Renata Giorgi

Hemopressin is an inverse agonist of CB1 cannabinoid receptors.

To date, the endogenous ligands described for cannabinoid receptors have been derived from membrane lipids. To identify a peptide ligand for CB1 cannabinoid receptors, we used the recently described conformation-state sensitive antibodies and screened a panel of endogenous peptides from rodent brain or adipose tissue. This led to the identification of hemopressin (PVNFKFLSH) as a peptide ligand that selectively binds CB1 cannabinoid receptors. We find that hemopressin is a CB1 receptor-selective antagonist, because it is able to efficiently block signaling by CB1 receptors but not by other members of family A G protein-coupled receptors (including the closely related CB2 receptors). Hemopressin also behaves as an inverse agonist of CB1 receptors, because it is able to block the constitutive activity of these receptors to the same extent as its well characterized antagonist, rimonabant. Finally, we examine the activity of hemopressin in vivo using different models of pain and find that it exhibits antinociceptive effects when administered by either intrathecal, intraplantar, or oral routes, underscoring hemopressins therapeutic potential. These results represent a demonstration of a peptide ligand for CB1 cannabinoid receptors that also exhibits analgesic properties. These findings are likely to have a profound impact on the development of novel therapeutics targeting CB1 receptors.

Antinociception induced by epidural motor cortex stimulation in naive conscious rats is mediated by the opioid system.

Epidural motor cortex stimulation (MCS) has been used for treating patients with neuropathic pain resistant to other therapeutic approaches. Experimental evidence suggests that the motor cortex is also involved in the modulation of normal nociceptive response, but the underlying mechanisms of pain control have not been clarified yet. The aim of this study was to investigate the effects of epidural electrical MCS on the nociceptive threshold of naive rats. Electrodes were placed on epidural motor cortex, over the hind paw area, according to the functional mapping accomplished in this study. Nociceptive threshold and general activity were evaluated under 15-min electrical stimulating sessions. When rats were evaluated by the paw pressure test, MCS induced selective antinociception in the paw contralateral to the stimulated cortex, but no changes were noticed in the ipsilateral paw. When the nociceptive test was repeated 15 min after cessation of electrical stimulation, the nociceptive threshold returned to basal levels. On the other hand, no changes in the nociceptive threshold were observed in rats evaluated by the tail-flick test. Additionally, no behavioral or motor impairment were noticed in the course of stimulation session at the open-field test. Stimulation of posterior parietal or somatosensory cortices did not elicit any changes in the general activity or nociceptive response. Opioid receptors blockade by naloxone abolished the increase in nociceptive threshold induced by MCS. Data shown herein demonstrate that epidural electrical MCS elicits a substantial and selective antinociceptive effect, which is mediated by opioids.

Antinociceptive effect of the calcium-binding protein MRP-14 and the role played by neutrophils on the control of inflammatory pain

Macrophages secrete a variety of chemical mediators that play a central role in the pathophysiology of inflammatory pain. Therefore, the activation or deactivation of these cells in an inflammatory focus could modulate the intensity of the algogenic response. Based on these premises and on our previous demonstration that the calcium‐binding protein MRP‐14, highly expressed in neutrophils, deactivates activated macrophages in vitro, we decided to investigate the role of MRP‐14 and of neutrophils in the control of inflammatory pain in mice. Our results show that this protein is endowed with antinociceptive activity. When tested in the writhing model it was able to inhibit pain response but did not change the behavior of the animals in the hot plate test. This observation indicates that MRP‐14 down‐regulates inflammatory but not central pain. Using a model of acute neutrophilic peritonitis induced by glycogen, a close correlation between neutrophil migration and antinociception was detected. Surgical adrenalectomy demonstrated that the antinociceptive response induced by glycogen was not due to endogenous liberation of glucocorticoids. The treatment of animals either with a monoclonal antibody anti‐MRP‐14 or a monoclonal antibody that depletes the animals of neutrophils reverts the antinociceptive response observed in the glycogen‐induced peritonitis. These data define the calcium‐binding protein MRP‐14 as a novel mediator for the control of inflammatory pain and consequently discloses an anti‐inflammatory role for the neutrophil. J. Leukoc. Biol. 64: 214–220; 1998.

