RESUMEN
This study aims to evaluate and compare cellular therapy with human Wharton's jelly (WJ) mesenchymal stem cells (MSCs) and neural precursors (NPs) in experimental autoimmune encephalomyelitis (EAE), a preclinical model of Multiple Sclerosis. MSCs were isolated from WJ by an explant technique, differentiated to NPs, and characterized by cytometry and immunocytochemistry analysis after ethical approval. Forty-eight rats were EAE-induced by myelin basic protein and Freund's complete adjuvant. Forty-eight hours later, the animals received intraperitoneal injections of 250 ng/dose of Bordetella pertussis toxin. Fourteen days later, the animals were divided into the following groups: a. non-induced, induced: b. Sham, c. WJ-MSCs, d. NPs, and e. WJ-MSCs plus NPs. 1 × 105. Moreover, the cells were placed in a 10 µL solution and injected via a stereotaxic intracerebral ventricular injection. After ten days, the histopathological analysis for H&E, Luxol, interleukins, and CD4/CD8 was carried out. Statistical analyses demonstrated a higher frequency of clinical manifestation in the Sham group (15.66%) than in the other groups; less demyelination was seen in the treated groups than the Sham group (WJ-MSCs, p = 0.016; NPs, p = 0.010; WJ-MSCs + NPs, p = 0.000), and a lower cellular death rate was seen in the treated groups compared with the Sham group. A CD4/CD8 ratio of <1 showed no association with microglial activation (p = 0.366), astrocytes (p = 0.247), and cell death (p = 0.577) in WJ-MSCs. WJ-MSCs and NPs were immunomodulatory and neuroprotective in cellular therapy, which would be translated as an adjunct in demyelinating diseases.
Asunto(s)
Encefalomielitis Autoinmune Experimental , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Esclerosis Múltiple , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/patología , Ratas , Esclerosis Múltiple/terapia , Esclerosis Múltiple/patología , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Humanos , Femenino , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Células-Madre Neurales , Modelos Animales de Enfermedad , Gelatina de Wharton/citologíaRESUMEN
Multiple sclerosis (MS) is a demyelinating central nervous system (CNS) disorder that is associated with functional impairment and accruing disability. There are multiple U.S. Food and Drug Administration (FDA)-approved drugs that effectively dampen inflammation and slow disability progression. However, these agents do not work well for all patients and are associated with side effects that may limit their use. The vagus nerve (VN) provides a direct communication conduit between the CNS and the periphery, and modulation of the inflammatory reflex via electrical stimulation of the VN (VNS) shows efficacy in ameliorating pathology in several CNS and autoimmune disorders. We therefore investigated the impact of VNS in a rat experimental autoimmune encephalomyelitis (EAE) model of MS. In this study, VNS-mediated neuroimmune modulation is demonstrated to effectively decrease EAE disease severity and duration, infiltration of neutrophils and pathogenic lymphocytes, myelin damage, blood-brain barrier disruption, fibrinogen deposition, and proinflammatory microglial activation. VNS modulates expression of genes that are implicated in MS pathogenesis, as well as those encoding myelin proteins and transcription factors regulating new myelin synthesis. Together, these data indicate that neuroimmune modulation via VNS may be a promising approach to treat MS, that not only ameliorates symptoms but potentially also promotes myelin repair (remyelination).
