chronic-post-ischemia-pain-model-for-complex-regional-pain-syndrome

Chronic Post-Ischemia Pain Model for Complex Regional Pain Syndrome Type-I in Rats

Complex regional pain syndrome type-I (CRPS-I) is a neurological disease that causes severe pain among patients and remains an unresolved medical ...

Zhejiang Chinese Medical University

Oxidation products of the naturally occurring phospholipid 1-palmitoyl-2-arachidonoyl-sn-glycerol-3-phosphatidylcholine (PAPC), which are known as OxPAPC, accumulate in atherosclerotic lesions and at other sites of inflammation in conditions such as septic inflammation and acute lung injury to exert pro- or anti-inflammatory effects. It is currently unknown whether OxPAPC also contributes to inflammatory pain and peripheral neuronal excitability in these conditions. Here, we observed that OxPAPC dose-dependently and selectively activated human TRPA1 nociceptive ion channels expressed in HEK293 cells in vitro, without any effect on other TRP channels, including TRPV1, TRPV4 and TRPM8. OxPAPC agonist activity was dependent on essential cysteine and lysine residues within the N-terminus of the TRPA1 channel protein. OxPAPC activated calcium influx into a subset of mouse sensory neurons which were also sensitive to the TRPA1 agonist mustard oil. Neuronal OxPAPC responses were largely abolished in neurons isolated from TRPA1-deficient mice. Intraplantar injection of OxPAPC into the mouse hind paw induced acute pain and persistent mechanical hyperalgesia and this effect was attenuated by the TRPA1 inhibitor, HC-030031. More importantly, we found levels of OxPAPC to be significantly increased in inflamed tissue in a mouse model of chronic inflammatory pain, identified by the binding of an OxPAPC-specific antibody. These findings suggest that TRPA1 is a molecular target for OxPAPC and OxPAPC may contribute to chronic inflammatory pain through TRPA1 activation. Targeting against OxPAPC and TRPA1 signaling pathway may be promising in inflammatory pain treatment.

Oxidized Phospholipid OxPAPC Activates TRPA1 and Contributes to Chronic Inflammatory Pain in Mice.

Poison ivy-induced allergic contact dermatitis (ACD) is the most common environmental allergic condition in the United States. Case numbers of poison ivy ACD are increasing due to growing biomass and geographical expansion of poison ivy and increasing content of the allergen, urushiol, likely attributable to rising atmospheric CO Severe and treatment-resistant itch is the major complaint of affected patients. However, because of limited clinical data and poorly characterized models, the pruritic mechanisms in poison ivy ACD remain unknown. Here, we aim to identify the mechanisms of itch in a mouse model of poison ivy ACD by transcriptomics, neuronal imaging, and behavioral analysis. Using transcriptome microarray analysis, we identified IL-33 as a key cytokine up-regulated in the inflamed skin of urushiol-challenged mice. We further found that the IL-33 receptor, ST2, is expressed in small to medium-sized dorsal root ganglion (DRG) neurons, including neurons that innervate the skin. IL-33 induces Ca influx into a subset of DRG neurons through neuronal ST2. Neutralizing antibodies against IL-33 or ST2 reduced scratching behavior and skin inflammation in urushiol-challenged mice. Injection of IL-33 into urushiol-challenged skin rapidly exacerbated itch-related scratching via ST2, in a histamine-independent manner. Targeted silencing of neuronal ST2 expression by intrathecal ST2 siRNA delivery significantly attenuated pruritic responses caused by urushiol-induced ACD. These results indicate that IL-33/ST2 signaling is functionally present in primary sensory neurons and contributes to pruritus in poison ivy ACD. Blocking IL-33/ST2 signaling may represent a therapeutic approach to ameliorate itch and skin inflammation related to poison ivy ACD.

IL-33/ST2 signaling excites sensory neurons and mediates itch response in a mouse model of poison ivy contact allergy.

