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Joint pain is one of the most debilitating symptoms of rheumatoid arthritis (RA) and patients frequently rate improvements in pain management as their priority. RA is hallmarked by the presence of anti-modified protein autoantibodies (AMPA) against post-translationally modified citrullinated, carbamylated and acetylated proteins. It has been suggested that autoantibody-mediated processes represent distinct mechanisms contributing to pain in RA. In this study, we investigated the pronociceptive properties of monoclonal AMPA 1325:01B09 (B09 mAb) derived from the plasma cell of an RA patient. We found that B09 mAb induces pain-like behavior in mice that is not associated with any visual, histological or transcriptional signs of inflammation in the joints, and not alleviated by non-steroidal anti-inflammatory drugs (NSAIDs). Instead, we found that B09 mAb is retained in dorsal root ganglia (DRG) and alters the expression of several satellite glia cell (SGC), neuron and macrophage-related factors in DRGs. Using mice that lack activating FcγRs, we uncovered that FcγRs are critical for the development of B09-induced pain-like behavior, and partially drive the transcriptional changes in the DRGs. Finally, we observed that B09 mAb binds SGC in vitro and in combination with external stimuli like ATP enhances transcriptional changes and protein release of pronociceptive factors from SGCs. We propose that certain RA antibodies bind epitopes in the DRG, here on SGCs, form immune complexes and activate resident macrophages via FcγR cross-linking. Our work supports the growing notion that autoantibodies can alter nociceptor signaling via mechanisms that are at large independent of local inflammatory processes in the joint.
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Artrite Reumatoide , Autoanticorpos , Animais , Camundongos , Receptores de IgG , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico , DorRESUMO
Type-1 diabetes mellitus (T1DM) is a chronic condition characterized by long-term hyperglycemia that results in several complications such as painful peripheral neuropathy, bone deterioration, and increased risk of bone fractures. Lithium, a first-line therapy for bipolar disorder, has become an attractive agent for attenuating peripheral neuropathy and menopause-induced bone loss. Therefore, our aim was to determine the effect of chronic lithium treatment on mechanical hypersensitivity and trabecular bone loss induced by T1DM in mice. T1DM was induced in male C57BL/6J mice by intraperitoneal injection of streptozotocin (STZ, 50 mg/kg/day, for 5 consecutive days). 12 weeks after T1DM-induction, mice received a daily intraperitoneal injection of vehicle, 30 or 60 mg/kg lithium (as LiCl) for 6 weeks. Throughout the treatment period, blood glucose levels and mechanical sensitivity were evaluated every 2 weeks. After lithium treatment, the femur and L5 vertebra were harvested for microcomputed tomography (microCT) analysis. T1DM mice showed significant hyperglycemia, mechanical hypersensitivity, and significant trabecular bone loss as compared with the control group. Chronic lithium treatment did not revert the hindpaw mechanical hypersensitivity nor hyperglycemia associated to T1DM induced by STZ. In contrast, microCT analysis revealed that lithium reverted, in a dose-dependent manner, the loss of trabecular bone associated to T1DM induced by STZ at both the distal femur and L5 vertebra. Lithium treatment by itself did not affect any trabecular bone parameter in non-diabetic mice.
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Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1 , Hiperglicemia , Animais , Glicemia , Osso Esponjoso/diagnóstico por imagem , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/tratamento farmacológico , Feminino , Hiperglicemia/induzido quimicamente , Lítio/farmacologia , Compostos de Lítio/efeitos adversos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estreptozocina , Microtomografia por Raio-XRESUMO
OBJECTIVE: An interesting and so far unexplained feature of chronic pain in autoimmune disease is the frequent disconnect between pain and inflammation. This is illustrated well in rheumatoid arthritis (RA) where pain in joints (arthralgia) may precede joint inflammation and persist even after successful anti-inflammatory treatment. In the present study, we have addressed the possibility that autoantibodies against citrullinated proteins (ACPA), present in RA, may be directly responsible for the induction of pain, independent of inflammation. METHODS: Antibodies purified from human patients with RA, healthy donors and murinised monoclonal ACPA were injected into mice. Pain-like behaviour was monitored for up to 28â days, and tissues were analysed for signs of pathology. Mouse osteoclasts were cultured and stimulated with antibodies, and supernatants analysed for release of factors. Mice were treated with CXCR1/2 (interleukin (IL) 8 receptor) antagonist reparixin. RESULTS: Mice injected with either human or murinised ACPA developed long-lasting pronounced pain-like behaviour in the absence of inflammation, while non-ACPA IgG from patients with RA or control monoclonal IgG were without pronociceptive effect. This effect was coupled to ACPA-mediated activation of osteoclasts and release of the nociceptive chemokine CXCL1 (analogue to human IL-8). ACPA-induced pain-like behaviour was reversed with reparixin. CONCLUSIONS: The data suggest that CXCL1/IL-8, released from osteoclasts in an autoantibody-dependent manner, produces pain by activating sensory neurons. The identification of this new pain pathway may open new avenues for pain treatment in RA and also in other painful diseases associated with autoantibody production and/or osteoclast activation.