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Encefalomielitis Autoinmune Experimental , Esclerosis Múltiple , Estimulación del Nervio Vago , Nervio Vago , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/inmunología , Ratas , Esclerosis Múltiple/terapia , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/patología , Estimulación del Nervio Vago/métodos , Inflamación/terapia , Inflamación/patología , Modelos Animales de Enfermedad , Femenino , Vaina de Mielina/metabolismo , Barrera HematoencefálicaRESUMEN
INTRODUCTION: Multiple sclerosis (MS), as a destructive pathology of myelin in central nervous system (CNS), causes physical and mental complications. Experimental autoimmune encephalomyelitis (EAE) is laboratory model of MS widely used for CNS-associated inflammatory researches. Cell therapy using macrophage M2 (MPM2) is a cell type with anti-inflammatory characteristics for all inflammatory-based neuropathies. This experimental study investigated the probable therapeutic anti-inflammatory effects of intraperitoneal (IP) injection of MPM2 on alleviation of motor defect in EAE-affected animals. MATERIALS AND METHODS: 24 C57/BL6 female mice were divided into four groups of EAE, EAE + Dexa, EAE + PBS, and EAE + MP2. EAE was induced through deep cervical injection of spinal homogenate of guinea pigs. MPM2 cells were harvested from bone marrow and injected (106cells/ml) in three days of 10, 13 and 16 post-immunizations (p.i). Clinical score (CS), anti-inflammatory cytokines (IL-4, IL-10), pro-inflammatory gene expression (TNF-α, IL-1ß) and histopathological investigations (HE, Nissl and Luxol Fast Blue) were considered. Data were analyzed using SPSS software (v.19) and p < 0.05 was considered significant level. RESULTS: During EAE induction, the mean animal weight was decreased (p < 0.05); besides, following MPM2 injection, the weight gain was applied (p < 0.05) in EAE + MPM2 groups than control. Increased (p < 0.05) levels of CS was found during EAE induction in days 17-28 in EAE animals; besides, CS was decreased (p < 0.05) in EAE + MPM2 group than EAE animals. Also, in days 25-28 of experiment, the CS was decreased (p < 0.05) in EAE + MPM2 than EAE + Dexa. Histopathological assessments revealed low density of cell nuclei in corpus callosum, microscopically. LFB staining also showed considerable decrease in white matter density of corpus callosum in EAE group. Acceleration of white matter density was found in EAE + MPM2 group following cell therapy procedure. Genes expression of TNF-α, IL-1ß along with IL-4 and IL-10 were decreased (p < 0.05) in EAE + MPM2 group. CONCLUSION: IP injection of MPM2 to EAE-affected female mice can potentially reduce the CNS inflammation, neuronal death and myelin destruction. MPM2 cell therapy can improve animal motor defects.
Asunto(s)
Encefalomielitis Autoinmune Experimental , Células Madre Mesenquimatosas , Ratones Endogámicos C57BL , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/inmunología , Femenino , Ratones , Medios de Cultivo Condicionados/farmacología , Células Madre Mesenquimatosas/metabolismo , Inyecciones Intraperitoneales , Monocitos/inmunología , Cobayas , Citocinas/metabolismo , Células Cultivadas , Trasplante de Células Madre Mesenquimatosas , Esclerosis Múltiple/terapia , Esclerosis Múltiple/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Modelos Animales de Enfermedad , HumanosRESUMEN
Dendritic cells (DCs) are essential orchestrators of immune responses and represent potential targets for immunomodulation in autoimmune diseases. Human amniotic fluid secretome is abundant in immunoregulatory factors, with extracellular vesicles (EVs) being a significant component. However, the impact of these EVs on dendritic cells subsets remain unexplored. In this study, we investigated the interaction between highly purified dendritic cell subsets and EVs derived from amniotic fluid stem cell lines (HAFSC-EVs). Our results suggest that HAFSC-EVs are preferentially taken up by conventional dendritic cell type 2 (cDC2) through CD29 receptor-mediated internalization, resulting in a tolerogenic DC phenotype characterized by reduced expression and production of pro-inflammatory mediators. Furthermore, treatment of cDC2 cells with HAFSC-EVs in coculture systems resulted in a higher proportion of T cells expressing the regulatory T cell marker Foxp3 compared to vehicle-treated control cells. Moreover, transfer of HAFSC-EV-treated cDC2s into an EAE mouse model resulted in the suppression of autoimmune responses and clinical improvement. These results suggest that HAFSC-EVs may serve as a promising tool for reprogramming inflammatory cDC2s towards a tolerogenic phenotype and for controlling autoimmune responses in the central nervous system, representing a potential platform for the study of the effects of EVs in DC subsets.
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Líquido Amniótico , Células Dendríticas , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental , Vesículas Extracelulares , Esclerosis Múltiple , Animales , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/inmunología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Ratones , Líquido Amniótico/citología , Líquido Amniótico/metabolismo , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/metabolismo , Humanos , Esclerosis Múltiple/terapia , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/metabolismo , Femenino , Células Madre/metabolismo , Células Madre/citología , Ratones Endogámicos C57BLRESUMEN
Proinflammatory T-lymphocytes recruited into the brain and spinal cord mediate multiple sclerosis (MS) and currently there is no cure for MS. IFN-γ-producing Th1 cells induce ascending paralysis in the spinal cord while IL-17-producing Th17 cells mediate cerebellar ataxia. STAT1 and STAT3 are required for Th1 and Th17 development, respectively, and the simultaneous targeting of STAT1 and STAT3 pathways is therefore a potential therapeutic strategy for suppressing disease in the spinal cord and brain. However, the pharmacological targeting of STAT1 and STAT3 presents significant challenges because of their intracellular localization. We have developed a STAT-specific single-domain nanobody (SBT-100) derived from camelids that targets conserved residues in Src homolog 2 (SH2) domains of STAT1 and STAT3. This study investigated whether SBT-100 could suppress experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. We show that SBT-100 ameliorates encephalomyelitis through suppressing the expansion of Th17 and Th1 cells in the brain and spinal cord. Adoptive transfer experiments revealed that lymphocytes from SBT-100-treated EAE mice have reduced capacity to induce EAE, indicating that the immunosuppressive effects derived from the direct suppression of encephalitogenic T-cells. The small size of SBT-100 makes this STAT-specific nanobody a promising immunotherapy for CNS autoimmune diseases, including multiple sclerosis.
