Cadmium exposure exacerbates immunological abnormalities in a BTBR T+ Itpr3tf/J autistic mouse model by upregulating inflammatory mediators in CD45R-expressing cells.

Autism spectrum disorder (ASD) is a neurodevelopmental illness characterized by behavior, learning, communication, and social interaction abnormalities in various situations. Individuals with impairments usually exhibit restricted and repetitive actions. The actual cause of ASD is yet unknown. It is believed, however, that a mix of genetic and environmental factors may play a role in its development. Certain metals have been linked to the development of neurological diseases, and the prevalence of ASD has shown a positive association with industrialization. Cadmium chloride (Cd) is a neurotoxic chemical linked to cognitive impairment, tremors, and neurodegenerative diseases. The BTBR T+ Itpr3tf/J (BTBR) inbred mice are generally used as a model for ASD and display a range of autistic phenotypes. We looked at how Cd exposure affected the signaling of inflammatory mediators in CD45R-expressing cells in the BTBR mouse model of ASD. In this study, we looked at how Cd affected the expression of numerous markers in the spleen, including IFN-γ, IL-6, NF-κB p65, GM-CSF, iNOS, MCP-1, and Notch1. Furthermore, we investigated the effect of Cd exposure on the expression levels of numerous mRNA molecules in brain tissue, including IFN-γ, IL-6, NF-κB p65, GM-CSF, iNOS, MCP-1, and Notch1. The RT-PCR technique was used for this analysis. Cd exposure increased the number of CD45R+IFN-γ+, CD45R+IL-6+, CD45R+NF-κB p65+, CD45R+GM-CSF+, CD45R+GM-CSF+, CD45R+iNOS+, and CD45R+Notch1+ cells in the spleen of BTBR mice. Cd treatment also enhanced mRNA expression in brain tissue for IFN-γ, IL-6, NF-κB, GM-CSF, iNOS, MCP-1, and Notch1. In general, Cd increases the signaling of inflammatory mediators in BTBR mice. This study is the first to show that Cd exposure causes immune function dysregulation in the BTBR ASD mouse model. As a result, our study supports the role of Cd exposure in the development of ASD.

Histamine H4 Receptor Antagonist Ameliorates the Progression of Experimental Autoimmune Encephalomyelitis via Regulation of T-Cell Imbalance.

Multiple sclerosis (MS) is a degenerative condition characterized by immune-mediated attacks on the central nervous system (CNS), resulting in demyelination and recurring T-cell responses. The histamine H4 receptor (H4R) is mainly expressed in cellular populations and plays a vital role in inflammation and immunological responses. The role of H4R in neurons of the CNS has recently been revealed. However, the precise role of H4R in neuronal function remains inadequately understood. The objective of this work was to investigate the impact of JNJ 10191584 (JNJ), a highly effective and specific H4R antagonist, on the development of experimental autoimmune encephalomyelitis (EAE) and to gain insight into the underlying mechanism involved. In this study, we examined the potential impact of JNJ therapy on the course of EAE in SJL/J mice. EAE mice were administered an oral dose of JNJ at a concentration of 6 mg/kg once a day, starting from day 10 and continuing until day 42. Afterward, the mice's clinical scores were assessed. In this study, we conducted additional research to examine the impact of JNJ on several types of immune cells, specifically Th1 (IFN-γ and T-bet), Th9 (IL-9 and IRF4), Th17 (IL-17A and RORγt), and regulatory T (Tregs; Foxp3 and TGF-ß1) cells in the spleen. In this study, we further investigated the impact of JNJ on the mRNA expression levels of IFN-γ, T-bet, IL-9, IRF4, IL-17A, RORγt, Foxp3, and TGF-ß1 in the brain. Daily treatment of JNJ effectively reduced the development of EAE in mice. The percentages of CD4+IFN-γ+, CD4+T-bet+, CD4+IL-9+, CD4+IRF4+, CD4+IL-17A+, and CD4+RORγt+ cells were shown to decrease, whereas the percentages of CD4+TGF-ß1+ and CD4+Foxp3+ cells were observed to increase in EAE mice treated with JNJ. Therefore, the HR4 antagonist positively affected the course of EAE by modulating the signaling of transcription factors. The identified results include possible ramifications in the context of MS treatment.

MAP kinase inhibitor PD98059 regulates Th1, Th9, Th17, and natural T regulatory cells in an experimental autoimmune encephalomyelitis mouse model of multiple sclerosis.