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Artralgia/imunologia , Autoanticorpos/imunologia , Quimiocina CXCL1/imunologia , Citrulina/imunologia , Interleucina-8/imunologia , Nociceptividade/fisiologia , Osteoclastos/imunologia , Animais , Autoanticorpos/farmacologia , Comportamento Animal/efeitos dos fármacos , Estudos de Casos e Controles , Quimiocina CXCL1/efeitos dos fármacos , Quimiocinas , Inflamação , Interleucina-8/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Nociceptividade/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Receptores de Interleucina-8/antagonistas & inibidores , Sulfonamidas/farmacologiaRESUMO
OBJECTIVES: Rheumatoid arthritis (RA)-specific anti-citrullinated protein/peptide antibodies (ACPAs) appear before disease onset and are associated with bone destruction. We aimed to dissect the role of ACPAs in osteoclast (OC) activation and to identify key cellular mediators in this process. METHODS: Polyclonal ACPA were isolated from the synovial fluid (SF) and peripheral blood of patients with RA. Monoclonal ACPAs were isolated from single SF B-cells of patients with RA. OCs were developed from blood cell precursors with or without ACPAs. We analysed expression of citrullinated targets and peptidylarginine deiminases (PAD) enzymes by immunohistochemistry and cell supernatants by cytometric bead array. The effect of an anti-interleukin (IL)-8 neutralising antibody and a pan-PAD inhibitor was tested in the OC cultures. Monoclonal ACPAs were injected into mice and bone structure was analysed by micro-CT before and after CXCR1/2 blocking with reparixin. RESULTS: Protein citrullination by PADs is essential for OC differentiation. Polyclonal ACPAs enhance OC differentiation through a PAD-dependent IL-8-mediated autocrine loop that is completely abolished by IL-8 neutralisation. Some, but not all, human monoclonal ACPAs derived from single SF B-cells of patients with RA and exhibiting distinct epitope specificities promote OC differentiation in cell cultures. Transfer of the monoclonal ACPAs into mice induced bone loss that was completely reversed by the IL-8 antagonist reparixin. CONCLUSIONS: We provide novel insights into the key role of citrullination and PAD enzymes during OC differentiation and ACPA-induced OC activation. Our findings suggest that IL8-dependent OC activation may constitute an early event in the initiation of the joint specific inflammation in ACPA-positive RA.
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Artrite Reumatoide/imunologia , Autoanticorpos/imunologia , Reabsorção Óssea/imunologia , Osso e Ossos/imunologia , Citrulina/imunologia , Hidrolases/metabolismo , Interleucina-8/imunologia , Osteoclastos/imunologia , Animais , Linfócitos B/imunologia , Reabsorção Óssea/diagnóstico por imagem , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/efeitos dos fármacos , Técnicas de Cultura de Células , Quimiocinas/imunologia , Feminino , Humanos , Hidrolases/antagonistas & inibidores , Imuno-Histoquímica , Interleucina-8/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Osteoclastos/efeitos dos fármacos , Desiminases de Arginina em Proteínas , Receptores de Interleucina-8/antagonistas & inibidores , Sulfonamidas/farmacologia , Líquido Sinovial , Microtomografia por Raio-XRESUMO
A rapid, sensitive and simple high-performance liquid chromatographic assay with ultraviolet detection was developed for the quantification of levofloxacin in microsamples (100 µL) of human plasma. The extraction procedure included a protein precipitation technique and a short chromatographic running time (4.5 min). Analyses were carried out on a Symmetry C18 column using a mixture of acetonitrile and 0.01 m potassium dihydrogen aqueous solution (pH 3.4; 14:86 v/v) as mobile phase. The method provided specificity and was linear (r ≥ 0.9992) over the concentration range 0.1-12 µg/mL. The average absolute recovery was 93.59%. The intra- and inter-day coefficients of variation were <6%. Additionally, levofloxacin was stable in all evaluations. The usefulness of this method was demonstrated in a pharmacokinetic study of levofloxacin in healthy adult volunteers. The present method offers two main advantages: (a) the use of microsamples reduces the total volume of blood to be collected from patients; and (b) it provides a good cost-effectiveness ratio. It is concluded that the method is rapid, simple, sensitive, economical and suitable for the determination of levofloxacin in human plasma using a small volume of sample.