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Encefalomielitis Autoinmune Experimental , Anticuerpos de Dominio Único , Células Th17 , Animales , Femenino , Ratones , Camélidos del Nuevo Mundo , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Ratones Endogámicos C57BL , Enfermedades Neuroinflamatorias/inmunología , Enfermedades Neuroinflamatorias/tratamiento farmacológico , Anticuerpos de Dominio Único/farmacología , Anticuerpos de Dominio Único/inmunología , Anticuerpos de Dominio Único/uso terapéutico , Médula Espinal/patología , Médula Espinal/efectos de los fármacos , Médula Espinal/inmunología , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT3/metabolismo , Células TH1/inmunología , Células TH1/efectos de los fármacos , Células Th17/inmunología , Células Th17/efectos de los fármacosRESUMEN
The most common central nervous system (CNS) inflammatory disease is multiple sclerosis (MS), modeled using experimental autoimmune encephalomyelitis (EAE). Mesenchymal stem cells (MSCs) exhibit potent immunomodulatory capabilities, including the suppression of immune cell functions and anti-inflammatory cytokine production. Female C57BL/6 mice (8-10 weeks old) were divided into three groups: 1. Control, 2. Allogeneic MSCs (ALO) treatment, and 3. Syngeneic MSCs (SYN) treatment. To induce EAE, myelin oligodendrocyte glycoprotein was injected subcutaneously with complete Freund's adjuvant, followed by intraperitoneal pertussis toxin. On Days 6 and 12 postimmunization, the treatment groups received intraperitoneal injections of 2 × 106 MSCs. Daily clinical and weight assessments were performed, and on Day 25, the mice were euthanized. At the end of the period, brain histological analysis was conducted to quantify lymphocyte infiltration. T-cell characteristics were determined using enzyme-linked immunosorbent assay and Real-time polymerase chain reaction (RT-PCR). The assessment of transcription factor expression levels in the CNS was also performed using RT-PCR. Compared to the control group, both the allogeneic (ALO) and syngeneic (SYN) groups demonstrated significantly reduced disease progression. The maximum clinical scores for the control, ALO, and SYN groups were 4.4 ± 0.1, 2.4 ± 0.2, and 2.1 ± 0.2, respectively (ALO and SYN vs. Control: p < .001). In comparison to the control group, histological studies demonstrated that the allogeneic and syngeneic groups had less lymphocytic infiltration (ALO: 1.4 ± 0.1, SYN: 1.2 ± 0.2, and control: 2.8 ± 0.15; p < .001) and demyelination (ALO: 1.2 ± 0.15, SYN: 1.1 ± 0.1 and control: 2.9 ± 0.1, p < .001). ALO and SYN groups had lower expression of Th1 and Th17 cytokines and transcription factors (IFN-γ: 0.067, 0.051; STAT4: 0.189, 0.162; T-bet: 0.175, 0.163; IL-17: 0.074, 0.061; STAT3: 0.271, 0.253; ROR-γt: 0.163, 0.149, respectively) compared to the control group on Day 25 following EAE induction. Additionally, ALO and SYN groups compared to the control group, expressed more Th2 and Treg cytokines and transcription factors (IL-4: 4.25, 4.63; STAT6: 2.78, 2.96; GATA3: 2.91, 3.08; IL-27: 2.32, 2.46, IL-33: 2.71, 2.85; TGF-ß: 4.8, 5.05; IL-10: 4.71, 4.93; CTLA-4: 7.72, 7.95; PD1: 4.12,4.35; Foxp3: 3.82,4.08, respectively). This research demonstrated that MSCs possess the potential to be a therapeutic option for MS and related CNS inflammatory disorders. Their immunomodulatory properties, coupled with the observed reductions in disease severity, lymphocytic infiltration, and demyelination, indicate that MSCs could play a crucial role in altering the course of MS by mitigating inflammatory immune responses and promoting regulatory immune processes. These findings open up new possibilities for the development of MSC-based therapies for MS, and further investigation and clinical trials may be warranted to explore their efficacy and safety in human patients.