Experimental autoimmune encephalitis (EAE), an animal model of multiple sclerosis (MS), provides significant insights into the mechanisms that initiate and drive autoimmunity. MS is a chronic autoimmune disease of the central nervous system, characterized by inflammatory infiltration associated with demyelination. T lymphocyte cells play a crucial role in MS, whereas natural T regulatory (nTreg) cells prevent autoimmune inflammation by suppressing lymphocyte activity. This study sought to investigate the role of PD98059, a selective MAP kinase inhibitor, in Th1, Th9, Th17, and nTreg cells using the SJL/J mouse model of EAE. Following EAE development, the mice were intraperitoneally administered PD98059 (5 mg/kg for two weeks) daily. We evaluated the effects of PD98059 on Th1 (IFN-γ and T-bet), Th9 (IL-9 and IRF4), Th17 (IL-17A and RORγT), and nTreg (FoxP3 and Helios) cells in the spleen using flow cytometry. Moreover, we explored the effects of PD98059 on the IFN-γ, T-bet, IL-9, IRF4, IL-17A, RORγT, FoxP3, and Helios mRNA and protein levels in brain tissues using qRT-PCR and Western blot analyses. PD98059 treatment significantly decreased the proportion of CD4+IFN-γ+, CD4+T-bet+, CD4+IL-9+, CD4+IRF4+, CD4+IL-17A+, CD4+RORγT+, CD4+IL-17A+, and CD4+RORγT+ cells while increasing that of CD4+FoxP3+ and CD4+Helios+ cells. In addition, PD98059 administration decreased the mRNA and protein levels of IFN-γ, T-bet, IL-9, IRF4, IL-17A, and RORγT but increased those of FoxP3 and Helios in the brain tissue of EAE mice. Our findings suggest that PD98059 corrects immune dysfunction in EAE mice, which is concurrent with the modulation of multiple signaling pathways.

Rituximab exerts its anti-arthritic effects via inhibiting NF-κB/GM-CSF/iNOS signaling in B cells in a mouse model of collagen-induced arthritis.

Rheumatoidarthritis (RA) is an autoimmune disease characterized by uncontrolled joint inflammation and damage to bone and cartilage. B cells are known to play a crucial role in the pathogenesis and development of arthritis. Previous studies have found that B cells may be a potential target for treating RA. Rituximab, a monoclonal antibody targeting B cells, has induced long-term clinical responses in RA. Collagen-induced arthritis (CIA) mouse model is a widely studied autoimmune model of RA. CIA mouse model was used to investigate the effect of rituximab on the RA severity in the mice. Following induction of CIA, animals were treated with rituximab (250 mg/kg/week) intraperitoneally on the days 28, 35, 42, 49, 56, and 63 after collagen induction. We investigated the effect of rituximab on NF-κB p65, IκBα, GM-CSF, MCP-1, iNOS, TNF-α, and IL-6 cells in splenic CD19+ and CD45R+ B cells using flow cytometry. We also assessed the effect of rituximab on NF-κB p65, GM-CSF, IκBα, MCP-1, iNOS, TNF-α, and IL-6 at mRNA levels using RT-PCR analyses of knee tissues. Rituximab treatment significantly decreased CD19+NF-κB p65+, CD45R+NF-κB p65+, CD19+GM-CSF+, CD45R+GM-CSF+, CD19+MCP-1+, CD45R+MCP-1+, CD19+TNF-α+, CD45R+TNF-α+, CD19+iNOS+, CD45R+iNOS+, CD19+IL-6+, and CD45R+IL-6+, and increased CD45R+IκBα+ in spleen cells of CIA mice. We further observed that rituximab treatment downregulated NF-κB p65, GM-CSF, MCP-1, iNOS, TNF-α, and IL-6, whereas it upregulated IκBα, mRNA level. All these findings suggest that rituximab may be a novel therapeutic target for the treatment of RA.

CXCR3 antagonist NBI-74330 mitigates joint inflammation in Collagen-Induced arthritis model in DBA/1J mice.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by uncontrolled synovial proliferation, pannus formation, cartilage injury, and bone destruction. We used the CXCR3-specific antagonist NBI-74330 to block T-cell-mediated signaling in a DBA/1J mouse model of collagen-induced arthritis (CIA). After CIA induction, DBA/1J mice were treated with NBI-74330 (100 mg/kg) daily from day 21 until day 34 and evaluated for arthritic score and histopathological changes. Furthermore, using flow cytometry, we investigated the effects of NBI-74330 on Th1 (IFN-γ, TNF-α, T-bet, STAT4, Notch-3, and RANKL), Th17 (IL-21, IL-17A, STAT3, and RORγt), and Th22 (IL-22) cells in splenic CD4+ and CXCR3+T-cells. We also used RT-PCR to assess the effect of mRNA levels of IFN-γ, TNF-α, T-bet, RANKL, IL-17A, RORγt, and IL-22 in knee tissues. The IFN-γ, TNF-α, and IL-17A serum protein levels were measured using ELISA. Compared to vehicle-treated CIA mice, the severity of arthritic scores and histological severity of inflammation decreased significantly in NBI-74330-treated CIA mice. Moreover, compared to vehicle-treated CIA mice, the percentages of CD4+IFN-γ+, CD4+TNF-α+, CD4+T-bet+, CD4+STAT4+, CD4+Notch-3+, CXCR3+IFN-γ+, CXCR3+TNF-α+, CXCR3+T-bet+, CXCR3+STAT4+, CXCR3+Notch-3+, CD4+RANKL+, CD4+IL-21+, CD4+IL-17A+, CD4+STAT3+, CD4+RORγt+, and CD4+IL-22+ cells decreased in NBI-74330-treated CIA mice. Furthermore, NBI-74330-treatment downregulated IFN-γ, TNF-α, T-bet, RANKL, STAT3, IL-17A, RORγt, and IL-22 mRNA levels. Serum IFN-γ, TNF-α, and IL-17A levels were significantly lower in NBI-74330-treated CIA mice than in vehicle-treated CIA mice. This study demonstrates the antiarthritic effects of NBI-74330 in CIA mice. Therefore, these data suggest that NBI-74330 could be considered a potential RA treatment.

