Hypersensitivity Model Research Articles

Syngeneic red blood cell-induced extracellular vesicles suppress delayed-type hypersensitivity to self-antigens in mice.

At present, the role of autologous cells as antigen carriers inducing immune tolerance is appreciated. Accordingly, intravenous administration of haptenated syngeneic mouse red blood cells (sMRBC) leads to hapten-specific suppression of contact hypersensitivity (CHS) in mice, mediated by light chain-coated extracellular vesicles (EVs). Subsequent studies suggested that mice intravenously administered with sMRBC alone may also generate regulatory EVs, revealing the possible self-tolerogenic potential of autologous erythrocytes. The current study investigated the immune effects induced by mere intravenous administration of a high dose of sMRBC in mice. The self-tolerogenic potential of EVs was determined in a newly developed mouse model of delayed-type hypersensitivity (DTH) to sMRBC. The effects of EV's action on DTH effector cells were evaluated cytometrically. The suppressive activity of EVs, after coating with anti-hapten antibody light chains, was assessed in hapten-induced CHS in wild-type or miRNA-150-/- mice. Intravenous administration of sMRBC led to the generation of CD9+CD81+ EVs that suppressed sMRBC-induced DTH in a miRNA-150-dependent manner. Furthermore, the treatment of DTH effector cells with sMRBC-induced EVs decreased the activation of T cells but enhanced their apoptosis. Finally, EVs coated with antibody light chains inhibited hapten-induced CHS. The current study describes a newly discovered mechanism of self-tolerance induced by the intravenous delivery of a high dose of sMRBC that is mediated by EVs in a miRNA-150-dependent manner. This mechanism implies the concept of naturally occurring immune tolerance, presumably activated by overloading of the organism with altered self-antigens.

CPNE7 Induces Biological Dentin Sealing in a Dentin Hypersensitivity Model

Dentin hypersensitivity commonly occurs due to opened dentinal tubules for many reasons. In our previous study, copine 7 (CPNE7) could induce dentin formation for an indirect pulp-capping model in vivo. This study aims to investigate the formation of tertiary dentin when CPNE7 is applied to intentionally exposed dentin with nothing over it in vivo, whether it affects microleakage of the teeth, and the penetration ability of CPNE7 molecules through dentinal tubules in vitro. Cervical dentin areas of 6 maxillary incisors of 5 beagles were exposed to a class V–like lesion, and 1 side of 3 maxillary incisors was adapted with recombinant CPNE7 protein for 5 min as the experimental group. The other side was the control group, and there was no treatment of ethylenediaminetetraacetic acid (EDTA) and CPNE7 after preparation. The defects were exposed without any restorations, and all beagles were sacrificed after 4 wk. The fluid penetration of exposed dentin areas was investigated by a microleakage-testing device and confocal laser scanning microscope. Tertiary dentin formation was confirmed with histological scanning electronic microscopic analysis. Tertiary dentin formation reduces dentinal fluid flow due to occluded tubules or discontinuity with primary or secondary dentin. The in vivo hypersensitivity model with the anterior teeth of beagle dogs showed newly formed tertiary dentin at the dentin-pulp boundary in recombinant CPNE7–treated teeth when compared with the untreated control group in histologic analysis. Scanning electronic microscopic analysis revealed occluded sites with mineral deposition of intratubular dentin. In the permeability test, the mean microleakage value of the CPNE7-treated group was significantly lower than that of the control group (P < 0.05). The tubular penetration of rhodamine B–combined CPNE7 was confirmed under confocal laser scanning microscope. CPNE7 induces formation of tertiary dentin through shallowly exposed dentinal tubules, which decreases dentin permeability.

Central Sensitization, N-methyl-D-aspartate Receptors, and Human Experimental Pain Models: Bridging the Gap between Target Discovery and Drug Development.

Central Sensitization, N-methyl-D-aspartate Receptors, and Human Experimental Pain Models: Bridging the Gap between Target Discovery and Drug Development.