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Encefalomielitis Autoinmune Experimental , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Ratones Endogámicos C57BL , Linfocitos T Reguladores , Células TH1 , Células Th17 , Células Th2 , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/inmunología , Femenino , Células Madre Mesenquimatosas/inmunología , Células Madre Mesenquimatosas/metabolismo , Trasplante de Células Madre Mesenquimatosas/métodos , Células TH1/inmunología , Ratones , Células Th17/inmunología , Linfocitos T Reguladores/inmunología , Células Th2/inmunología , Tejido Adiposo/citología , Citocinas/metabolismoRESUMEN
Multiple sclerosis (MS) is an inflammatory demyelinating disorder of the central nervous system. Recent research has revealed that mesenchymal stem cell-derived extracellular vesicles (MSC-EVs), containing specific miRNAs, possess immunomodulatory properties and have demonstrated therapeutic potential in the treatment of MS. This study aimed to investigate the role MSC-EVs, containing microRNA-181a-5p (miR-181a-5p) in both experimental autoimmune encephalomyelitis (EAE), an established animal model of MS, and lipopolysaccharide-stimulated BV2 microglia. We evaluated clinical symptoms and inflammatory responses in EAE mice following intrathecal injections of MSC-EVs. MSC-EVs containing miR-181a-5p were co-cultured with microglia to explore their impact on inflammation and cell pyroptosis. We validated the interaction between miR-181a-5p and its downstream regulators and conducted in vivo verification by injecting manipulated EVs containing miR-181a-5p into EAE mice. Our results demonstrated that MSC-EVs, containing miR-181a-5p reduced the clinical symptoms of EAE mice. Furthermore, we observed downregulation of miR-181a-5p in EAE model mice, and its expression was restored after treatment with MSC-EVs, which corresponded to suppressed microglial inflammation and pyroptosis. Additionally, EVs containing miR-181a-5p mitigated spinal cord injury and demyelination in EAE mice. Mechanistically, ubiquitin-specific protease 15 (USP15) exhibited high expression in EAE mice, and miR-181a-5p was specifically targeted and bound to USP15, thereby regulating the RelA/NEK7 axis. In conclusion, MSC-EVs containing miR-181a-5p inhibit microglial inflammation and pyroptosis through the USP15-mediated RelA/NEK7 axis, thus alleviating the clinical symptoms of EAE. These findings present a potential therapeutic approach for the treatment of MS.
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Encefalomielitis Autoinmune Experimental , Vesículas Extracelulares , Ratones Endogámicos C57BL , MicroARNs , Microglía , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/inmunología , MicroARNs/genética , MicroARNs/metabolismo , Vesículas Extracelulares/metabolismo , Ratones , Microglía/metabolismo , Femenino , Células Madre Mesenquimatosas/metabolismo , Piroptosis , Línea Celular , Esclerosis Múltiple/terapia , Humanos , Modelos Animales de Enfermedad , Lipopolisacáridos , Enfermedades Desmielinizantes/terapiaRESUMEN
Multiple sclerosis (MS) is an autoimmune disease characterized by demyelination of the central nervous system (CNS). Autologous hematopoietic cell transplantation (HCT) shows promising benefits for relapsing-remitting MS in open-label clinical studies, but the cellular mechanisms underlying its therapeutic effects remain unclear. Using single-nucleus RNA sequencing, we identify a reactive myeloid cell state in chronic experimental autoimmune encephalitis (EAE) associated with neuroprotection and immune suppression. HCT in EAE mice results in an increase of the neuroprotective myeloid state, improvement of neurological deficits, reduced number of demyelinated lesions, decreased number of effector T cells and amelioration of reactive astrogliosis. Enhancing myeloid cell incorporation after a modified HCT further improved these neuroprotective effects. These data suggest that myeloid cell manipulation or replacement may be an effective therapeutic strategy for chronic inflammatory conditions of the CNS.
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Encefalomielitis Autoinmune Experimental , Ratones Endogámicos C57BL , Células Mieloides , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/patología , Ratones , Femenino , Trasplante de Células Madre Hematopoyéticas/métodos , Neuroprotección/fisiologíaRESUMEN
The clinical effect of human Mesenchymal stem cells (hMSCs) transplanted into EAE mice/MS patients is short lived due to poor survival of the transplanted cells. Since Granagard, a nanoformulation of pomegranate seed oil, extended the presence of Neuronal Stem cells transplanted into CJD mice brains, we tested whether this safe food supplement can also elongate the survival of hMSCs transplanted into EAE mice. Indeed, pathological studies 60 days post transplantation identified human cells only in brains of Granagard treated mice, concomitant with increased clinical activity. We conclude that Granagard may prolong the activity of stem cell transplantation in neurological diseases.