Cadmium Exposure Is Associated with Behavioral Deficits and Neuroimmune Dysfunction in BTBR T+ Itpr3tf/J Mice.

Autism spectrum disorders (ASD) are neurobehavioral disabilities characterized by impaired social interactions, poor communication skills, and restrictive/repetitive behaviors. Cadmium is a common heavy metal implicated in ASD. In this study, we investigated the effects of Cd exposure on BTBR T+ Itpr3tf/J (BTBR) mice, an ASD model. We looked for changes in repetitive behaviors and sociability through experiments. We also explored the molecular mechanisms underlying the effects of Cd exposure, focusing on proinflammatory cytokines and pathways. Flow cytometry measured IL-17A-, IL-17F-, IL-21-, TNF-α-, STAT3-, and RORγt-expressing CD4+ T cells from the spleens of experimental mice. We then used RT-PCR to analyze IL-17A, IL-17F, IL-21, TNF-α, STAT3, and RORγ mRNA expression in the brain. The results of behavioral experiments showed that Cd exposure significantly increased self-grooming and marble-burying in BTBR mice while decreasing social interactions. Cd exposure also significantly increased the number of CD4+IL-17A+, CD4+IL-17F+, CD4+IL-21+, CD4+TNF-α+, CD4+STAT3+, and CD4+RORγt+ cells, while upregulating the mRNA expression of the six molecules in the brain. Overall, our results suggest that oral exposure to Cd aggravates behavioral and immune abnormalities in an ASD animal model. These findings have important implications for ASD etiology and provide further evidence of heavy metals contributing to neurodevelopmental disorders through proinflammatory effects.

A potent and selective CXCR2 antagonist improves neuroimmune dysregulation through the inhibition of NF-κB and notch inflammatory signaling in the BTBR mouse model of autism.

Autism comprises a broad range of neurodevelopmental disorders characterized by social communication deficits and repetitive and stereotyped behaviors. Chemokine receptor CXCR2 is expressed on neurons and is upregulated in neurological disorders. BTBR T+ Itpr3tf/J (BTBR) mice, a model for autism that shows the core features of ASD. Here, we studied the anti-inflammatory effect of a potent and selective CXCR2 antagonist SB332235 in the BTBR mice. The CXCR2 antagonist represents a promising therapeutic agent for several neuroinflammatory disorders. In this study, we investigated the effects of SB332235 administration on NF-κB-, Notch-1-, Notch-3-, GM-CSF-, MCP-1-, IL-6-, and IL-2- and TGF-ß1-expressing CD40+ cells in BTBR and C57BL/6 (C57) mice in the spleen cells by flow cytometry. We further assessed the effect of SB332235 treatment on NF-κB, Notch-1, GM-CSF, MCP-1, IL-6, and IL-2 mRNA expression levels in the brain tissue by RT-PCR. We also explored the effect of SB332235 administration on NF-κB, GM-CSF, IL-6, and TGF-ß1 protein expression levels in the brain tissue by western blotting. The SB332235-treated BTBR mice significantly decreases in CD40 + NF-κB+, CD40 + Notch-1+, CD40 + Notch-3+, CD40 + GM-CSF+, CD40 + MCP-1+, CD40 + IL-6+, and CD40 + IL-2+, and increases in CD40 + TGF-ß1+ in the spleen cells. Our results further demonstrated that BTBR mice treated with SB332235 effectively decreased NF-κB, Notch-1, GM-CSF, MCP-1, IL-6, and IL-2, increasing TGF-ß1 mRNA and protein expression levels in the brain tissue. In conclusion, these results indicate that SB332235 elicits an anti-inflammatory response by downregulating the inflammatory mediators and NF-κB/Notch inflammatory signaling in BTBR mice. This could represent a promising novel therapeutic target for autism treatment.

S3I-201, a selective stat3 inhibitor, ameliorates clinical symptoms in a mouse model of experimental autoimmune encephalomyelitis through the regulation of multiple intracellular signalling in Th1, Th17, and treg cells.

