Endogenous control of inflammatory visceral pain by T cell-derived opioids in IL-10-deficient mice.

BACKGROUND: The opioid-mediated analgesic activity of mucosal CD4+ T lymphocytes in colitis has been reported in immunocompetent mice so far. Here, we investigated whether CD4+ T lymphocytes alleviate from inflammation-induced abdominal pain in mice with defective immune regulation. METHODS: Endogenous control of visceral pain by opioids locally produced in inflamed mucosa was assessed in IL-10-deficient mice. KEY RESULTS: CD4+ T lymphocytes but not F4/80+ macrophages isolated from the lamina propria of IL-10-deficient mice with colitis express enkephalin-containing opioid peptides as assessed by cytofluorometry. Colitis in IL-10-/- mice was not associated with abdominal pain. Intraperitoneal injection of naloxone-methiodide, a peripheral opioid receptor antagonist, induced abdominal hypersensitivity in IL-10-/- mice with colitis. CONCLUSION AND INFERENCES: Opioid-mediated analgesic activity of mucosal T lymphocytes remains operating in IL-10-/- mice with impaired immune regulation. The data suggest that endogenous T cell-derived opioids might reduce inflammation-induced abdominal pain in inflammatory bowel diseases associated with homozygous "loss of function mutations" in interleukin-10.

Opioids and the immune system - friend or foe.

Systemically administered opioids are among the most powerful analgesics for treating severe pain. Several negative side effects (respiratory depression, addiction, nausea and confusion) and the risk of opioid-induced hyperalgesia accompany opioid administration. One other side effect is the potential of opioids to suppress the immune response and thereby to increase the vulnerability to infections. The link between opioids and immunosuppression has been investigated both in vitro and in vivo as well as in patients. However, the results are inconsistent: Exogenous opioids such as morphine and fentanyl have been found to impair the function of macrophages, natural killer cells and T-cells and to weaken the gut barrier in vitro and in animal studies. In epidemiological studies, high doses and the initiation of opioid therapy for non-malignant pain have been correlated with a higher risk of infectious diseases such as pneumonia. However clear randomized controlled studies are missing. Furthermore, immune cells including neutrophils, macrophages and T-cells have been shown to secrete endogenous opioid peptides, which then bind to peripheral opioid receptors to relieve inflammatory and neuropathic pain. In addition to cytokines, hormones and bacterial products, the release of opioid peptides is stimulated by the application of exogenous opioids. In summary, there is a reciprocal interaction between the immune system and endogenous as well as exogenous opioids. Further to the existing epidemiological studies, controlled clinical studies are needed in the future to elucidate the role of the opioid-immune system interaction in patients and to determine its clinical relevance. LINKED ARTICLES: This article is part of a themed section on Emerging Areas of Opioid Pharmacology. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.14/issuetoc.

Opioid and Opiate Immunoregulatory Processes.

The discovery of the ability of the nervous system to communicate through "public" circuits with other systems of the body is attributed to Ernst and Berta Scharrer, who described the neurosecretory process in 1928. Indeed, the immune system has been identified as another important neuroendocrine target tissue. Opioid peptides are involved in this communication (i.e., neuroimmune) and with that of autoimmunoregulation (communication between immunocytes). The significance of opioid neuropeptide involvement with the immune system is ascertained from the presence of novel δ, µ., and κ receptors on inflammatory cells that result in modulation of cellular activity after activation, as well as the presence of specific enzymatic degradation and regulation processes. In contrast to the relatively uniform antinociceptive action of opiate and opioid signal molecules in neural tissues, the presence of naturally occurring morphine in plasma and a novel µ3 opiate-specific receptor on inflammatory cells adds to the growing knowledge that opioid and opiate signal molecules may have antagonistic actions in select tissues. In examining various disorders (e.g., human immunodeficiency virus, substance abuse, parasitism, and the diffuse inflammatory response associated with surgery) evidence has also been found for the involvement of opiate/opioid signaling in prominent mechanisms. In addition, the presence of similar mechanisms in man and organisms 500 million years divergent in evolution bespeaks the importance of this family of signal molecules. The present review provides an overview of recent advances in the field of opiate and opioid immunoregulatory processes and speculates as to their significance in diverse biological systems.

Nociceptin/orphanin FQ (N/OFQ) modulates immunopathology and airway hyperresponsiveness representing a novel target for the treatment of asthma.