Hydroquinone (HQ) is one of the most frequently used and effective skin-lightening products to treat skin hyperpigmentation disorders, including postinflammatory hyperpigmentation, melasma and solar lentigines. HQ is also widely used in cosmetic products for skin whitening. However, HQ treatment can evoke substantial skin irritation, a side effect that remains poorly understood. Here we demonstrate that HQ is an activator of the peripheral irritant receptor transient receptor potential (TRP) cation channel member A1 (TRPA1). HQ failed to activate TRPV1, TRPV4 or TRPM8. HQ-induced TRPA1 activation was dependent on essential redox-sensitive cysteine and lysine residues within N-terminus of channel protein. HQ elicited Ca influx in a subpopulation of mouse sensory neurons sensitive to the TRPA1 agonist, mustard oil. HQ-induced neuronal responses were significantly reduced by TRPA1 inhibitors, and reduced in neurons isolated from Trpa1-deficient mice. In mice, intraplantar injection of HQ at clinically relevant concentrations elicited both acute pain and persistent mechanical hyperalgesia which were almost completely abolished by TRPA1 inhibitors. These findings identify TRPA1 as a molecular target for HQ and provide insights into the mechanism of HQ-induced skin irritation. These findings also suggest that selective TRPA1 antagonists may be useful to counteract HQ-induced skin irritation.

Involvement of Transient Receptor Potential Cation Channel Member A1 activation in the irritation and pain response elicited by skin-lightening reagent hydroquinone.

A murine monoclonal antibody (MAb-1) specific for GM3 has been generated by immunizing β3Gn-T5 knockout mice with purified GM3 ganglioside. The binding specificity of MAb-1 (IgG subclass) was established by an enzyme-linked immunosorbent assay (ELISA) and FACS and the antibody showed high binding specificity with GM3. Cell viability assay showed that MAb-1 significantly suppressed cell growth. Immunohistochemistry analysis revealed that MAb-1 was strongly expressed in human ovarian cancer tissues, whereas it was hardly expressed in normal tissues. Finally, antibody-dependent cellular cytotoxicity (ADCC) activities were determined by measuring lactate dehydrogenase (LDH) releasing assay and the results showed high ADCC activities in two representative ovarian cancer cell lines (OVHM and ID8). All of these data indicate that MAb-1 may be potentially used as a therapeutic antibody against ovarian cancers in clinical trials.

Generation and characterization of a IgG monoclonal antibody specific for GM3 (NeuGc) ganglioside by immunizing β3Gn-T5 knockout mice.

Acute gout arthritis is one of the most painful inflammatory conditions. Treatments for gout pain are limited to colchicine, nonsteroidal anti-inflammatory drugs, and corticosteroids, which oftentimes result in severe adverse effects. Electroacupuncture (EA) has been proved to be effective in relieving many inflammatory pain conditions with few side effects. Here, we aim to investigate the therapeutic potentials of EA on pain and inflammation of a rat model of acute gout arthritis and underlying mechanisms. We found that 2/100 Hz EA produced the most robust analgesic effect on mechanical hyperalgesia of acute gout arthritis rat model compared with 2 and 100 Hz. EA produced similar analgesic effect compared with indomethacin. 2/100 Hz EA also significantly alleviates the ongoing pain behavior, thermal hyperalgesia, and ankle edema. Locally applied  and -opioid receptor antagonists but not adenosine A receptor antagonist significantly abolished the analgesic effect of EA. Locally applied  and -opioid receptor agonists produced significant antiallodynia on acute gout arthritis rats, mimicking EA. Furthermore, 2/100 Hz EA upregulated -endorphin expression in inflamed ankle skin tissue. Our results demonstrated, for the first time, that EA can be used for relieving acute gout arthritis with effect dependent on peripheral opioid system and comparable with the one obtained with indomethacin.

Electroacupuncture Alleviates Pain Responses and Inflammation in a Rat Model of Acute Gout Arthritis.

