the increased induction of TAT in FU5-5 cells since the amount of agonist activity seen for dexamethasone mesylate (or its metabolites) depended not on the cell type used but rather on the glucocorticoid inducible enzyme monitored, i.e. TAT or glutamine synthetase. The combined data suggest that the hypersensitivity of FU5-5 cells towards glucocorticoid induction of TAT may be linked with the ability of both reversible and irreversible antiglucocorticoids to display increased TAT agonist activity in FU5-5 cells. This behavior was somewhat steroid specific since the antiandrogen cyproterone acetate did not display increased TAT agonist activity in FU5-5 cells compared to HTC cells and was only 2-fold less effective as an antiglucocorticoid in FU5-5.