Oral administration of lipoteichoic acid from Lactobacillus rhamnosus GG overcomes UVB‐induced immunosuppression and impairs skin tumor growth in mice

There is increasing evidence of the relevant connection and regulation between the gut and skin immune axis. In fact, oral administration of lipoteichoic acid (LTA) from Lactobacillus rhamnosus GG (LGG) prevents the development of UV-induced skin tumors in chronically exposed mice. Here we aim to evaluate whether this LTA is able to revert UV-induced immunosuppression as a mechanism involved in its anti-tumor effect and whether it has an immunotherapeutic effect against cutaneous squamous cell carcinoma. Using a mouse model of contact hypersensitivity, we demonstrate that LTA overcomes UV-induced skin immunosuppression. This effect was in part achieved by modulating the phenotype of lymph node resident dendritic cells (DC) and the homing of skin migratory DC. Importantly, oral LTA reduced significantly the growth of established skin tumors once UV radiation was discontinued, demonstrating that it has a therapeutic, besides the already demonstrated preventive antitumor effect. The data presented here strongly indicates that oral administration of LTA represents a promising immunotherapeutic approach for different conditions in which the skin immune system is compromised.

The Role of Conformational Dynamics in Abacavir-Induced Hypersensitivity Syndrome

Abacavir is an antiretroviral drug used to reduce human immunodeficiency virus (HIV) replication and decrease the risk of developing acquired immune deficiency syndrome (AIDS). However, its therapeutic value is diminished by the fact that it is associated with drug hypersensitivity reactions in up to 8% of treated patients. This hypersensitivity is strongly associated with patients carrying human leukocyte antigen (HLA)-B*57:01, but not patients carrying closely related alleles. Abacavir’s specificity to HLA-B*57:01 is attributed to its binding site within the peptide-binding cleft and subsequent influence of the repertoire of peptides that can bind HLA-B*57:01. To further our understanding of abacavir-induced hypersensitivity we used molecular dynamics (MD) to analyze the dynamics of three different peptides bound to HLA-B*57:01 in the presence and absence of abacavir or abacavir analogues. We found that abacavir and associated peptides bind to HLA-B*57:01 in a highly diverse range of conformations that are not apparent from static crystallographic snapshots, but observed no difference in either the conformations, nor degree of flexibility when compared to abacavir-unbound systems. Our results support hypersensitivity models in which abacavir-binding alters the conformational ensemble of neopeptides, so as to favour exposed peptide surfaces that are no longer recognized as self by circulating CD8+ T cells, and are conducive to TCR binding. Our findings highlight the need to also consider the role of dynamics in understanding drug-induced hypersensitivities at the molecular and mechanistic level. This additional insight can help inform the chemical modification of abacavir to prevent hypersensitivity reactions in HLA-B*57:01+ HIV patients whilst retaining potent antiretroviral activity.

The effect of Heweijiangni-decoction on esophageal morphology in a rat model of OVA-induced visceral hypersensitivity followed by acid exposure

Heweijiangni decoction (HWJND) is an effective traditional Chinese medicine prescription in clinical treatment of nonerosive reflux disease (NERD). Esophageal hypersensitivity and acid contribute to the disease. However, the exact underlying mechanism of action remains unclear. In this study, we observed the effect of HWJND on esophageal morphology in a rat model of ovalbumin (OVA)-induced visceral hypersensitivity followed by acid exposure. Esophageal morphology was assessed by measuring the extent of dilated intercellular spaces (DIS), desmosome disruption, and mitochondrial fragmentation. HWJND in low, moderate, and high doses relieved DIS and desmosome disruption in esophageal epithelium compared with model group (P&lt;0.05 for all doses). In addition, HWJND in high dose protected mitochondria from fragmentation (P&lt;0.05). Other findings suggest that DIS and mitochondrial fragmentation are independent events, and that omeprazole protects mitochondria. Overall, HWJND significantly resists esophageal morphology changes in OVA-induced and acid exposure rat model.

The discovery of novel 3-aryl-indazole derivatives as peripherally restricted pan-Trk inhibitors for the treatment of pain.

The design, synthesis, and biological evaluation of novel 3-aryl-indazole derivatives as peripherally selective pan-Trk inhibitors are described. Three strategies were used to obtain a potent compound exhibiting low central nervous system (CNS) penetration and high plasma exposure: 1) a structure-based drug design (SBDD) approach was used to improve potency; 2) a substrate for an efflux transporter for lowering brain penetration was explored; and 3) the most basic pKa (pKa-MB) value was used as an indicator to identify compounds with good membrane permeability. This enabled the identification of the peripherally targeted 17c with the potency, kinase-selectivity, and plasma exposure required to demonstrate in vivo efficacy in a Complete Freund's adjuvant (CFA)-induced thermal hypersensitivity model.