CD4+ T cells, specifically Th cells (Th1 and Th17) and regulatory T cells (Tregs), play a pivotal role in the pathogenesis of multiple sclerosis (MS), a demyelinating autoimmune disease of the CNS. STAT3 inhibitors are potential therapeutic targets for several immune disorders. In this study, we investigated the role of a well-known STAT3 inhibitor, S3I-201, in experimental autoimmune encephalomyelitis (EAE), a model of MS. Following induction of EAE, mice were intraperitoneally administered S3I-201 (10 mg/kg) each day, beginning on day 14 and continuing till day 35 and were evaluated for clinical signs. Flow cytometry was used to investigate further the effect of S3I-201 on Th1 (IFN-γ, STAT1, pSTAT1, and T-bet), Th17 (IL-17A, STAT3, pSTAT3, and RORγt), and regulatory T cells (Treg, IL-10, TGF-ß1, and FoxP3) expressed in splenic CD4+ T cells. Moreover, we analyzed the effects of S3I-201 on mRNA and protein expression of IFN-γ, T-bet, IL-17A, STAT1, STAT3, pSTAT1, pSTAT3, RORγ, IL-10, TGF-ß1, and FoxP3 in the brains of EAE mice. The severity of clinical scores decreased in S3I-201-treated EAE mice compared to vehicle-treated EAE mice. S3I-201 treatment significantly decreased CD4+IFN-γ+, CD4+STAT1+, CD4+pSTAT1+, CD4+T-bet+, CD4+IL-17A+, CD4+STAT3+, CD4+pSTAT3+, and CD4+RORγt+ and increased CD4+IL-10+, CD4+TGF-ß1+, and CD4+FoxP3+ in the spleens of EAE mice. Additionally, S3I-201 administration in EAE mice significantly decreased the mRNA and protein expression of Th1 and Th17 and increased those of Treg. These results suggest that S3I-201 may have novel therapeutic potential against MS.

Mitogen-activated protein kinase inhibitor PD98059 improves neuroimmune dysfunction in experimental autoimmune encephalomyelitis in SJL/J mice through the inhibition of nuclear factor-kappa B signaling in B cells.

Multiple sclerosis (MS) is a severe autoimmune disease leading to demyelination, followed by consequent axonal degeneration, causing sensory, motor, cognitive, and visual symptoms. Experimental autoimmune encephalomyelitis (EAE) is the most well-studied animal model of MS. Most current MS treatments are not completely effective, and severe side effects remain a great challenge. In this study, we report the therapeutic efficacy of PD98059, a potent mitogen-activated protein kinase inhibitor, on proteolipid protein (PLP)139-151-induced EAE in SJL/J mice. Following the induction of EAE, mice were intraperitoneally treated with PD98059 (5 mg/kg for 14 days) daily from day 14 to day 28. This study investigated the effects of PD98059 on C-C motif chemokine receptor 6 (CCR6), CD14, NF-κB p65, IκBα, GM-CSF, iNOS, IL-6, TNF-α in CD45R+ B lymphocytes using flow cytometry. Furthermore, we analyzed the effect of PD98059 on CCR6, CD14, NF-κB p65, GM-CSF, iNOS, IL-6, and TNF-α mRNA and protein expression levels using qRT-PCR analysis in brain tissues. Mechanistic investigations revealed that PD98059-treated in mice with EAE had reduced CD45R+CCR6+, CD45R+CD14+, CD45R+NF-κB p65+, CD45R+GM-CSF+, CD45R+iNOS+, CD45R+IL-6+, and CD45R+TNF-α+ cells and increased CD45R+IκBα+ cells compared with vehicle-treated control mice in the spleen. Moreover, downregulation of CCR6, CD14, NF-κB p65, GM-CSF, iNOS, IL-6, and TNF-α mRNA expression level was observed in PD98059-treated mice with EAE compared with vehicle-treated control mice in the brain tissue. The results of this study demonstrate that PD98059 modulates inflammatory mediators through multiple cellular mechanisms. The results of this study suggest that PD98059 may be pursued as a therapeutic agent for the treatment of MS.

CC chemokine receptor 5 antagonist alleviates inflammation by regulating IFN-γ/IL-10 and STAT4/Smad3 signaling in a mouse model of autoimmune encephalomyelitis.