BACKGROUND AND PURPOSE: There is evidence supporting a role for the nociceptin/orphanin FQ (N/OFQ; NOP) receptor and its endogenous ligand N/OFQ in the modulation of neurogenic inflammation, airway tone and calibre. We hypothesized that NOP receptor activation has beneficial effects upon asthma immunopathology and airway hyperresponsiveness. Therefore, the expression and function of N/OFQ and the NOP receptor were examined in healthy and asthmatic human airway tissues. The concept was further addressed in an animal model of allergic asthma. EXPERIMENTAL APPROACH: NOP receptor expression was investigated by quantitative real-time PCR. Sputum N/OFQ was determined by RIA. N/OFQ function was tested using several assays including proliferation, migration, collagen gel contraction and wound healing. The effects of N/OFQ administration in vivo were studied in ovalbumin (OVA)-sensitized and challenged mice. KEY RESULTS: NOP receptors were expressed on a wide range of human and mouse immune and airway cells. Eosinophils expressed N/OFQ-precursor mRNA and their number correlated with N/OFQ concentration. N/OFQ was found in human sputum and increased in asthma. Additionally, in asthmatic human lungs N/OFQ immunoreactivity was elevated. NOP receptor activation inhibited migration of immunocytes and increased wound healing in airway structural cells. Furthermore, N/OFQ relaxed spasmogen-stimulated gel contraction. Remarkably, these findings were mirrored in OVA-mice where N/OFQ treatment before or during sensitization substantially reduced airway constriction and immunocyte trafficking to the lung, in particular eosinophils. N/OFQ also reduced inflammatory mediators and mucin production. CONCLUSIONS AND IMPLICATIONS: We demonstrated a novel dual airway immunomodulator/bronchodilator role for N/OFQ and suggest targeting this system as an innovative treatment for asthma.

Endogenous opiates and behavior: 2013.

This paper is the thirty-sixth consecutive installment of the annual review of research concerning the endogenous opioid system. It summarizes papers published during 2013 that studied the behavioral effects of molecular, pharmacological and genetic manipulation of opioid peptides, opioid receptors, opioid agonists and opioid antagonists. The particular topics that continue to be covered include the molecular-biochemical effects and neurochemical localization studies of endogenous opioids and their receptors related to behavior, and the roles of these opioid peptides and receptors in pain and analgesia; stress and social status; tolerance and dependence; learning and memory; eating and drinking; alcohol and drugs of abuse; sexual activity and hormones, pregnancy, development and endocrinology; mental illness and mood; seizures and neurologic disorders; electrical-related activity and neurophysiology; general activity and locomotion; gastrointestinal, renal and hepatic functions; cardiovascular responses; respiration and thermoregulation; and immunological responses.

Endogenous regulation of visceral pain via production of opioids by colitogenic CD4(+) T cells in mice.

BACKGROUND & AIMS: A dysregulated response of CD4(+) T cells against the microbiota contributes to the development of inflammatory bowel disease. Effector CD4(+) T cells, generated in response to microbe-derived antigens, can reduce somatic inflammatory pain through the local release of opioids. We investigated whether colitogenic CD4(+) T cells that accumulate in the inflamed colon also produce opioids and are able to counteract inflammation-induced visceral pain in mice. METHODS: Colitis was induced via transfer of naive CD4(+)CD45RB(high) T cells to immune-deficient mice or by administration of dextran sulfate sodium. Mice without colitis were used as controls. Samples of colon tissue were collected, and production of opioids by immune cells from inflamed intestine was assessed by quantitative polymerase chain reaction and cytofluorometry analyses. The role of intestinal opioid tone in inflammation-induced visceral hypersensitivity was assessed by colorectal distention. RESULTS: In mice with T cell- or dextran sulfate sodium-induced colitis, colitogenic CD4(+) T cells (T-helper 1 and Th17 cells) accumulated in the inflamed intestine and expressed a high level of endogenous opioids. In contrast, macrophages and epithelial cells did not express opioids; opioid synthesis in the myenteric plexus was not altered on induction of inflammation. In mice with colitis, the local release of opioids by colitogenic CD4(+) T cells led to significant reduction of inflammation-associated visceral hypersensitivity. CONCLUSIONS: In mice, colitogenic Th1 and Th17 cells promote intestinal inflammation and colonic tissue damage but have simultaneous opioid-mediated analgesic activity, thereby reducing abdominal pain.

Immunomodulatory effects of endogenous and synthetic peptides activating opioid receptors.