APRIL Induces a Novel Subset of IgA+ Regulatory B Cells That Suppress Inflammation via Expression of IL-10 and PD-L1.

Regulatory B cells (Bregs) are immunosuppressive cells that modulate immune responses through multiple mechanisms. The signals required for the differentiation and activation of these cells remain still poorly understood. We have already shown that overexpression of A PRoliferation-Inducing Ligand (APRIL) reduces the incidence and severity of collagen-induced arthritis (CIA) in mice. Furthermore, we have described that APRIL, but not BAFF, promoted IL-10 production and regulatory functions in human B cells. Therefore, we hypothesized that APRIL, but not BAFF, may be involved in the induction and/or activation of IL-10 producing Bregs that suppress inflammatory responses in vitro and in vivo. Here, we describe that APRIL promotes the differentiation of naïve human B cells to IL-10-producing IgA+ B cells. These APRIL-induced IgA+ B cells display a Breg phenotype and inhibit T cell and macrophage responses through IL-10 and PD-L1. Moreover, APRIL-induced IL-10 producing Bregs suppress inflammation in vivo in experimental autoimmune encephalitis (EAE) and contact hypersensitivity (CHS) models. Finally, we showed a strong correlation between APRIL and IL-10 in the inflamed synovial tissue of inflammatory arthritis patients. Collectively, these observations indicate the potential relevance of this novel APRIL-induced IgA+ Breg population for immune homeostasis and immunopathology.

Action of MK-7264 (gefapixant) at human P2X3 and P2X2/3 receptors and in vivo efficacy in models of sensitisation.

The P2X3 receptor is an ATP-gated ion channel expressed by sensory afferent neurons and is used as a target to treat chronic sensitisation conditions. The first-in-class, selective P2X3 and P2X2/3 receptor antagonist, the diaminopyrimidine MK-7264 (gefapixant), has progressed to Phase III trials for refractory or unexplained chronic cough. We used patch clamp to elucidate the pharmacology and kinetics of MK-7264 and rat models of hypersensitivity and hyperalgesia to test its efficacy on these conditions. Whole-cell patch clamp of 1321N1 cells expressing human P2X3 and P2X2/3 receptors was used to determine mode of MK-7264 action, potency, and kinetics. The analgesic efficacy was assessed using paw withdrawal threshold and limb weight distribution in rat models of inflammatory, osteoarthritic, and neuropathic sensitisation. MK-7264 is a reversible allosteric antagonist at human P2X3 and P2X2/3 receptors. Experiments with the slowly desensitising P2X2/3 heteromer revealed concentration- and state-dependency to wash-on, with faster rates and greater inhibition when applied before agonist compared to during agonist application. The wash-on rate (τ value) for MK-7264 at maximal concentrations was much lower when applied before compared to during agonist application. In vivo, MK-7264 displayed efficacy comparable to naproxen in inflammatory and osteoarthritic sensitisation models and gabapentin in neuropathic sensitisation models, increasing paw withdrawal threshold and decreasing weight-bearing discomfort. MK-7264 is a reversible and selective P2X3 and P2X2/3 antagonist, exerting allosteric antagonism via preferential activity at closed channels. Its efficacy in rat models supports its clinical investigation for chronic sensitisation conditions.

Sa1312 – Ameliorating Effects of Sacral Nerve Stimulation and Mechanisms Involving Nerve Growth Factor and Trpv1 in a Rodent Model of Visceral Hypersensitivity

Sa1312 – Ameliorating Effects of Sacral Nerve Stimulation and Mechanisms Involving Nerve Growth Factor and Trpv1 in a Rodent Model of Visceral Hypersensitivity

Su1578 – Chronic Intracolonic Administration of Linaclotide Inhibits Nociceptive Signaling in a Mouse Model of Chronic Visceral Hypersensitivity

Su1578 – Chronic Intracolonic Administration of Linaclotide Inhibits Nociceptive Signaling in a Mouse Model of Chronic Visceral Hypersensitivity