Multiple sclerosis (MS) is an immunopathological disease that causes demyelination and recurrent episodes of T cell-mediated immune attack in the central nervous system. Experimental autoimmune encephalomyelitis (EAE) is a well-established mouse model of MS. The roles of T cells in MS/EAE have been well investigated, but little is known about the role of CCR5+ cells. In the present study, we investigated whether treatment with DAPTA, a selective CCR5 antagonist, could modulate the progression of EAE in the SJL/J mice. EAE mice were treated with DAPTA (0.01 mg/kg) intraperitoneally daily from day 14 to day 42, and the clinical scores were evaluated. We further investigated the effects of DAPTA on IFN-γ-, TGF-ß-, IL-10-, IL-17A-, IL-22-, T-bet, STAT4-, RORγT-, AhR-, Smad3-, and Foxp3-expressing CCR5+ spleen cells using flow cytometry analysis. We further explored the effects of DAPTA on mRNA/protein expression of IFN-γ, IL-10, IL-17A, IL-22, TGF-ß, T-bet, STAT4, RORγT, AhR, Foxp3, and NF-H in the brain tissue. The severity of clinical scores decreased in DAPTA-treated EAE mice as compared to that in the EAE control mice. Moreover, the percentage of CCR5+IFN-γ+, CCR5+T-bet+, CCR5+STAT4+, CCR5+IL-17A+, CCR5+RORγt+, CCR5+IL-22+, and CCR5+AhR+ cells decreased while CCR5+TGF-ß+, CCR5+IL-10+, CCR5+Smad3+, and CCR5+Foxp3+ increased in DAPTA-treated EAE mice. Furthermore, DAPTA treatment significantly mitigated the EAE-induced expression of T-bet, STAT4, IL-17A, RORγT, IL-22, and AhR but upregulated Foxp3, IL-10, and NF-H expression in the brain tissue. Taken together, our data demonstrated that DAPTA could ameliorate EAE progression through the downregulation of the inflammation-related cytokines and transcription factors signaling, which may be useful for the clinical therapy of MS.

Lead (Pb) exposure exacerbates behavioral and immune abnormalities by upregulating Th17 and NF-κB-related signaling in BTBR T+ Itpr3tf/J autistic mouse model.

Autism spectrum disorder (ASD) is a highly prevalent neurodevelopmental disorder that are characterized by abnormal social interaction impairments in communication and repetitive and restricted activities or interests. Even though the exact etiology of ASD remains unknown. Lead (Pb) is a toxin known to harm many organs in the body, it is one of the most ubiquitous metal exposures which is associated with neurological deficits. Previous studies have shown that the exposure to Pb may play a role in ASD. BTBR T+ Itpr3tf/J (BTBR) mouse model is commonly used as a preclinical model for ASD. In this study, we investigated the effects of Pb exposure on sociability, self-grooming and marble burying behaviors tests in BTBR mice. We further examined the effects of Pb on IL-17A- RORγT-, STAT3-, NF-κB p65-, iNOS-, TLR-2- and TLR-4-producing CD45+ cells in spleen using flow cytometry. We also explored the effects of Pb on IL-17A, RORγT, STAT3, NF-κB p65, and TLR-2 mRNA expression in the brain tissue using RT-PCR analysis. Our results demonstrated that Pb exposure substantially increased repetitive behavior, marble burying and decrease social interactions in BTBR mice. In addition, in spleen cells, Pb exposure exaggerated CD45+IL-17A+, CD45+RORγT+, CD45+STAT3+, CD45+NF-κB p65+, CD45+iNOS+, CD45+TLR-2+ and CD45+TLR-4+ in BTBR mice. We also found that Pb significantly increased IL-17A, RORγT, STAT3, NF-κB p65, and TLR-2 mRNA in the brain tissue. Therefore, Pb exposure exacerbates behavioral and neuroimmune function in BTBR mice, suggesting a potentially strong role for Pb in ASD.

CXCR2 antagonist SB332235 mitigates deficits in social behavior and dysregulation of Th1/Th22 and T regulatory cell-related transcription factor signaling in male BTBR T+ Itpr3tf/J mouse model of autism.

Autism spectrum disorders is a complex neurodevelopmental disorder characterized by abnormal social interaction, defective communication, repetitive and stereotyped patterns of behaviors or interests. The BTBR T+ Itpr3tf/J (BTBR) inbred mice are generally used as a model for ASD, display a range of autistic phenotypes. Recent studies suggest that the CXCR2 antagonist is crucial for targets in the treatment of inflammatory and neurodegenerative diseases. In this study, we investigated the potential effects of the CXCR2 antagonist SB332235 on sociability behaviors, marble burying, and self-grooming, we also explored the treatment of SB332235 on Th1 (IFN-γ, Stat1, and T-bet), Th22 (IL-22, TNF-α, and AhR), and T regulatory (Treg, IL-10, Helios and Foxp3) production in CD4+ T cells in male BTBR and C57BL/6 (C57) mice in spleen. We also investigated the effects of SB332235 on IFN-γ, IL-10, IL-22, T-bet, AhR, and Foxp3 mRNA expression levels in the brain tissues. The SB332235-treated mice significantly improve behavioral abnormalities in BTBR mice. In addition, SB332235 administration causes a significantly decreases in IFN-γ, Stat1, T-bet, IL-22, TNF-α, and AhR, and increases in IL-10, Foxp3 and Helios production CD4+ T cells in BTBR mice. We further observed that SB332235 downregulated IFN-γ, IL-10, IL-22, T-bet, and AhR, and upregulated IL-10 and Foxp3 mRNA expression in the brain tissues. Our findings demonstrated that SB332235 treatment attenuated behavior deficits, through inhibiting Th1/Th22 and upregulating Treg cell-related transcription factors signaling pathway. Therefore, CXCR2 antagonist administration may be a promising therapeutic agent to attenuate behavior deficits via its anti-inflammatory effect.