The main role of endogenous opioid peptides is the modulation of pain. Opioid peptides exert their analgesic activity by binding to the opioid receptors distributed widely in the central nervous system (CNS). However, opioid receptors are also found on tissues and organs outside the CNS, including the cells of the immune system, indicating that opioids are capable of exerting additional effects in periphery. Morphine, which is a gold standard in the treatment of chronic pain, is well-known for its immunosuppressive effects. Much less is known about the immunomodulatory effects exerted by endogenous (enkephalins, endorphins, dynorphins and endomorphins) and synthetic peptides activating opioid receptors. In this review we tried to summarize opioid peptide-mediated modulation of immune cell functions which can be stimulatory as well as inhibitory.

[Antibodies to endogenous bioregulators and their association with age and sex in chronic pain syndrome].

Authors studied changes in the levels of antibodies to endogenous bioregulators (Ab) to Β-endorphin, orphanin, serotonin, dopamine and angiotensin in 36 healthy people and 109 patients with dorsopathy with chronic pain syndrome. The association of these immunological indicators with age and sex was found. It has been concluded that the levels of Ab to endogenous bioregulators may be considered as a marker of algic system pathology that does not depend on age and is sex-related.

Nociceptin/orphanin FQ receptor activation decreases the airway hyperresponsiveness induced by allergen in sensitized mice.

Several studies suggest that the N/OFQ (nociceptin/orphanin FQ)-NOP (N/OFQ peptide) receptor pathway is involved in airway physiology. We previously demonstrated a modulation of the endogenous N/OFQ levels in allergen-sensitized mice. Here, we investigated the effects of NOP receptor activation in allergen sensitization using a murine model of allergen-induced airway hyperresponsiveness (AHR). BALB/c mice were intraperitoneally treated with the NOP receptor agonist UFP-112, either during the sensitization phase (30 min before ovalbumin administration) or at the end of sensitization process (15 min before bronchopulmonary reactivity evaluation). At day 21 from the first allergen exposure, bronchopulmonary reactivity and total and differential cell count in bronchoalveolar lavage fluid were evaluated. In a separate set of experiments cell proliferation in lymphocytes, cytokine levels, IgE serum levels, and the effect of UFP-112 on IL-13-induced AHR were evaluated. Pretreatment with UFP-112, during the sensitization phase, caused a significant reduction in allergen-induced AHR and total cell lung infiltration. No effect on allergen-induced AHR was observed when the treatment was performed at the end of sensitization process, on tissues harvested from OVA-sensitized mice and on IL-13-induced AHR. The in vitro proliferative response of lymphocytes was significantly reduced by pretreatment during the sensitization phase with UFP-112. This effect was paralleled by a significant modulation of cytokine secretion in pulmonary tissues and lymphocytes. In conclusion, we demonstrated a role for the NOP receptor and N/OFQ pathway in the AHR induced by allergen, probably through a modulation of the immune response that triggers the development of AHR that involves pro- and anti-inflammatory cytokines.

The novel roles of neutrophils via opioid peptides: regulation of the estrous cycle and pain.

Neutrophils are excreted into the vaginal vault at metestrus during the estrous cycle, and this phenomenon has long been used to determine the phase of the estrous cycle. A much smaller number of neutrophils are also detected in the uterus and the ovary. Recently, we provided several lines of evidence supporting the notion that neutrophils infiltrate into the ovary to regulate the estrous cycle by opioid peptides. Upon inflammation, on the other hand, neutrophils infiltrate into the site of infection to suppress pain by opioid peptides. Thus, opioid peptides are key molecules by which neutrophils play a novel role in regulation of the pain and estrous cycle. In both cases, opioid peptides appear to be secreted by neutrophils stimulated with chemokines, such as MIP-2 and KC in mouse, corticotropin-releasing hormone and IL-1.

Endogenous opiates and behavior: 2011.

This paper is the thirty-fourth consecutive installment of the annual review of research concerning the endogenous opioid system. It summarizes papers published during 2011 that studied the behavioral effects of molecular, pharmacological and genetic manipulation of opioid peptides, opioid receptors, opioid agonists and opioid antagonists. The particular topics that continue to be covered include the molecular-biochemical effects and neurochemical localization studies of endogenous opioids and their receptors related to behavior (Section 2), and the roles of these opioid peptides and receptors in pain and analgesia (Section 3); stress and social status (Section 4); tolerance and dependence (Section 5); learning and memory (Section 6); eating and drinking (Section 7); alcohol and drugs of abuse (Section 8); sexual activity and hormones, pregnancy, development and endocrinology (Section 9); mental illness and mood (Section 10); seizures and neurologic disorders (Section 11); electrical-related activity and neurophysiology (Section 12); general activity and locomotion (Section 13); gastrointestinal, renal and hepatic functions (Section 14); cardiovascular responses (Section 15); respiration (Section 16); and immunological responses (Section 17).