Endothelin-1 (ET-1) is a potent endogenous vasoconstrictor that has been widely known as a pain mediator involved in various pain states. Evidence indicates that ET-1 sensitizes transient receptor potential cation channel, subfamily A, member 1 (TRPA1) in vivo. But the molecular mechanisms still remain unknown. We aim to explore whether ET-1 sensitizes TRPA1 in primary sensory neurons and the molecular mechanisms. Ca imaging, immunostaining, electrophysiology, animal behavioral assay combined with pharmacological experiments were performed. ET-1 sensitized TRPA1-mediated Ca responses in human embryonic kidney (HEK)293 cells as well as in cultured native mouse dorsal root ganglion (DRG) neurons. ET-1 also sensitized TRPA1 channel currents. ET-1 sensitized TRPA1 activated by endogenous agonist HO. ET receptor (ETR) colocalized with TRPA1 in DRG neurons. ET-1-induced TRPA1 sensitization in vivo was mediated via ETR and protein kinase A (PKA) pathway in HEK293 cells and DRG neurons. Pharmacological blocking of ETR, PKA, and TRPA1 significantly attenuated ET-1-induced mechanical hyperalgesia in mice. Our results suggest that TRPA1 acts as a molecular target for ET-1, and sensitization of TRPA1 through ETR-PKA pathway contributes to ET-1-induced mechanical hyperalgesia. Pharmacological targeting of TRPA1 and ETR-PKA pathway may provide effective strategies to alleviate pain conditions associated with ET-1.

ETR and protein kinase A pathway mediate ET-1 sensitization of TRPA1 channel: A molecular mechanism of ET-1-induced mechanical hyperalgesia.

Complex regional pain syndrome type 1 (CRPS-I) is a debilitating pain condition that significantly affects life quality of patients. It remains a clinically challenging condition and the mechanisms of CRPS-I have not been fully elucidated. Here, we investigated the involvement of TRPV1, a non-selective cation channel important for integrating various painful stimuli, in an animal model of CRPS-I. A rat model of chronic post-ischemia pain (CPIP) was established to mimic CRPS-I. TRPV1 expression was significantly increased in hind paw tissue and small to medium-sized dorsal root ganglion (DRG) neurons of CPIP rats. CPIP rats showed increased TRPV1 current density and capsaicin responding rate in small-sized nociceptive DRG neurons. Local pharmacological blockage of TRPV1 with the specific antagonist AMG9810, at a dosage that does not produce hyperthermia or affect thermal perception or locomotor activity, effectively attenuated thermal and mechanical hypersensitivity in bilateral hind paws of CPIP rats and reduced the hyperexcitability of DRG neurons induced by CPIP. CPIP rats showed bilateral spinal astrocyte and microglia activations, which were significantly attenuated by AMG9810 treatment. These findings identified an important role of TRPV1 in mediating thermal and mechanical hypersensitivity in a CRPS-I animal model and further suggest local pharmacological blocking TRPV1 may represent an effective approach to ameliorate CRPS-I.

TRPV1 Channel Contributes to the Behavioral Hypersensitivity in a Rat Model of Complex Regional Pain Syndrome Type 1.

 Complex regional pain syndrome type-I (CRPS-I) is a progressive and devastating pain condition, which remains clinically challenging. The mechanisms of CRPS-I still remain largely unknown. We aim to identify transcriptome profiles of genes relevant to pain mechanisms and major pathways involved in CRPS-I.  A rat model of chronic post-ischemia pain (CPIP) was established to mimic CRPS-I. RNA-sequencing (RNA-Seq) was used to profile transcriptome of L4-6 dorsal root ganglia (DRGs) of a rat model of CRPS-I.  CPIP model rats developed persistent mechanical/thermal hyperalgesia in ipsilateral hind paw. RNA-Seq identified a total of 295 differentially expressed genes (DEGs), including 195 up- and 100 downregulated, in ipsilateral DRGs of CPIP rats compared with sham rats. The expression of several representative genes was confirmed by qPCR. Functional analysis of DEGs revealed that the most significant enriched biological processes of upregulated genes include response to lipopolysaccharide, inflammatory response and cytokine activity, which are all important mechanisms mediating pain. We further screened DEGs implicated in pain progress, genes enriched in small- to medium-sized sensory neurons and enriched in TRPV1-lineage nociceptors. By comparing our dataset with other published datasets of neuropathic or inflammatory pain models, we identified a core set of genes and pathways that extensively participate in CPIP and other neuropathic pain states.  Our study identified transcriptome gene changes in DRGs of an animal model of CRPS-I and could provide insights into identifying promising genes or pathways that can be potentially targeted to ameliorate CRPS-I.