Smo-Shh Agonist Purmorphamine Prevents Neurobehavioral and Neurochemical Defects in 8-OH-DPAT-Induced Experimental Model of Obsessive-Compulsive Disorder.

Obsessive-compulsive disorder is a mental disorder characterized by repetitive, unwanted thoughts and behavior due to abnormal neuronal corticostriatal-thalamocortical pathway and other neurochemical changes. Purmorphamine is a smoothened-sonic-hedgehog agonist that has a protective effect against many neurological diseases due to its role in maintaining functional connectivity during CNS development and its anti-inflammatory and antioxidant properties. As part of our current research, we investigated the neuroprotective effects of PUR against behavioral and neurochemical changes in 8-hydroxy-2-(di-n-propylamino)-tetralin-induced obsessive-compulsive disorder in rats. Additionally, the effect of PUR was compared with the standard drug for OCD, i.e., fluvoxamine. The intra-dorsal raphe-nucleus injection of 8-OH-DPAT in rats for seven days significantly showed OCD-like repetitive and compulsive behavior along with increased oxidative stress, inflammation, apoptosis, as well as neurotransmitter imbalance. These alterations were dose-dependently attenuated by long-term purmorphamine treatment at 5 mg/kg and 10 mg/kg i.p. In this study, we assessed the level of various neurochemical parameters in different biological samples, including brain homogenate, blood plasma, and CSF, to check the drug's effect centrally and peripherally. These effects were comparable to the standard oral treatment withfluvoxamine at 10 mg/kg. However, when fluvoxamine was given in combination with purmorphamine, there was a more significant restoration of these alterations than the individualtreatmentswithfluvoxamine and purmorphamine. All the above findings demonstrate that the neuroprotective effect of purmorphamine in OCD can be strong evidence for developing a new therapeutic target for treating and managing OCD.

Methylmercury chloride exposure exacerbates existing neurobehavioral and immune dysfunctions in the BTBR T+ Itpr3tf/J mouse model of autism.

Autism spectrum disorder (ASD) is a neurodevelopmental disease characterized by impaired communication, impaired reciprocal social interaction, restricted sociability deficits, and the presence of stereotyped patterns of behaviors. Immune dysregulation has been suggested to play a possible etiological role in ASD. Recent studies have demonstrated that exposure to methylmercury chloride (MeHgCl) leads to abnormal gait, motor deficits, impaired hearing, and memory deficits; however, its effects on behavioral and immunological responses have not been adequately investigated in ASD. In this study, we investigated the effects of MeHgCl exposure on marble burying, self-grooming behaviors, sociability tests, and locomotor activities in BTBR T+ Itpr3tf/J (BTBR) mice. We also explored the possible molecular mechanism underlying the effects of MeHgCl administration on IFN-γ-, T-bet-, IL-9-, and IL-17A-producing CD4+, CXCR5+, CXCR6+, and CCR9+ cells isolated from spleens. Furthermore, the effects of MeHgCl exposure on the mRNA expression and levels of pro-inflammatory cytokines in the brain tissue and serum samples were also assessed. Our results demonstrated that MeHgCl exposure caused a significant increase in marble burying, self-grooming behaviors and a decrease in social interactions and adverse effects on locomotor activity in BTBR mice. MeHgCl exposure also significantly increased the production of CD4+IFN-γ+, CD4+T-bet+, CCR9+T-bet+, CXCR5+IL-9+, CD4+IL-9+, CXCR6+IL-17A+, and CD4+IL-17A+ cells in the spleen. Furthermore, MeHgCl exposure increased mRNA and protein levels of pro-inflammatory cytokines in the brain and serum respectively in BTBR mice. In conclusion, MeHgCl administration aggravated existing behavioral and immune abnormalities in BTBR mice.