Pain inhibition by blocking leukocytic and neuronal opioid peptidases in peripheral inflamed tissue.

Inflammatory pain can be controlled by endogenous opioid peptides. Here we blocked the degradation of opioids in peripheral injured tissue to locally augment this physiological system. In rats with hindpaw inflammation, inhibitors of aminopeptidase N (APN; bestatin) or neutral endopeptidase (NEP; thiorphan), and a dual inhibitor, NH(2)-CH-Ph-P(O)(OH)CH(2)-CH-CH(2)Ph(p-Ph)-CONH-CH-CH(3)-COOH (P8B), were applied to injured paws. Combined bestatin (1.25-5 mg)/thiorphan (0.2-0.8 mg) or P8B (0.0625-1 mg) alone elevated mechanical nociceptive thresholds to 307 and 227% of vehicle-treated controls, respectively. This analgesia was abolished by antibodies to methionine-enkephalin, leucine-enkephalin, and dynorphin A 1-17, by peripherally restricted and by selective µ-, δ-, and κ-opioid receptor antagonists. Flow cytometry and photospectrometry revealed expression and metabolic activity of APN and NEP on macrophages, granulocytes, and sciatic nerves from inflamed tissue. Radioimmunoassays showed that inhibition of leukocytic APN and NEP by bestatin (5-500 µM)/thiorphan (1-100 µM) combinations or by P8B (1-100 µM) prevented the degradation of enkephalins. Blockade of neuronal peptidases by bestatin (0.5-10 mM)/thiorphan (0.1-5 mM) or by P8B (0.1-10 mM) additionally hindered dynorphin A 1-17 catabolism. Thus, leukocytes and peripheral nerves are important sources of APN and NEP in inflamed tissue, and their blockade promotes peripheral opioid analgesia.

Preparation and evaluation of an immunoaffinity sorbent for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis-mass spectrometry.

In this study, we explored a procedure for the preparation of an immunoaffinity (IA) sorbent for the analysis of opioid peptides by on-line immunoaffinity solid-phase extraction capillary electrophoresis-mass spectrometry (IA-SPE-CE-MS). We followed a site-specific antibody immobilization approach based on the covalent attachment of the oxidized antibodies through their carbohydrate moieties to hydrazide silica particles, using a polyclonal antibody against Endomorphin 1 and 2 (End1 and End2). The main features of the IA sorbent were studied, such as the amount of hydrazide groups and antibodies attached onto oxidized diol silica particles. Once the procedure was optimized, standard solutions of End1 and End2 were used in order to establish the IA-SPE-CE-MS methodology. Acceptable repeatability, reproducibility and linearity range values were obtained for the proposed methodology. The limits of detection (LODs) of 1 ng mL(-1) were approximately 100-fold better than those obtained by CE-MS. Selectivity of the IA sorbent was good but some cross-reactivity against Dynorphin A (1-7) was observed when a mixture of several opioid peptides was analyzed. Human plasma samples spiked with End1 and End2 were also analyzed and both peptides could be detected down to 100 ng mL(-1).

Influence of endogenous opioid systems on T lymphocytes as assessed by the knockout of mu, delta and kappa opioid receptors.

Here, we evaluated the influence of endogenous opioid activation on immune responses by examining consequences of all three opioid receptor gene (mu, delta and kappa) inactivation. In triple-opioid receptor knockout mice, splenocytes and thymocytes numbers, lymphocyte subsets as well as proliferation and cytokines induced by in vitro stimulation of T lymphocytes were measured. Compared with wild-type mice, similar lymphocyte distribution in thymus and spleen as well as comparable T lymphocyte proliferation were observed, while lower levels of IL-2 and IFNγ as well as higher levels of IL-4 and IL-10 were found in triple-opioid receptor knockout mice. Together, our results indicate a shift from TH1 to TH2 cytokines in triple-opioid receptor knockout animals, suggesting that global endogenous opioid tone drives T lymphocytes toward a TH1 profile under non-pathological conditions.

Endogenous opioid peptides in regulation of innate immunity cell functions.

Endogenous opioid peptides comprise a group of bioregulatory factors involved in regulation of functional activity of various physiological systems of an organism. One of most important functions of endogenous opioids is their involvement in the interaction between cells of the nervous and immune systems. Summary data on the effects of opioid peptides on regulation of functions of innate immunity cells are presented.

Endogenous opioid-mediated analgesia is dependent on adaptive T cell response in mice.