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Cromatografia Líquida de Alta Pressão/métodos , Levofloxacino/sangue , Levofloxacino/farmacocinética , Adulto , Cromatografia Líquida de Alta Pressão/instrumentação , Humanos , Masculino , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
This study was performed to evaluate whether early, middle, or late treatment of zoledronate, an approved bisphosphonate that blocks bone resorption, can reduce nociceptive behaviors in a mouse arthritis model. Arthritis was produced by repeated intra-articular knee injections of complete Freund's adjuvant (CFA). A dose-response curve with zoledronate (3, 30, 100, and 300 µg/kg, i.p., day 4 to day 25, twice weekly for 3 weeks) was performed, and the most effective dose of zoledronate (100 µg/kg, i.p.) was initially administered at different times of disease progression: day 4 (early), day 15 (middle), or day 21 (late) and continued until day 25 after the first CFA injection. Flinching of the injected extremity (spontaneous nociceptive behavior), vertical rearings and horizontal activity (functional outcomes), and knee edema were assessed. Zoledronate improved both functional outcomes and reduced flinching behavior. At day 25, the effect of zoledronate on flinching behavior and vertical rearings was greater in magnitude when it was given early or middle rather than late in the treatment regimen. Chronic zoledronate did not reduce knee edema in CFA-injected mice nor functional outcomes in naïve mice by itself. These results suggest that zoledronate may have a positive effect on arthritis-induced nociception and functional disabilities.
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Artrite Experimental/complicações , Artrite Experimental/tratamento farmacológico , Difosfonatos/administração & dosagem , Difosfonatos/uso terapêutico , Imidazóis/administração & dosagem , Imidazóis/uso terapêutico , Atividade Motora/efeitos dos fármacos , Dor Nociceptiva/complicações , Dor Nociceptiva/tratamento farmacológico , Animais , Artrite Experimental/induzido quimicamente , Conservadores da Densidade Óssea/administração & dosagem , Conservadores da Densidade Óssea/uso terapêutico , Difosfonatos/farmacologia , Relação Dose-Resposta a Droga , Esquema de Medicação , Edema/complicações , Edema/tratamento farmacológico , Adjuvante de Freund , Imidazóis/farmacologia , Masculino , Camundongos , Medição da Dor/efeitos dos fármacos , Ácido ZoledrônicoRESUMO
The relationship between blood levels of ketoprofen and its anti-hyperalgesic effects was examined in rat using the carrageenan-evoked thermal hyperalgesia model. Female adult Wistar rats were injected with carrageenan into the plantar surface of the right hind paw. Immediately after, rats were administered with ketoprofen po and hindpaw withdrawal latency measured and micro-whole blood samples were obtained over six hours via a cannula inserted in the caudal artery. Ketoprofen levels were measured by HPLC. Ketoprofen concentration increased in a dose-dependent manner and was reflected in dose-dependent anti-hyperalgesic effect. The pharmacokinetic and pharmacodynamic parameters expressed as mean ± s.e.m. following administration of 1, 3.2, and 10 mg/kg ketoprofen were: Cmax 1.27 ± 0.08, 3.44 ± 0.20 and 11.76 ± 0.81 µg/mL; AUClast 4.16 ± 0.17, 11.63 ± 0.65 and 28.15 ± 1.32 µg h/mL; and Emax observed (AUCE ): 65.41 ± 7.79, 92.06 ± 6.46 and 98.42 ± 7.53%. A direct relationship between blood concentrations and the anti-hyperalgesic effect of ketoprofen followed a maximum effect model equation. The results indicate that the anti-hyperalgesic effect of ketoprofen in the carrageenan pain model can be predicted by the pharmacokinetic properties of ketoprofen.