Skin Microbiome Modulates the Effect of Ultraviolet Radiation on Cellular Response and Immune Function

SummaryThe skin is colonized by a diverse microbiome intricately involved in various molecular and cellular processes within the skin and beyond. UV radiation is known to induce profound changes in the skin and modulate the immune response. However, the role of the microbiome in UV-induced immune suppression has been overlooked. By employing the standard model of contact hypersensitivity (using germ-free mice) we found diminished UV-induced systemic immune suppression in the presence of microbiome. Upon UV exposure, we found enhanced epidermal hyperplasia and neutrophilic infiltration in the presence and enhanced numbers of mast cells and monocyte or macrophages in the absence of microbiome. Transcriptome analysis revealed a predominant expression of cytokine genes related to pro-inflammatory milieu in the presence versus immunosuppressive milieu (with increased interleukin-10) in the absence of microbiome. Collectively, microbiome abrogates the immunosuppressive response to UV by modulating gene expression and cellular microenvironment of the skin.

051 Dermal dendritic cells present neo-self antigens induced by ultraviolet B exposure to expand Foxp3+ regulatory T cells

Dendritic cells play a key role in controlling homeostasis Foxp3+ regulatory T cells (Tregs), which constitute about 5-10% of peripheral CD4+T cells. The literature showed that Tregs contributed to immunosuppression induced by ultraviolet B (UVB) exposure using contact hypersensitivity models. Here we found that Tregs were expanded to more than half of CD4+T cells in the murine skin after UVB exposure without hapten-application. Upon UVB exposure, Tregs actively proliferated via interacting with dermal dendritic cells in the absence of exogenous antigen in vivo. Moreover, Treg proliferation were stimulated by a subset of dermal dendritic cells from UVB-exposed skin without exogenous antigen in vitro, indicating that neo-self antigens induced by UVB exposure are presented by the dendritic cell subset. RNA sequencing analysis showed that the dermal dendritic cell subset highly expressed immunological tolerance related genes such as Areg, Tnfsf9, and PD-L1. UVB-expanded Tregs in the skin also contributed to the homeostasis of Tregs in systemic lymphoid organs. These results suggest that immunological self-tolerance is regulated by Tregs expanded by a special subset of dendritic cells presenting neo-self antigens induced by UVB exposure.

Dietary coconut oil ameliorates skin contact hypersensitivity through mead acid production in mice.

Coconut oil is used as a dietary oil worldwide, and its healthy effects are recognized by the fact that coconut oil is easy to digest, helps in weight management, increases healthy cholesterol, and provides instant energy. Although topical application of coconut oil is known to reduce skin infection and inflammation, whether dietary coconut oil has any role in decreasing skin inflammation is unknown. In this study, we showed the impact of dietary coconut oil in allergic skin inflammation by using a mouse model of contact hypersensitivity (CHS). Mice maintained on coconut oil showed amelioration of skin inflammation and increased levels of cis-5, 8, 11-eicosatrienoic acid (mead acid) in serum. Intraperitoneal injection of mead acid inhibited CHS and reduced the number of neutrophils infiltrating tothe skin. Detailed mechanistic studies unveiled that mead acid inhibited the directional migration of neutrophils by inhibiting the filamentous actin polymerization and leukotriene B4 production required for secondary recruitment of neutrophils. Our findings provide valuable insights into the preventive roles of coconut oil and mead acid against skin inflammation, thereby offering attractive therapeutic possibilities.

Reversal of Peripheral Neuropathic Pain by the Small-Molecule Natural Product Physalin F via Block of CaV2.3 (R-Type) and CaV2.2 (N-Type) Voltage-Gated Calcium Channels