Transcriptome profiling of dorsal root ganglia in a rat model of complex regional pain syndrome type-I reveals potential mechanisms involved in pain.

In the United States, poison ivy exposure is the most common naturally occurring allergen to cause allergic contact dermatitis (ACD). The immune and pruritic mechanisms associated with poison ivy ACD remain largely unexplored. Here, we compared skin whole transcriptomes and itch mediator levels in mouse ACD models induced by the poison ivy allergen, urushiol, and the synthetic allergen, oxazolone. The urushiol model produced a Th2-biased immune response and scratching behavior, resembling findings in poison ivy patients. Urushiol-challenged skin contained elevated levels of the cytokine thymic stromal lymphopoietin (TSLP), a T-cell regulator and itch mediator, and pruritogenic serotonin (5-HT) and endothelin (ET-1), but not substance P (SP) or histamine. The oxazolone model generated a mixed Th1/Th2 response associated with increased levels of substance P, 5-HT, ET-1, but not TSLP or histamine. Injections of a TSLP monoclonal neutralizing antibody, serotonergic or endothelin inhibitors, but not SP inhibitors or antihistamines, reduced scratching behaviors in urushiol-challenged mice. Our findings suggest that the mouse urushiol model may serve as a translational model of human poison ivy ACD study. Inhibiting signaling by TSLP and other cytokines may represent alternatives to the standard steroid/antihistamine regimen for steroid-resistant or -intolerant patients and in exaggerated systemic responses to poison ivy.

Transcriptome profiling reveals Th2 bias and identifies endogenous itch mediators in poison ivy contact dermatitis.

Gout arthritis, which is provoked by monosodium urate (MSU) crystal accumulation in the joint and periarticular tissues, induces severe pain and affects life quality of the patients. Eucalyptol (1, 8-cineol), the principal component in the essential oils of eucalyptus leaves, has been known to possess anti-inflammatory and analgesic properties. We aimed to examine the therapeutic effects of eucalyptol on gout arthritis and related mechanisms.

Article	Year
Oxidized Phospholipid OxPAPC Activates TRPA1 and Contributes to Chronic Inflammatory Pain in Mice. PloS one\| PubMed ID: 27812120	2016
IL-33/ST2 signaling excites sensory neurons and mediates itch response in a mouse model of poison ivy contact allergy. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 27821781	2016
Involvement of Transient Receptor Potential Cation Channel Member A1 activation in the irritation and pain response elicited by skin-lightening reagent hydroquinone. Scientific reports\| PubMed ID: 28790335	2017
Generation and characterization of a IgG monoclonal antibody specific for GM3 (NeuGc) ganglioside by immunizing β3Gn-T5 knockout mice. Scientific reports\| PubMed ID: 29416099	2018
Electroacupuncture Alleviates Pain Responses and Inflammation in a Rat Model of Acute Gout Arthritis. Evidence-based complementary and alternative medicine : eCAM\| PubMed ID: 29743920	2018
ETR and protein kinase A pathway mediate ET-1 sensitization of TRPA1 channel: A molecular mechanism of ET-1-induced mechanical hyperalgesia. Molecular pain\| PubMed ID: 30990108	2019
TRPV1 Channel Contributes to the Behavioral Hypersensitivity in a Rat Model of Complex Regional Pain Syndrome Type 1. Frontiers in pharmacology\| PubMed ID: 31105572	2019
Transcriptome profiling of dorsal root ganglia in a rat model of complex regional pain syndrome type-I reveals potential mechanisms involved in pain. Journal of pain research\| PubMed ID: 31114302	2019
Transcriptome profiling reveals Th2 bias and identifies endogenous itch mediators in poison ivy contact dermatitis. JCI insight\| PubMed ID: 31184997	2019
Eucalyptol Alleviates Inflammation and Pain Responses in a Mouse Model of Gout Arthritis. British journal of pharmacology\| PubMed ID: 31883118	2019

Yan Tai

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