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Analgésicos/sangue , Analgésicos/uso terapêutico , Hiperalgesia/sangue , Hiperalgesia/tratamento farmacológico , Cetoprofeno/sangue , Cetoprofeno/uso terapêutico , Analgésicos/farmacocinética , Animais , Carragenina , Feminino , Temperatura Alta , Hiperalgesia/induzido quimicamente , Cetoprofeno/farmacocinética , Ratos WistarRESUMO
BACKGROUND: Prolonged fetal exposure to hyperglycemia may increase the risk of developing abnormal glucose metabolism and type-2 diabetes during childhood, adolescence, and adulthood; however, the mechanisms by which gestational diabetes mellitus (GDM) predisposes offspring to metabolic disorders remain unknown. AIM: To quantify the nerve axons, macrophages, and vasculature in the pancreas from adult offspring born from mouse dams with GDM. METHODS: GDM was induced by i.p. administration of streptozotocin (STZ) in ICR mouse dams. At 12 wk old, fasting blood glucose levels were determined in offspring. At 15 wk old, female offspring born from dams with and without GDM were sacrificed and pancreata were processed for immunohistochemistry. We quantified the density of sensory [calcitonin gene-related peptide (CGRP)] and tyrosine hydroxylase (TH) axons, blood vessels (endomucin), and macro-phages (CD68) in the splenic pancreas using confocal microscopy. RESULTS: Offspring mice born from STZ-treated dams had similar body weight and blood glucose values compared to offspring born from vehicle-treated dams. However, the density of CGRP+ and TH+ axons, endomucin+ blood vessels, and CD68+ macrophages in the exocrine pancreas was significantly greater in offspring from mothers with GDM vs control offspring. Likewise, the microvasculature in the islets was significantly greater, but not the number of macrophages within the islets of offspring born from dams with GDM compared to control mice. CONCLUSION: GDM induces neuronal, vascular, and inflammatory changes in the pancreas of adult progeny, which may partially explain the higher propensity for offspring of mothers with GDM to develop metabolic diseases.
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Worldwide over 12 million people were diagnosed with cancer (excluding non-melanoma skin cancer) and 8 million individuals died from cancer in 2008. Recent data indicate that 75-90% of patients with advanced stage diseases or metastatic cancer will experience significant cancer pain. Bone cancer pain is common in patients with advanced breast, prostate, and lung cancer as these tumors have a marked affinity to metastasize to bone. Once tumors metastasize to bone, they are a major cause of morbidity and mortality as the tumor induces significant skeletal remodeling, fractures, pain and anemia; all of which reduce the functional status, quality of life and survival of the patient. Currently, the factors that drive cancer pain are poorly understood, however, several recently introduced models of bone cancer pain that mirror the human condition, are providing insight into the mechanisms that drive bone cancer pain and guiding the development of novel therapies to treat the cancer pain. Several of these therapies have recently been approved by the FDA to treat bone cancer pain (bisphosphonates, denosumab) and others are currently being evaluated in human clinical trials (tanezumab). These new mechanism-based therapies are enlarging the repertoire of modalities available to treat bone cancer pain and improving the quality of life and functional status of patients with bone cancer.
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Neoplasias Ósseas/complicações , Dor/tratamento farmacológico , Dor/etiologia , Animais , Ensaios Clínicos como Assunto , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , HumanosRESUMO
Gastric injury is mainly described by inflammation of the gastric epithelium. Recently, our group of work demonstrated that Prosthechea karwinskii leaves extract induces both an in vitro antioxidative action and an in vivo gastroprotective effect in a rat. However, the molecules involved in the gastroprotective action by Prosthechea karwinskii are not known. Thus, the aim of this study is to determine whether Prosthechea karwinskii extract modifies anti-inflammatory and antioxidative biomarkers in an in vivo rat model of indomethacin-induced gastric injury. Rats were orally administered with indomethacin and Prosthechea karwinskii leaf extract. Our results suggest that the gastroprotective effect of Prosthechea karwinskii leaf extract is related to the reduction in leukocyte infiltration and antioxidative action in a model of indomethacin-induced gastric injury. Further studies are warranted to investigate the role of the compounds identified in the gastroprotective action of Prosthechea karwinskii leaves extract.