No universally efficacious therapy exists for chronic pain, a disease affecting one-fifth of the global population. An overreliance on the prescription of opioids for chronic pain despite their poor ability to improve function has led to a national opioid crisis. In 2018, the NIH launched a Helping to End Addiction Long-term plan to spur discovery and validation of novel targets and mechanisms to develop alternative nonaddictive treatment options. Phytochemicals with medicinal properties have long been used for various treatments worldwide. The natural product physalin F, isolated from the Physalis acutifolia (family: Solanaceae) herb, demonstrated antinociceptive effects in models of inflammatory pain, consistent with earlier reports of its anti-inflammatory and immunomodulatory activities. However, the target of action of physalin F remained unknown. Here, using whole-cell and slice electrophysiology, competition binding assays, and experimental models of neuropathic pain, we uncovered a molecular target for physalin F's antinociceptive actions. We found that physalin F (i) blocks CaV2.3 (R-type) and CaV2.2 (N-type) voltage-gated calcium channels in dorsal root ganglion (DRG) neurons, (ii) does not affect CaV3 (T-type) voltage-gated calcium channels or voltage-gated sodium or potassium channels, (iii) does not bind G-protein coupled opioid receptors, (iv) inhibits the frequency of spontaneous excitatory postsynaptic currents (EPSCs) in spinal cord slices, and (v) reverses tactile hypersensitivity in models of paclitaxel-induced peripheral neuropathy and spinal nerve ligation. Identifying CaV2.2 as a molecular target of physalin F may spur its use as a tool for mechanistic studies and position it as a structural template for future synthetic compounds.

Cerebrolysin improves peripheral inflammatory pain: Sex differences in two models of acute and chronic mechanical hypersensitivity.

Chronic inflammatory pain is a major health problem worldwide with high prevalence in women. Cerebrolysin is a multimodal neuropeptide preparation that crosses the blood brain barrier and displays neuroprotective properties in aging and disease. Previously, we showed that cerebrolysin reduced mechanical allodynia in a model of persistent inflammation and pain. We aim to build upon the findings of our previous study by investigating the response to acute administration of cerebrolysin in two models of peripheral inflammation and assessing sex differences. We utilized the complete Freund's adjuvant (CFA) that produces maximal oedema and mechanical allodynia within days and carrageenan that produces similar effects within hours. Cerebrolysin reversed the mechanical allodynia in both sexes in CFA-treated rats. On the other hand, in rats treated with carrageenan, cerebrolysin was only effective in reducing mechanical allodynia in female rats. In conclusion, the present study shows that cerebrolysin effects may be sex-specific depending on different mechanisms that are at play in these two models of peripheral inflammatory pain. Further investigations are required to determine the factors contributing to sex differences.

Future Directions for Understanding Adolescent Bipolar Spectrum Disorders: A Reward Hypersensitivity Perspective

The idea that bipolar spectrum disorders (BSDs) are characterized by enhanced sensitivity to rewarding stimuli is at the core of the reward hypersensitivity model, one of the most prominent and well-supported theories of BSDs. In this article, we present the reward hypersensitivity model of BSDs, review evidence supporting it, discuss its relevance to explaining why BSDs typically begin and consolidate during the period of adolescence, and consider three major unresolved issues for this model that provide important directions for future research. Finally, we present integrations of the reward hypersensitivity model with circadian rhythm and immune system models that should provide greater understanding of the mechanisms involved in BSDs, and then suggest additional directions for future research deriving from these integrated models.

Inhibition of Cav 3.2 calcium channels: A new target for colonic hypersensitivity associated with low-grade inflammation.

Abdominal pain associated with low-grade inflammation is frequently encountered in irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) during remission. Current treatments are not very effective and new therapeutic approaches are needed. The role of CaV 3.2 channels, which are important in other chronic pain contexts, was investigated in a murine model of colonic hypersensitivity (CHS) associated with low-grade inflammation. Low doses of dextran sulfate sodium (DSS; 0.5%) were chronically administered to C57BL/6j mice in drinking water. Their inflammatory state was assessed by systemic and local measures of IL-6, myeloperoxidase, and lipocalin-2 using elisa. Colonic sensitivity was evaluated by the visceromotor responses to colorectal distension. Functional involvement of CaV 3.2 channels was assessed with different pharmacological (TTA-A2, ABT-639, and ethosuximide) and genetic tools. DSS induced low-grade inflammation associated with CHS in mice. Genetic or pharmacological inhibition of CaV 3.2 channels reduced CHS. Cav3.2 channel deletion in primary nociceptive neurons in dorsal root ganglia (CaV 3.2Nav1.8 KO mice) suppressed CHS. Spinal, but not systemic, administration of ABT-639, a peripherally acting T-type channel blocker, reduced CHS. ABT-639 given intrathecally to CaV 3.2Nav1.8 KO mice had no effect, demonstrating involvement of CaV 3.2 channels located presynaptically in afferent fibre terminals. Finally, ethosuximide, which is a T-type channel blocker used clinically, reduced CHS. These results suggest that ethosuximide represents a promising drug reposition strategy and that inhibition of CaV 3.2 channels is an attractive therapeutic approach for relieving CHS in IBS or IBD.