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Encefalomielitis Autoinmune Experimental , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Esclerosis Múltiple , Humanos , Animales , Ratones , Esclerosis Múltiple/terapia , Esclerosis Múltiple/patología , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/patología , Encéfalo/patología , Trasplante de Células Madre , Factores Inmunológicos , Células Madre Mesenquimatosas/patología , Células Madre Mesenquimatosas/fisiologíaRESUMEN
BACKGROUND AIMS: Adipose mesenchymal stem cells (ASCs) represent a promising therapeutic approach in inflammatory neurological disorders, including multiple sclerosis (MS). Recent lines of evidence indicate that most biological activities of ASCs are mediated by the delivery of soluble factors enclosed in extracellular vesicles (EVs). Indeed, we have previously demonstrated that small EVs derived from ASCs (ASC-EVs) ameliorate experimental autoimmune encephalomyelitis (EAE), a murine model of MS. The precise mechanisms and molecular/cellular target of EVs during EAE are still unknown. METHODS: To investigate the homing of ASC-EVs, we intravenously injected small EVs loaded with ultra-small superparamagnetic iron oxide nanoparticles (USPIO) at disease onset in EAE-induced C57Bl/6J mice. Histochemical analysis and transmission electron microscopy were carried out 48 h after EV treatment. Moreover, to assess the cellular target of EVs, flow cytometry on cells extracted ex vivo from EAE mouse lymph nodes was performed. RESULTS: Histochemical and ultrastructural analysis showed the presence of labeled EVs in lymph nodes but not in lungs and spinal cord of EAE injected mice. Moreover, we identified the cellular target of EVs in EAE lymph nodes by flow cytometry: ASC-EVs were preferentially located in macrophages, with a consistent amount also noted in dendritic cells and CD4+ T lymphocytes. CONCLUSIONS: This represents the first direct evidence of the privileged localization of ASC-EVs in draining lymph nodes of EAE after systemic injection. These data provide prominent information on the distribution, uptake and retention of ASC-EVs, which may help in the development of EV-based therapy in MS.
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Encefalomielitis Autoinmune Experimental , Vesículas Extracelulares , Células Madre Mesenquimatosas , Esclerosis Múltiple , Ratones , Animales , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/patología , Esclerosis Múltiple/terapia , Esclerosis Múltiple/patología , Ganglios Linfáticos , Ratones Endogámicos C57BLRESUMEN
Beyond the encouraging results and broad clinical applicability of immune checkpoint (ICP) inhibitors in cancer therapy, ICP-based immunotherapies in the context of autoimmune disease, particularly multiple sclerosis (MS), have garnered considerable attention and hold great potential for developing effective therapeutic strategies. Given the well-established immunoregulatory role of ICPs in maintaining a balance between stimulatory and inhibitory signaling pathways to promote immune tolerance to self-antigens, a dysregulated expression pattern of ICPs has been observed in a significant proportion of patients with MS and its animal model called experimental autoimmune encephalomyelitis (EAE), which is associated with autoreactivity towards myelin and neurodegeneration. Consequently, there is a rationale for developing immunotherapeutic strategies to induce inhibitory ICPs while suppressing stimulatory ICPs, including engineering immune cells to overexpress ligands for inhibitory ICP receptors, such as program death-1 (PD-1), or designing fusion proteins, namely abatacept, to bind and inhibit the co-stimulatory pathways involved in overactivated T-cell mediated autoimmunity, and other strategies that will be discussed in-depth in the current review. Video Abstract.
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Encefalomielitis Autoinmune Experimental , Esclerosis Múltiple , Animales , Humanos , Esclerosis Múltiple/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/terapia , Linfocitos T , Inmunoterapia , AutoinmunidadRESUMEN
INTRODUCTION: Clinical and experimental studies highlighted the significant therapeutic role of Mesenchymal stem cells (MSCs) in neurodegenerative diseases. MSCs possess potent immunomodulatory properties by releasing exosomes, which generate a suitable microenvironment. microRNAs (miRNAs), as one of several effective bioactive molecules of exosomes, influence cellular communication and activities in recipient cells. Recent studies revealed that miRNAs could control the progression of multiple sclerosis (MS) via differentiation and function of T helper cells (Th). METHODS: Here, we investigated the therapeutic effects of syngeneic-derived BM-MSC in experimental autoimmune encephalomyelitis (EAE) mouse model of MS by evaluating expression profile of miRNAs, pro- and anti-inflammatory in serum and brain tissues. Three-time scheme groups (6th day, 6th & 12th days, and 12th day, of post-EAE induction) were applied to determine the therapeutic effects of intraperitoneally received 1*106 of BM-MSCs. RESULTS: The expression levels of mature isoforms of miR-193, miR-146a, miR-155, miR-21, and miR-326 showed that BM-MSCs treatment attenuated the EAE clinical score and reduced clinical inflammation as well as demyelination. The improved neurological functional outcome associated with enhanced expression of miR-193 and miR-146a, but decreased expression levels of miR-155, miR-21, and miR-326 were followed by suppressing effects on Th1/Th17 immune responses (reduced levels of IFN-γand IL-17 cytokine expression) and induction of Treg cells, immunoregulatory responses (increase of IL-10, TGF-ß, and IL-4) in treatment groups. CONCLUSION: Our findings suggest that BM-MSCs administration might change expression patterns of miRNAs and downstream interactions followed by immune system modulation. However, there is a need to carry out future human clinical trials and complementary experiments.