Pain is an inherent component of inflammation often accompanying immune response. A large spectrum of molecules released within the inflamed tissue induces pain by stimulating primary afferent neurons in situ. Activity of primary sensitive fibers can be counteracted by local opioid release by leukocytes. In this study, we investigated the endogenous regulation of CFA-induced inflammatory pain in the context of adaptive T cell immune response. The nociceptive response to mechanical stimuli was studied using von Frey filaments in mice immunized with OVA in CFA. The nociceptive response of nude versus wild-type mice was dramatically increased, demonstrating T cell deficiency associated with increased pain sensitivity. Based on adoptive transfer experiments of OVA-specific CD4(+) T lymphocytes into nude mice, we show that Ag-specific activated, but not resting T lymphocytes are responsible for the spontaneous relief of inflammation-induced pain following Ag challenge. The analgesia was dependent on opioid release by Ag-primed CD4(+) T lymphocytes at the inflammatory site. Indeed, T cell-mediated analgesia was inhibited by local injection of an opioid receptor antagonist, unable to cross the blood-brain barrier. Notably, we found opioid precursor mRNA to be >7-fold increased in Ag-specific activated CD4(+) T lymphocytes, as compared with resting T lymphocytes in vivo. Taken together, our results show that CD4(+) T lymphocytes acquire antinociceptive effector properties when specifically primed by Ag and point out analgesia as a property linked to the effector phase of adaptive T cell response.

Endogenous µ-opioid peptides modulate immune response towards malignant melanoma.

Opioids exert major effects not only in the central nervous system but also in immune responses. We investigated the effects of µ-opioid peptides, secreted by tumor cells, on anti-tumor immune responses. For this purpose, tumor growth was studied in wild-type and µ-opioid receptor-deficient (MOR-/-) mice injected with B16 melanoma cells. The ability of these cells to produce opioids was studied by Western blots in vitro. Finally, biopsy material from human melanomas was investigated by immunohistochemistry for ß endorphin expression. Injection of B16 melanoma cells, producing endogenous ß endorphin, in the flank of MOR-/- mice revealed a profound reduction in tumor growth, paralleled by a significantly higher infiltration of immune cells into the tumors, when compared to tumor growth after injection of B16 melanoma cells into wild-type mice. Opioids present in B16 cell supernatant significantly reduced the proliferation of normal but not MOR-/- leucocytes. Immunohistochemical analyses of biopsies from human melanoma tissues showed a positive correlation between expression of ß endorphin and tumor progression. Our data provide evidence that µ-opioid peptides may play a major role in cancer progression by modulating immune response. This finding may have implications for the future optimization of immunointerventions for cancer.

Mechanisms of peripheral immune-cell-mediated analgesia in inflammation: clinical and therapeutic implications.

Peripheral mechanisms of endogenous pain control are significant. In peripheral inflamed tissue, an interaction between immune-cell-derived opioids and opioid receptors localized on sensory nerve terminals results in potent, clinically measurable analgesia. Opioid peptides and the mRNA encoding their precursor proteins are present in immune cells. These cells 'home' preferentially to injured tissue, where they secrete opioids to reduce pain. Investigation of the mechanisms underlying the migration of opioid-containing immune cells to inflamed tissue is an active area of research, with recent data demonstrating the importance of cell adhesion molecules in leukocyte adhesion to both the endothelium in vascular transmigration and to neurons within peripheral inflamed tissue. This review summarizes the physiological mechanisms and clinical significance of this unique endogenous peripheral analgesic pathway and discusses therapeutic implications for the development of novel targeted peripheral analgesics.

Opioid receptors and opioid peptide-producing leukocytes in inflammatory pain--basic and therapeutic aspects.

This review summarizes recent findings on neuro-immune mechanisms underlying opioid-mediated inhibition of pain. The focus is on events occurring in peripheral injured tissues that lead to the sensitization and excitation of primary afferent neurons, and on the modulation of such mechanisms by immune cell-derived opioid peptides. Primary afferent neurons are of particular interest from a therapeutic perspective because they are the initial generators of impulses relaying nociceptive information towards the spinal cord and the brain. Thus, if one finds ways to inhibit the sensitization and/or excitation of peripheral sensory neurons, subsequent central events such as wind-up, sensitization and plasticity may be prevented. This is in part achieved by endogenously released immune cell-derived opioid peptides within inflamed tissue. In addition, exogenous opioid receptor ligands that selectively modulate primary afferent function and do not cross the blood-brain barrier, avoid centrally mediated untoward side effects of conventional analgesics (e.g., opioids, anticonvulsants). This article discusses peripheral opioid receptors and their signaling pathways, opioid peptide-producing/secreting inflammatory cells and arising therapeutic perspectives.