Cathepsin B inhibitor alleviates Th1, Th17, and Th22 transcription factor signaling dysregulation in experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS) is an autoimmune disease characterized by inflammatory infiltration in association with demyelination in the central nervous system. Among the factors involved in the immunological mechanisms of MS, Th1, Th17, and Th22 cells play a critical role. In the present study, we investigated the role of CA-074, a potent Cathepsin B inhibitor, in MS progression, using the SJL/J mouse model of experimental autoimmune encephalomyelitis (EAE). Following induction of EAE, mice were administered CA-074 (10 mg/kg) intraperitoneally each day, beginning on day 14 and continuing until day 28, and were evaluated for clinical signs. We further investigated the effect of CA-074 on Th1 (T-bet/STAT4), Th17 (IL-17A/RORγT), Th22 (TNF-α/IL-22), and regulatory T (Treg/Foxp3) cells in the spleen, using flow cytometry. We also analyzed the effect of CA-074 on T-bet, IL-17A, RORγT, IL-22, and mRNA and protein levels using RT-PCR and western blot analysis for brain tissues. Cathepsin B expression were also assessed by western blot in the brain tissues. The severity of clinical scores decreased significantly in CA-074-treated mice compared with that in EAE control mice. Moreover, the percentage of CD4+T-bet+, CXCR5+T-bet+, CD4+STAT4+, CD4+IL-17A+, CXCR5+IL-17A+, CD4+RORγT+, CCR6+RORγT+, CD4+TNF-α+, CD4+IL-22+, and CCR6+IL-22+ cells decreased while CD25+Foxp3+ increased in CA-074-treated EAE mice as compared to vehicle-treated EAE mice. Further, CA-074-treated EAE mice had downregulated Cathepsin B protein expression which was associated with decreased T-bet, IL-17A, RORγT, and IL-22 mRNA/protein expression. These results suggest that Cathepsin B could be a novel therapeutic candidate against for the treatment of MS.

CCR1 antagonist ameliorates experimental autoimmune encephalomyelitis by inhibition of Th9/Th22-related markers in the brain and periphery.

Multiple sclerosis (MS) is an immune-mediated disease of the central nervous system. The disease manifestation is associated with the proliferation and activation of lymphocytes and astrocytes, leading to demyelination and neuronal damage. Most of the current therapies are not completely effective, and few target the underlying pathophysiology of MS. T helper 9 (Th9)- and Th22-dominant cells have been proven to play a pathogenic role in experimental autoimmune encephalomyelitis (EAE), an animal model of MS. The goal of the present study was to investigate the therapeutic efficacy of J-113863, a novel CCR1 chemokine receptor, on PLP139-151-induced EAE in SJL/J mice. Following induction of EAE, mice were treated with J-113863 (10 mg/kg) or saline intraperitoneally daily from day 14 until day 25, and the clinical score was evaluated. We further investigated the effect of J-113863 on IL-9, IRF4, IL-22, IFN-γ, STAT3, AhR, and IL-17A in CD3+, CD4+, CCR6+, and CCR8+ spleen cells using flow cytometry. We also analyzed the effect of J-113863 on IL-9, IRF4, IL-22, IFN-γ, STAT3, AhR, and IL-17A mRNA expression levels. Our results revealed that J-113863 treatment notably attenuated the severity of clinical scores in EAE mice. J-113863 treatment decreased the percentage expression of CD4+IL-9+, CCR8+IL-9+, CD4+IRF4+, CD3+IL-22+, CCR6+IL-22+, CD3+IFN-γ+, CCR6+IFN-γ+, CD3+STAT3+, CCR6+STAT3+, CD4+IL-17A+, and CCR6+IL-17A+, and increased the percentage of CD3+AhR+, and CCR6+AhR+ cells in the spleen. These results confirmed that J-113863 suppressed Th9/Th22 cells to reduce demyelination in EAE mice, suggesting its potential role as a novel drug candidate for MS treatment.

5-Aminoisoquinolinone, a PARP-1 Inhibitor, Ameliorates Immune Abnormalities through Upregulation of Anti-Inflammatory and Downregulation of Inflammatory Parameters in T Cells of BTBR Mouse Model of Autism.

Autism spectrum disorder (ASD) covers a range of neurodevelopmental disorders involving impairments in communication and repetitive and stereotyped patterns of behavior and reciprocal social interaction. 5-Aminoisoquinolinone (5-AIQ), a PARP-1 inhibitor, has neuroprotective and anti-inflammatory effects. We investigated the influence of 5-AIQ-treatment in BTBR T+ Itpr3tf/J (BTBR) mice as an autism model and used flow cytometry to assess the effect of 5-AIQ on FOXP3, Helios, GATA3, IL-9, IL-10 and IL-17A production by CXCR6+ and CD4+ T cells in the spleen. We also confirmed the effect of 5-AIQ treatment on expression of FOXP3, Helios, GATA3, IL-17A, IL-10, and IL-9 mRNA and protein expression levels in the brain tissue by quantitative PCR and western blotting. Our results demonstrated that 5-AIQ-treated BTBR mice had significantly increased numbers of CXCR6+FOXP3+, CXCR6+IL-10+, and CXCR6+Helios+ cells and decreased numbers of CD4+GATA3+, CD4+IL-9+, and CD4+IL-17A+ cells as compared with those in untreated BTBR mice. Our results further demonstrated that treatment with 5-AIQ in BTBR mice increased expression for FOXP3, IL-10, and Helios, and decreased expression for GATA3, IL-17A, and IL-9 mRNA. Our findings support the hypotheses that 5-AIQ has promising novel therapeutic effects on neuroimmune dysfunction in autism and is associated with modulation of Treg and Th17 cells.