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Encefalomielitis Autoinmune Experimental , Células Madre Mesenquimatosas , MicroARNs , Esclerosis Múltiple , Animales , Ratones , Humanos , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/terapia , MicroARNs/genética , MicroARNs/metabolismo , Esclerosis Múltiple/genética , Esclerosis Múltiple/terapia , Inflamación/metabolismo , Células Madre Mesenquimatosas/metabolismo , Ratones Endogámicos C57BL , Células de la Médula ÓseaRESUMEN
Inducing antigen-specific tolerance during an established immune response typically requires non-specific immunosuppressive signalling molecules. Hence, standard treatments for autoimmunity trigger global immunosuppression. Here we show that established antigen-specific responses in effector T cells and memory T cells can be suppressed by a polymer glycosylated with N-acetylgalactosamine (pGal) and conjugated to the antigen via a self-immolative linker that allows for the dissociation of the antigen on endocytosis and its presentation in the immunoregulatory environment. We show that pGal-antigen therapy induces antigen-specific tolerance in a mouse model of experimental autoimmune encephalomyelitis (with programmed cell-death-1 and the co-inhibitory ligand CD276 driving the tolerogenic responses), as well as the suppression of antigen-specific responses to vaccination against a DNA-based simian immunodeficiency virus in non-human primates. Our findings show that pGal-antigen therapy invokes mechanisms of immune tolerance to resolve antigen-specific inflammatory T-cell responses and suggest that the therapy may be applicable across autoimmune diseases.
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Encefalomielitis Autoinmune Experimental , Tolerancia Inmunológica , Animales , Ratones , Autoinmunidad , Glicosilación , Acetilgalactosamina , Encefalomielitis Autoinmune Experimental/terapiaRESUMEN
Dysregulation of the interleukin-1 (IL-1) pathway leads to immune diseases that can result in chronic tissue and organ inflammation. Although IL-1 blockade has shown promise in ameliorating these symptoms and improving patients' quality of life, there is an urgent need for more effective, long-lasting treatments. We developed a lentivirus (LV)-mediated gene transfer strategy using transplanted autologous hematopoietic stem/progenitor cells (HSPCs) as a source of IL-1 receptor antagonist (IL-1RA) for systemic delivery to tissues and organs. Transplantation of mouse and human HSPCs transduced with an IL-1RA-encoding LV ensured stable IL-1RA production while maintaining the clonogenic and differentiation capacities of HSPCs in vivo. We examined the efficacy of cell-mediated IL-1RA delivery in three models of IL-1-dependent inflammation, for which treatment hindered neutrophil recruitment in an inducible model of gout, prevented systemic and multi-tissue inflammation in a genetic model of cryopyrin-associated periodic syndromes, and reduced disease severity in an experimental autoimmune encephalomyelitis model of multiple sclerosis. Our findings demonstrate HSPC-mediated IL-1RA delivery as a potential therapeutic modality that can be exploited to suppress tissue and organ inflammation in diverse immune-related diseases involving IL-1-driven inflammation.