Transcription factor Sp4 is required for hyperalgesic state persistence.

Understanding how painful hypersensitive states develop and persist beyond the initial hours to days is critically important in the effort to devise strategies to prevent and/or reverse chronic painful states. Changes in nociceptor transcription can alter the abundance of nociceptive signaling elements, resulting in longer-term change in nociceptor phenotype. As a result, sensitized nociceptive signaling can be further amplified and nocifensive behaviors sustained for weeks to months. Building on our previous finding that transcription factor Sp4 positively regulates the expression of the pain transducing channel TRPV1 in Dorsal Root Ganglion (DRG) neurons, we sought to determine if Sp4 serves a broader role in the development and persistence of hypersensitive states in mice. We observed that more than 90% of Sp4 staining DRG neurons were small to medium sized, primarily unmyelinated (NF200 neg) and the majority co-expressed nociceptor markers TRPV1 and/or isolectin B4 (IB4). Genetically modified mice (Sp4+/-) with a 50% reduction of Sp4 showed a reduction in DRG TRPV1 mRNA and neuronal responses to the TRPV1 agonist—capsaicin. Importantly, Sp4+/- mice failed to develop persistent inflammatory thermal hyperalgesia, showing a reversal to control values after 6 hours. Despite a reversal of inflammatory thermal hyperalgesia, there was no difference in CFA-induced hindpaw swelling between CFA Sp4+/- and CFA wild type mice. Similarly, Sp4+/- mice failed to develop persistent mechanical hypersensitivity to hind-paw injection of NGF. Although Sp4+/- mice developed hypersensitivity to traumatic nerve injury, Sp4+/- mice failed to develop persistent cold or mechanical hypersensitivity to the platinum-based chemotherapeutic agent oxaliplatin, a non-traumatic model of neuropathic pain. Overall, Sp4+/- mice displayed a remarkable ability to reverse the development of multiple models of persistent inflammatory and neuropathic hypersensitivity. This suggests that Sp4 functions as a critical control point for a network of genes that conspire in the persistence of painful hypersensitive states.

Ameliorating effects of optimized gastric electrical stimulation and mechanisms involving nerve growth factor and opioids in a rodent model of gastric hypersensitivity

Gastric electrical stimulation (GES) has been applied to treat gastric motility disorders for decades. This study was designed to investigate the effects and mechanisms of GES for visceral hypersensitivity in a rodent model of functional dyspepsia (FD). Male Sprague-Dawley rat pups at 10-days old received 0.1% iodoacetamide (IA) daily for 6days. The experiments were performed when the rats reached 8-11weeks of age, and visceral hypersensitivity was established. Then, GES parameters were optimized and the chronic effects of GES on gastric hypersensitivity were assessed by electromyogram (EMG). Naloxone (3mg/kg), D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP, 1mg/kg), and anti-NGF (16μg/kg) were individually intraperitoneally injected to investigate opioid and nerve growth factor (NGF) mechanisms. Tissues were analyzed for NGF expression. In the IA-treated rats, the visceromotor response to gastric distension was significantly increased, and both acute GES with optimized stimulation parameters (0.25seconds on, 0.25seconds off, 100Hz, 0.25ms, 6mA) and chronic GES (7days, 2 hours/day) normalized gastric hypersensitivity. The inhibitory effect of GES on gastric hypersensitivity was blocked by naloxone and CTOP. Anti-NGF normalized EMG responses in IA-treated rats. The expressions of NGF in the tissues of IA-treated rats were dramatically increased, and these increases were suppressed with GES. GES with optimized parameters improves gastric hypersensitivity induced by neonatal treatment of IA mediated peripherally by suppressing NGF and via the opioid mechanism involving the µ receptor. GES as a potential therapy for treating visceral pain may be explored in clinical studies.