Antinociceptive action of hemopressin in experimental hyperalgesia.

Endogenous hemorphins, derived from degradation of the beta-chain of hemoglobin, lower arterial blood pressure and exert an antinociceptive action in experimental models of nociception. Hemopressin, derived from the alpha-chain of hemoglobin, also decreases blood pressure, but its effects on pain have not been studied. In this work, we examined the influence of hemopressin on inflammatory pain. Hemopressin reverted the hyperalgesia induced by either carrageenin or bradykinin when injected concomitantly or 2.5 h after the phlogistic agents. Hemopressin administered systemically also reverted the hyperalgesia induced by carrageenin. Naloxone did not prevent the antinociceptive action of this peptide. These data suggest that hemopressin inhibits peripheral hyperalgesic responses by mechanisms independent of opioid receptor activation.

Neutrophils and the calcium-binding protein MRP-14 mediate carrageenan-induced antinociception in mice.

BACKGROUND: We have previously shown that the calcium-binding protein MRP-14 secreted by neutrophils mediates the antinociceptive response in an acute inflammatory model induced by the intraperitoneal injection of glycogen in mice. AIM: In an attempt to broaden the concept that neutrophils and MRP-14 controls inflammatory pain induced by different type of irritants, in the present study, after demonstrating that carrageenan (Cg) also induces atinociception in mice, we investigated the participation of both neutrophils and MRP-14 in the phenomenon. METHODS: Male Swiss mice were injected intraperitoneally with Cg and after different time intervals, the pattern of cell migration of the peritoneal exudate and the nociceptive response of animals submitted to the writhing test were evaluated. The participation of neutrophils and of the MRP-14 on the Cg effect was evaluated by systemic inoculation of monoclonal antibodies anti-granulocyte and anti-MRP-14. RESULTS: Our results demonstrate that the acute neutrophilic peritonitis evoked by Cg induced antinociception 2, 4 and 8 h after inoculation of the irritant. Monoclonal antibodies anti-granulocyte or anti-MRP-14 reverts the antinociceptive response only 2 and 8 h after Cg injection. The antibody anti-MRP-14 partially reverts the antinociception observed after 4 h of Cg injection while the anti-granulocyte antibody enhances this effect. This effect is reverted by simultaneous treatment of the animals with both antibodies. After 4 h of Cg injection in neutrophil-depleted mice a significant expression of the calcium-binding protein MRP-14 was detected in the cytoplasm of peritoneal macrophages. This suggests that the enhancement of the effect observed after treatment with the anti-neutrophil antibody may be due to secretion of MRP-14 by macrophages. It has also been demonstrated that endogenous opioids and glucocorticoids are not involved in the antinociception observed at the 4th hour after Cg injection. CONCLUSION: These data support the hypothesis that neutrophils and the calcium-binding protein MRP-14 are participants of the endogenous control of inflammatory pain in mice despite the model of acute inflammation used.

The C-terminus of murine S100A9 inhibits hyperalgesia and edema induced by jararhagin

The effect of a synthetic peptide (H92-G110) identical to the C-terminus of murine S100A9 (mS100A9p) was investigated on hyperalgesia and edema induced by either jararhagin or papain in the rat paw. mS100A9p not only reverted hyperalgesia and edema induced by jararhagin, but also the highest concentration induced antinociception. Hemorrhage induced by jararhagin and its hydrolytic activity were inhibited by mS100A9p. These data suggest that mS100A9p might block jararhagin-induced hyperalgesia and edema by inhibiting jararhagin catalytic activity, since papain-induced hyperalgesia and edema were not inhibited by mS100A9p.