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Antiulcerosos , Úlcera Gástrica , Ratos , Animais , Indometacina/efeitos adversos , Óxido Nítrico/farmacologia , Leucotrieno B4/efeitos adversos , Úlcera Gástrica/induzido quimicamente , Úlcera Gástrica/tratamento farmacológico , Úlcera Gástrica/prevenção & controle , Antiulcerosos/farmacologia , Extratos Vegetais/uso terapêutico , Mucosa Gástrica , Antioxidantes/farmacologia , Folhas de PlantaRESUMO
While the long-term complications of gestational diabetes mellitus (GDM) in the cardiovascular, endocrine, and central nervous systems from offspring have been widely studied, less is known about the long-term outcomes of GDM on the peripheral nervous system. Thus, here we assessed the mechanical sensitivity and density of nerve fibers of the hind paw from middle-aged offspring born from dams with GDM. GDM was induced by the intraperitoneal administration of streptozotocin (STZ) in mouse dams. Mechanical sensitivity in male and female offspring was bi-weekly evaluated from week 18 to week 40 of age. At 40 weeks old, offspring were sacrificed and glabrous hind paw skin was processed for immunohistochemistry to determine the density of intraepidermal CGRP and PGP9.5 positive nerve fibers. Offspring mice born from STZ-treated dams had significantly greater mechanical sensitivity from 18 to 40 weeks of age compared to offspring born from vehicle-treated dams (control group). The density of intraepidermal CGRP+ and PGP9.5+ nerve fibers were significantly lower in the hind paw skin of female but not male offspring, born from STZ-treated dams versus the control group. These results suggest that GDM has long-term sex-dependent complications on the nociceptive system. Further studies are necessary to elucidate the mechanisms underlying the GDM-induced long-term consequences.
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Diabetes Mellitus Experimental , Diabetes Gestacional , Gravidez , Humanos , Camundongos , Feminino , Animais , Diabetes Gestacional/induzido quimicamente , Estreptozocina , Peptídeo Relacionado com Gene de Calcitonina , Fibras NervosasRESUMO
Cancer-induced bone pain (CIBP) is the most common and devastating symptom of bone metastatic cancer that substantially disrupts patients' quality of life. Currently, there are few effective analgesic treatments for CIBP other than opioids which come with severe side effects. In order to better understand the factors and mechanisms responsible for CIBP it is essential to have clinically relevant animal models that mirror pain-related symptoms and disease progression observed in patients with bone metastatic cancer. In the current study, we characterize a syngeneic mouse model of prostate cancer induced bone pain. We transfected a prostate cancer cell line (RM1) with green fluorescent protein (GFP) and luciferase reporters in order to visualize tumor growth longitudinally in vivo and to assess the relationship between sensory neurons and tumor cells within the bone microenvironment. Following intra-femoral injection of the RM1 prostate cancer cell line into male C57BL/6 mice, we observed a progressive increase in spontaneous guarding of the inoculated limb between 12 and 21 days post inoculation in tumor bearing compared to sham operated mice. Daily running wheel performance was evaluated as a measure of functional impairment and potentially movement evoked pain. We observed a progressive reduction in the distance traveled and percentage of time at optimal velocity between 12 and 21 days post inoculation in tumor bearing compared to sham operated mice. We utilized histological, radiographic and µCT analysis to examine tumor induced bone remodeling and observed osteolytic lesions as well as extra-periosteal aberrant bone formation in the tumor bearing femur, similar to clinical findings in patients with bone metastatic prostate cancer. Within the tumor bearing femur, we observed reorganization of blood vessels, macrophage and nerve fibers within the intramedullary space and periosteum adjacent to tumor cells. Tumor bearing mice displayed significant increases in the injury marker ATF3 and upregulation of the neuropeptides SP and CGRP in the ipsilateral DRG as well as increased measures of central sensitization and glial activation in the ipsilateral spinal cord. This immunocompetent mouse model will be useful when combined with cell type selective transgenic mice to examine tumor, immune cell and sensory neuron interactions in the bone microenvironment and their role in pain and disease progression associated with bone metastatic prostate cancer.
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Many types of solid tumors metastasize to the bone, where it causes significant morbidity and mortality in patients with advanced disease. Bone metastases are not only incurable but also affect bone health which impairs patients' quality of life. In order to understand the mechanisms and develop effective treatments for bone-metastatic disease, it is first necessary to develop animal models that permit the assessment of tumor growth in the bone and progressive structural changes of the bone simultaneously. Longitudinal analysis of bone tumor progression is generally performed by bioluminescent imaging; however, this method is not able to assess progressive structural changes of the bone. Here, we describe a simple method for assessment of bone lesions using a scoring system that takes into account disease burden and bone destruction using longitudinal radiographs.