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Encefalomielitis Autoinmune Experimental , Proteína Antagonista del Receptor de Interleucina 1 , Animales , Humanos , Encefalomielitis Autoinmune Experimental/terapia , Inflamación/terapia , Interleucina-1 , Lentivirus , Calidad de Vida , RatonesRESUMEN
Remyelination and neurodegeneration prevention mitigate disability in Multiple Sclerosis (MS). We have shown acute intermittent hypoxia (AIH) is a novel, non-invasive and effective therapy for peripheral nerve repair, including remyelination. Thus, we posited AIH would improve repair following CNS demyelination and address the paucity of MS repair treatments. AIH's capacity to enhance intrinsic repair, functional recovery and alter disease course in the experimental autoimmune encephalomyelitis (EAE) model of MS was assessed. EAE was induced by MOG35-55 immunization in C57BL/6 female mice. EAE mice received either AIH (10 cycles-5 min 11% oxygen alternating with 5 min 21% oxygen) or Normoxia (control; 21% oxygen for same duration) once daily for 7d beginning at near peak EAE disease score of 2.5. Mice were followed post-treatment for an additional 7d before assessing histopathology or 14d to examine maintenance of AIH effects. Alterations in histopathological correlates of multiple repair indices were analyzed quantitatively in focally demyelinated ventral lumbar spinal cord areas to assess AIH impacts. AIH begun at near peak disease significantly improved daily clinical scores/functional recovery and associated histopathology relative to Normoxia controls and the former were maintained for at least 14d post-treatment. AIH enhanced correlates of myelination, axon protection and oligodendrocyte precursor cell recruitment to demyelinated areas. AIH also effected a dramatic reduction in inflammation, while polarizing remaining macrophages/microglia toward a pro-repair state. Collectively, this supports a role for AIH as a novel non-invasive therapy to enhance CNS repair and alter disease course following demyelination and holds promise as a neuroregenerative MS strategy.
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Encefalomielitis Autoinmune Experimental , Esclerosis Múltiple , Remielinización , Encefalomielitis Autoinmune Experimental/patología , Encefalomielitis Autoinmune Experimental/terapia , Esclerosis Múltiple/patología , Esclerosis Múltiple/terapia , Animales , Ratones , Ratones Endogámicos C57BL , Anaerobiosis , Oxígeno , FemeninoRESUMEN
Analogs of immunodominant myelin peptides involved in multiple sclerosis (MS: the most common autoimmune disease) have been extensively used to modify the immune response over the progression of the disease. The immunodominant 35-55 epitope of myelin oligodendrocyte glycoprotein (MOG35-55 ) is an autoantigen appearing in MS and stimulates the encephalitogenic T cells, whereas mannan polysaccharide (Saccharomyces cerevisiae) is a carrier toward the mannose receptor of dendritic cells and macrophages. The conjugate of mannan-MOG35-55 has been extensively studied for the inhibition of chronic experimental autoimmune encephalomyelitis (EAE: an animal model of MS) by inducing antigen-specific immune tolerance against the clinical symptoms of EAE in mice. Moreover, it presents a promising approach for the immunotherapy of MS under clinical investigation. In this study, a competitive enzyme-linked immunosorbent assay (ELISA) was developed to detect the MOG35-55 peptide that is conjugated to mannan. Intra- and inter-day assay experiments proved that the proposed ELISA methodology is accurate and reliable and could be used in the following applications: (i) to identify the peptide (antigen) while it is conjugated to mannan and (ii) to adequately address the alterations that the MOG35-55 peptide may undergo when it is bound to mannan during production and stability studies.
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Epítopos Inmunodominantes , Esclerosis Múltiple , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Péptidos , Animales , Ratones , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/terapia , Ensayo de Inmunoadsorción Enzimática , Epítopos Inmunodominantes/análisis , Mananos/química , Ratones Endogámicos C57BL , Glicoproteína Mielina-Oligodendrócito/análisis , Fragmentos de Péptidos/análisis , Esclerosis Múltiple/metabolismo , Esclerosis Múltiple/terapiaRESUMEN
Multiple sclerosis (MS) is a chronic autoimmune disease that affects the central nervous system and is marked by inflammation and damage to the myelin sheath surrounding nerve fibers. Recent studies have highlighted the therapeutic value of exosomes (Exos) obtained from bone marrow mesenchymal stem cells (BMSCs) in MS treatment. These BMSC-Exos contain biologically active molecules that show promising results in preclinical evaluations. The aim of this study was to investigate the mechanism of BMSC-Exos containing miR-23b-3p in both LPS-stimulated BV2 microglia and in experimental autoimmune encephalomyelitis (EAE), an animal model for MS. Exos were isolated from BMSCs, and their effects were evaluated in vitro by co-culturing with BV2 microglia. The interaction between miR-23b-3p and its downstream targets was also explored. The efficacy of BMSC-Exos was further verified in vivo by injecting the Exos into EAE mice. The results showed that BMSC-Exos containing miR-23b-3p reduced microglial pyroptosis in vivo by specifically binding to and suppressing the expression of NEK7. In vivo, BMSC-Exos containing miR-23b-3p alleviated the severity of EAE by decreasing microglial inflammation and pyroptosis via the repression of NEK7. These findings provide new insights into the therapeutic potential of BMSC-Exos containing miR-23b-3p for MS.