Dysregulation of Ki-67 Expression in T Cells of Children with Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by behavioral abnormalities such as impairments in social function and deficits in communication. The etiology of autism is unknown in most cases, but many studies have pointed towards the immune system as a causative agent in autism. Specific studies implicated lymphocytes, natural killer (NK) cells, monocytes, cytokines, and specific transcription factors in the development of ASD. The protein Ki-67 is n expressed in the proliferating cells and is used as a tool in several disorders. Ki-67 plays a crucial role in many neurological diseases. However, Ki-67 role in ASD is not fully understood. In this study, we investigated the possible role of Ki-67 expression in autistic children. We compared Ki-67 production in CD3+, CD4+, CD8+, CXCR4+, CXCR7+, CD45R+, HLA-DR+, GATA3+, Helios+, and FOXP3+ peripheral blood mononuclear cells (PBMCs) in autistic children to typically developing (TD) controls using immunofluorescence staining. We also determined Ki-67 mRNA levels in PBMCs using RT-PCR. The results revealed that autistic children had significantly increased numbers of CD3+Ki-67+, CD4+Ki-67+, CD8+Ki-67+, CXCR4+Ki-67+, CXCR7+Ki-67+, CD45R+Ki-67+, HLA-DR+Ki-67+, CXCR4+GATA3+, GATA3+Ki-67+ cells and decreased Helios+Ki-67+ and FOXP3+Ki-67+ cells compared with TD controls. In addition, the autistic children showed upregulation of Ki-67 mRNA levels compared with TD controls. Further studies need to be carried out to assess the exact role of Ki-67 and its therapeutic potential in ASD.

Upregulation of interleukin (IL)-31, a cytokine producing CXCR1 peripheral immune cells, contributes to the immune abnormalities of autism spectrum disorder.

Autism spectrum disorders (ASD) are a group of neurodevelopmental disorders characterized by communication deficits, impaired social interactions, and restricted stereotypical behaviors. Several immune cells are associated with immune dysfunction in ASD; however, IL-31 has not been explored in ASD. This study aims to investigate the role of inflammatory cytokines and transcription factors of the CXCR1 cells in children with ASD. In the current study, we investigated the cytokines and transcription factors produced by CXCR1+ cells (IL-31, IL-9, IL-21R, IL-21, NF-κB p65, RORγT, STAT1, and FoxP3) in peripheral blood mononuclear cells (PBMCs), from children with ASD and typically developing (TD) control children, using flow cytometric analysis. In addition, we measured mRNA and protein expression levels of IL-31 using quantitative real-time PCR and western blot analyses in PBMCs. In our study, children with ASD had increased CXCR1+IL-31+, CXCR1+IL-9+, CXCR1+IL-21R+, CXCR1+IL-21+, CXCR1+NF-κB+ p65, CXCR1+RORγT+, and CXCR1+STAT1+, and decreased CXCR1+FoxP3+ cells as compared with cells from the TD control samples. Similarly, children with ASD showed increased IL-31 mRNA and protein expression levels as compared to those of TD control samples. Our results suggest that upregulated production of inflammatory cytokines and transcription factors in CXCR1+ cells cause immunological imbalance in children with ASD. Therefore, attenuation of inflammatory cytokines/mediators and transcription factors could have a therapeutic potential in the treatment of ASD.

3-Aminobenzamide alleviates elevated DNA damage and DNA methylation in a BTBR T+Itpr3tf/J mouse model of autism by enhancing repair gene expression.

Little is known about genetic and epigenetic alterations in autism spectrum disorder. Moreover, the efficiency of DNA repair in autism must be improved to correct these alterations. We examined whether 3-aminobenzamide (3-AB) could reverse these alterations. We conducted experiments to clarify the molecular mechanism underlying these ameliorations. An assessment of genetic and epigenetic alterations by a modified comet assay showed elevated levels of oxidative DNA strand breaks and DNA hypermethylation in BTBR T+Itpr3tf/J (BTBR) mice used as a model of autism. Oxidative DNA strand breaks and DNA methylation were further quantified fluorometrically, and the results showed similar changes. Conversely, 3-AB treated BTBR mice showed a significant reduction in these alterations compared with untreated mice. The expressions of 43 genes involved in DNA repair were altered in BTBR mice. RT2 Profiler PCR Array revealed significantly altered expression of seven genes, which was confirmed by RT-PCR analyses. 3-AB treatment relieved these disturbances and significantly improved Ogg1 and Rad1 up-regulation. Moreover, autism-like behaviors were also mitigated in BTBR animals by 3-AB treatment without alterations in locomotor activities. The simultaneous effects of reduced DNA damage and DNA methylation levels as well as the regulation of repair gene expression indicate the potential of 3-AB as a therapeutic agent to decrease the levels of DNA damage and DNA methylation in autistic patients. The current data may help in the development of therapies that ultimately provide a better quality of life for individuals suffering from autism.