Pro-inflammatory effects of the aqueous extract of Echinometra lucunter sea urchin spines

The sea urchin, Echinometra lucunter, can be found along the Western Central Atlantic shores. In Brazil, it is responsible by circa 50% of the accidents caused by marine animals. The symptoms usually surpass trauma and may be pathologically varied and last differently, ranging from spontaneous healing in a few days, to painful consequences lasting for weeks. In this work, we have mimicked the sea urchin accident by administering an aqueous extract of the spine into mice and rats and evaluated the pathophysiological developments. Our data clearly indicate that the sea urchin accident is indeed a pro-inflammatory event, triggered by toxins present in the spine that can cause edema and alteration in the leukocyte–endothelial interaction. Moreover, the spine extract was shown to exhibit a hyperalgesic effect. The extract is rich in proteins, as observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but also contains other molecules that can be analyzed by reversed phase high-performance liquid chromatography. Altogether, these effects corroborate that an E. lucunter encounter is an accident and not an incident, as frequently reported by the victims.

Involvement of proteinase-activated receptors 1 and 2 in spreading and phagocytosis by murine adherent peritoneal cells: Modulation by the C-terminal of S100A9 protein

Proteinase-activated receptors (PAR) are widely recognized for their modulatory properties in inflammatory and immune responses; however, their direct role on phagocyte effector functions remains unknown. S100A9, a protein secreted during inflammatory responses, deactivates activated peritoneal macrophages, and its C-terminal portion inhibits spreading and phagocytosis of adherent peritoneal cells. Herein, the effect of PAR1 and PAR2 agonists was investigated on spreading and phagocytosis by adherent peritoneal cells, as well as the ability of murine C-terminal of S100A9 peptide (mS100A9p) to modulate this effect. Adherent peritoneal cells obtained from mouse abdominal cavity were incubated with PAR1 and PAR2 agonists and spreading and phagocytosis of Candida albicans particles were evaluated. PAR1 agonists increased both the spreading and the phagocytic activity, but PAR2 agonists only increased the spreading index. mS100A9p reverted both the increased spreading and phagocytosis induced by PAR1 agonists, but no interference in the increased spreading induced by PAR2 agonists was noticed. The shorter homologue peptide to the C-terminal of mS100A9p, corresponding to the H(92)-E(97) region, also reverted the increased spreading and phagocytosis induced by PAR1 agonists. These findings show that proteinase-activated receptors have an important role for spreading and phagocytosis of adherent peritoneal cells, and that the peptide corresponding to the C-terminal of S100A9 protein is a remarkable candidate for use as a novel compound to modulate PAR1 function.

Effect of the C-terminus of murine S100A9 protein on experimental nociception

Calcium-binding protein S100A9 induces antinociception in mice evaluated by the writhing test. Similarly, a peptide identical to the C-terminus of murine S100A9 (mS100A9p) inhibits the hyperalgesia induced by jararhagin, a metalloprotease. Thus, we investigated the effect of mS100A9p on different models used to evaluate nociception. mS100A9p induced a dose-dependent inhibitory effect on the writhing test, and on mechanical hyperalgesia induced by carrageenan. mS100A9p inhibited thermal hyperalgesia induced by carrageenan. mS100A9p did not modify the nociceptive response in hot plate or tail-flick tests. These data demonstrate that the C-terminus of S100A9 protein interferes with control mechanisms of inflammatory pain.

The C-terminus of murine S100A9 protein inhibits hyperalgesia induced by the agonist peptide of protease-activated receptor 2 (PAR2).

S100A9 protein induces anti‐nociception in rodents, in different experimental models of inflammatory pain. Herein, we investigated the effects of a fragment of the C‐terminus of S100A9 (mS100A9p), on the hyperalgesia induced by serine proteases, through the activation of protease‐activated receptor‐2 (PAR2).