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Neoplasias Ósseas , Qualidade de Vida , Animais , Neoplasias Ósseas/patologia , Osso e Ossos/patologia , Modelos Animais de Doenças , Humanos , Camundongos , RadiografiaRESUMO
ABSTRACT: Several bone conditions, eg, bone cancer, osteoporosis, and rheumatoid arthritis (RA), are associated with a risk of developing persistent pain. Increased osteoclast activity is often the hallmark of these bony pathologies and not only leads to bone remodeling but is also a source of pronociceptive factors that sensitize the bone-innervating nociceptors. Although historically bone loss in RA has been believed to be a consequence of inflammation, both bone erosion and pain can occur years before the symptom onset. Here, we have addressed the disconnection between inflammation, pain, and bone erosion by using a combination of 2 monoclonal antibodies isolated from B cells of patients with RA. We have found that mice injected with B02/B09 monoclonal antibodies (mAbs) developed a long-lasting mechanical hypersensitivity that was accompanied by bone erosion in the absence of joint edema or synovitis. Intriguingly, we have noted a lack of analgesic effect of naproxen and a moderate elevation of few inflammatory factors in the ankle joints suggesting that B02/B09-induced pain-like behavior does not depend on inflammatory processes. By contrast, we found that inhibiting osteoclast activity and acid-sensing ion channel 3 signaling prevented the development of B02/B09-mediated mechanical hypersensitivity. Moreover, we have identified secretory phospholipase A2 and lysophosphatidylcholine 16:0 as critical components of B02/B09-induced pain-like behavior and shown that treatment with a secretory phospholipase A2 inhibitor reversed B02/B09-induced mechanical hypersensitivity and bone erosion. Taken together, our study suggests a potential link between bone erosion and pain in a state of subclinical inflammation and offers a step forward in understanding the mechanisms of bone pain in diseases such as RA.
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Canais Iônicos Sensíveis a Ácido , Artrite Reumatoide , Osteoclastos , Dor , Canais Iônicos Sensíveis a Ácido/metabolismo , Animais , Anticorpos Monoclonais/farmacologia , Artrite Reumatoide/complicações , Artrite Reumatoide/tratamento farmacológico , Inflamação/complicações , Camundongos , Osteoclastos/patologia , Dor/patologiaRESUMO
PURPOSE: Gestational diabetes mellitus (GDM) induces cardiovascular and metabolic disturbances in offspring. However, the effects of GDM in pain processing in offspring and whether male and female offspring are equally affected is not well known. Thus, we determined: i) whether GDM in mice affects offspring hindpaw mechanical sensitivity, capsaicin-induced spontaneous pain-like behaviors, and epidermal nerve fiber density (ENFD); and ii) whether there is sexual dimorphism in these parameters in offspring from GDM dams. METHODS: GDM was induced in pregnant ICR mice via i.p. streptozotocin (STZ). Then, glucose levels from dams and offspring were determined. Male and female offspring 2-3 months of age were evaluated for: a) baseline mechanical sensitivity of the hind paw by using von Frey filaments; b) number of flinches and time spent guarding induced by intraplantar capsaicin (0.1%); and c) density of PGP-9.5 and CGRP axons in the epidermis from the hind paw glabrous skin. RESULTS: Prepartum levels of glucose in STZ-treated dams were significantly increased compared to vehicle-treated dams; however, GDM or vehicle offspring displayed normal and similar blood glucose levels. Male and female GDM offspring showed significantly greater mechanical sensitivity and capsaicin-induced pain behaviors compared to vehicle offspring. Male GDM offspring displayed a slightly more intense nociceptive phenotype in the capsaicin test. PGP-9.5 and CGRP ENFD in hind paw glabrous skin were greater in male and female GDM offspring versus their controls. Sexual dimorphism was generally not observed in GDM offspring in most of the studied parameters. CONCLUSION: These results suggest GDM induced greater pain-like behaviors in adult offspring regardless of sex along with an increased ENFD of PGP-9.5 and CGRP in the hind paw glabrous skin. We show that GDM peripheral neuropathy differs from diabetic peripheral neuropathy acquired in adulthood and set the foundation to further study this in human babies exposed to GDM.
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The adult K/BxN transgenic mouse develops spontaneous autoimmune arthritis with joint remodeling and profound bone loss. We report that both males and females display a severe sustained tactile allodynia which is reduced by gabapentin but not the potent cyclooxygenase inhibitor ketorolac. In dorsal horn, males and females show increased GFAP+ astrocytic cells; however, only males demonstrate an increase in Iba1+ microglia. In dorsal root ganglia (DRG), there is an increase in CGRP+, TH+, and Iba1+ (macrophage) labeling, but no increase in ATF3+ cells. At the ankle there is increased CGRP+, TH+, and GAP-43+ fiber synovial innervation. Thus, based on the changes in dorsal horn, DRG and peripheral innervation, we suggest that the adult K/BxN transgenic arthritic mice display a neuropathic phenotype, an assertion consistent with the analgesic pharmacology seen in this animal. These results indicate the relevance of this model to our understanding of the nociceptive processing which underlies the chronic pain state that evolves secondary to persistent joint inflammation.
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Artrite Experimental/complicações , Gânglios Espinais/patologia , Hiperalgesia/patologia , Articulação do Joelho/patologia , Tecido Nervoso/patologia , Dor Nociceptiva/patologia , Analgésicos/farmacologia , Animais , Artrite Experimental/fisiopatologia , Feminino , Gabapentina/farmacologia , Gânglios Espinais/efeitos dos fármacos , Hiperalgesia/tratamento farmacológico , Hiperalgesia/etiologia , Articulação do Joelho/efeitos dos fármacos , Masculino , Camundongos , Camundongos Transgênicos , Tecido Nervoso/efeitos dos fármacos , Neuralgia/patologia , Dor Nociceptiva/tratamento farmacológico , Dor Nociceptiva/etiologia , FenótipoRESUMO
Bone pain is a serious and debilitating symptom of multiple myeloma (MM) that impairs the quality of life of patients. The underlying mechanisms of the pain are unknown and understudied, and there is a need for immunocompetent preclinical models of myeloma-induced bone pain. The aim of this study was to provide the first in-depth behavioral characterization of an immunocompetent mouse model of MM presenting the clinical disease features: osteolytic bone disease and bone pain. We hypothesized that a widely used syngeneic model of MM, established by systemic inoculation of green fluorescent protein-tagged myeloma cells (5TGM1-GFP) in immunocompetent C57Bl/KaLwRijHsd (BKAL) mice, would present pain-related behaviors. Disease phenotype was confirmed by splenomegaly, high serum paraprotein, and tumor infiltration in the bone marrow of the hind limbs; however, myeloma-bearing mice did not present pain-related behaviors or substantial bone disease. Thus, we investigated an alternative model in which 5TGM1-GFP cells were directly inoculated into the intrafemoral medullary cavity. This localized myeloma model presented the hallmarks of the disease, including high serum paraprotein, tumor growth, and osteolytic bone lesions. Compared with control mice, myeloma-bearing mice presented myeloma-induced pain-related behaviors, a phenotype that was reversed by systemic morphine treatment. Micro-computed tomography analyses of the myeloma-inoculated femurs showed bone disease in cortical and trabecular bone. Repeated systemic bisphosphonate treatment induced an amelioration of the nociceptive phenotype, but did not completely reverse it. Furthermore, intrafemorally injected mice presented a profound denervation of the myeloma-bearing bones, a previously unknown feature of the disease. This study reports the intrafemoral inoculation of 5TGM1-GFP cells as a robust immunocompetent model of myeloma-induced bone pain, with consistent bone loss. Moreover, the data suggest that myeloma-induced bone pain is caused by a combinatorial mechanism including osteolysis and bone marrow denervation. © 2019 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.
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Background: Painful distal symmetrical polyneuropathy (DPN) is a frequent complication of type-2 diabetes mellitus (T2DM) that commonly presents as neuropathic pain and loss of skin nerve fibers. However, there are limited therapies to effectively treat DPN and many of the current animal models of T2DM-induced DPN do not appear to mirror the human disease. Thus, we validated a DPN mouse model induced by a cafeteria-style diet plus low-doses of streptozotocin (STZ). Methods: Female C57BL/6J mice were fed either standard (STD) diet or obesogenic cafeteria (CAF) diet for 32 weeks, starting at 8 weeks old. Eight weeks after starting diets, CAF or STD mice received either four low-doses of STZ or vehicle. Changes in body weight, blood glucose and insulin levels, as well as oral glucose- and insulin-tolerance tests (OGTT and ITT) were determined. The development of mechanical hypersensitivity of the hindpaws was determined using von Frey filaments. Moreover, the effect of the most common neuropathic pain drugs was evaluated on T2DM-induced mechanical allodynia. Finally, the density of PGP -9.5+ (a pan-neuronal marker) axons in the epidermis from the hindpaw glabrous skin was quantified. Results: At 22-24 weeks after STZ injections, CAF + STZ mice had significantly higher glucose and insulin levels compared to CAF + VEH, STD + STZ, and STD + VEH mice, and developed glucose tolerance and insulin resistance. Skin mechanical sensitivity was detected as early as 12 weeks post-STZ injections and it was significantly attenuated by intraperitoneal acute treatment with amitriptyline, gabapentin, tramadol, duloxetine, or carbamazepine but not by diclofenac. The density of PGP-9.5+ nerve fibers was reduced in CAF + STZ mice compared to other groups. Conclusion: This reverse translational study provides a painful DPN mouse model which may help in developing a better understanding of the factors that generate and maintain neuropathic pain and denervation of skin under T2DM and to identify mechanism-based new treatments.
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Rheumatoid arthritis-associated joint pain is frequently observed independent of disease activity, suggesting unidentified pain mechanisms. We demonstrate that antibodies binding to cartilage, specific for collagen type II (CII) or cartilage oligomeric matrix protein (COMP), elicit mechanical hypersensitivity in mice, uncoupled from visual, histological and molecular indications of inflammation. Cartilage antibody-induced pain-like behavior does not depend on complement activation or joint inflammation, but instead on tissue antigen recognition and local immune complex (IC) formation. smFISH and IHC suggest that neuronal Fcgr1 and Fcgr2b mRNA are transported to peripheral ends of primary afferents. CII-ICs directly activate cultured WT but not FcRγ chain-deficient DRG neurons. In line with this observation, CII-IC does not induce mechanical hypersensitivity in FcRγ chain-deficient mice. Furthermore, injection of CII antibodies does not generate pain-like behavior in FcRγ chain-deficient mice or mice lacking activating FcγRs in neurons. In summary, this study defines functional coupling between autoantibodies and pain transmission that may facilitate the development of new disease-relevant pain therapeutics.
Assuntos
Anticorpos Monoclonais/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Artralgia/imunologia , Artrite Reumatoide/imunologia , Autoanticorpos/imunologia , Cartilagem/imunologia , Neurônios/metabolismo , Animais , Anticorpos Monoclonais/uso terapêutico , Artralgia/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Autoanticorpos/uso terapêutico , Comportamento Animal/efeitos dos fármacos , Proteína de Matriz Oligomérica de Cartilagem/imunologia , Colágeno Tipo II/imunologia , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Transgênicos , Receptores de IgG/deficiência , Receptores de IgG/genéticaRESUMO
Paclitaxel (Taxol) is a frontline antineoplastic agent used to treat a variety of solid tumors including breast, ovarian, or lung cancer. The major dose limiting side effect of paclitaxel is a peripheral sensory neuropathy that can last days to a lifetime. To begin to understand the cellular events that contribute to this neuropathy, we examined a marker of cell injury/regeneration (activating transcription factor 3; ATF3), macrophage hyperplasia/hypertrophy; satellite cell hypertrophy in the dorsal root ganglia (DRG) and sciatic nerve as well as astrocyte and microglial activation within the spinal cord at 1, 4, 6 and 10 days following intravenous infusion of therapeutically relevant doses of paclitaxel. At day 1 post-infusion, there was an up-regulation of ATF3 in a subpopulation of large and small DRG neurons and this up-regulation was present through day 10. In contrast, hypertrophy of DRG satellite cells, hypertrophy and hyperplasia of CD68(+) macrophages in the DRG and sciatic nerve, ATF3 expression in S100beta(+) Schwann cells and increased expression of the microglial marker (CD11b) and the astrocyte marker glial fibrillary acidic protein (GFAP) in the spinal cord were not observed until day 6 post-infusion. The present results demonstrate that using the time points and markers examined, DRG neurons show the first sign of injury which is followed days later by other neuropathological changes in the DRG, peripheral nerve and dorsal horn of the spinal cord. Understanding the cellular changes that generate and maintain this neuropathy may allow the development of mechanism-based therapies to attenuate or block this frequently painful and debilitating condition.