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Encefalomielitis Autoinmune Experimental , Células Madre Mesenquimatosas , MicroARNs , Esclerosis Múltiple , Ratones , Animales , Microglía/metabolismo , Encefalomielitis Autoinmune Experimental/terapia , Encefalomielitis Autoinmune Experimental/metabolismo , Piroptosis , Células Madre Mesenquimatosas/metabolismo , Inflamación/metabolismo , Esclerosis Múltiple/terapia , MicroARNs/genética , MicroARNs/metabolismoRESUMEN
Microglia are the resident immune cells of the central nervous system, playing a role in the inflammatory process development and resolution, presenting two main phenotypes, pro-inflammatory M1, and anti-inflammatory M2. Therapies affecting the microglia phenotype may be beneficial in treating inflammatory neurodegenerative diseases. In our experiments, we used the animal multiple sclerosis model, experimental allergic encephalomyelitis (EAE). Rats were treated during the pre- or symptomatic phase of the disease with cyclophosphamide, followed by hematopoietic stem cell transplantation, and with/without post-transplantation cyclophosphamide. Our study aimed to analyze the microglia phenotype in animals subjected to this treatment. The number of M1 cells in the spinal cord, and inducible nitric oxide synthase (iNOS) levels in the brain were similar in all experimental groups. The differences were observed in M2 cells number and arginase 1 (Arg1) levels, which were decreased in EAE animals, and increased after treatment in the symptomatic phase of EAE, and in the pre-symptomatic phase, but only with post-transplantation cyclophosphamide. Analysis of gene expression in the brain showed decreased iNOS expression in EAE animals treated in the symptomatic phase of EAE and no differences in Arg1 expression. Results indicate that treatment applied to experimental animals influences the microglia phenotype, promoting differentiation towards M2 cells.
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Encefalomielitis Autoinmune Experimental , Trasplante de Células Madre Hematopoyéticas , Ratas , Animales , Ratones , Encefalomielitis Autoinmune Experimental/terapia , Microglía/metabolismo , Fenotipo , Ciclofosfamida/uso terapéutico , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: Mesenchymal stem cells (MSCs) modulate immune responses, and their immunomodulatory potential can be enhanced using inflammatory cytokines. Here, the modulatory effects of IFN-γ-licensed MSCs on expression of T cell-related chemokines and chemokine receptors were evaluated using an experimental autoimmune encephalomyelitis (EAE) model. MATERIAL AND METHODS: EAE was induced in 3 groups of C57bl/6 mice and then treated with PBS, MSCs and IFN-γ-treated MSCs. The EAE manifestations were registered daily and finally, the brain and spinal cords were isolated for histopathological and gene expression studies. RESULTS: The clinical scores were lowered in MSCs and IFN-γ-licensed MSCs groups, however, mice treated with IFN-γ-licensed MSCs exhibited lower clinical scores than MSCs-treated mice. Leukocyte inï¬ltration into the brain was reduced after treatment with MSCs or IFN-γ-licensed MSCs compared to untreated group (P<0.05 and P<0.01, respectively). In comparison with untreated EAE mice, treatment with MSCs reduced CCL20 expression (P<0.001) and decreased CXCR3 and CCR6 expression (P<0.02 and P<0.04, respectively). In comparison with untreated EAE mice, treatment with IFN-γ-licensed MSCs reduced CXCL10, CCL17 and CCL20 expression (P<0.05, P<0.05, and P<0.001, respectively) as well as decreased CXCR3 and CCR6 expression (P<0.002 and P<0.02, respectively), whilst promoting expression of CCL22 and its receptor CCR4 (P<0.0001 and P<0.02, respectively). In comparison with MSC-treated group, mice treated with IFN-γ-licensed MSCs exhibited lower CXCL10 and CCR6 expression (P<0.002 and P<0.01, respectively), whereas greater expression of CCL22 and CCR4 (P<0.0001 and P<0.01, respectively). CONCLUSION: Priming the MSC with IFN-γ can be an efficient approach to enhance the immunomodulatory potential of MSCs.
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Encefalomielitis Autoinmune Experimental , Células Madre Mesenquimatosas , Animales , Ratones , Encefalomielitis Autoinmune Experimental/terapia , Interferón gamma , Receptores de Quimiocina/metabolismo , Receptores de Quimiocina/uso terapéutico , Quimiocinas/metabolismo , Quimiocinas/farmacología , Quimiocinas/uso terapéutico , Linfocitos T , Células Madre Mesenquimatosas/metabolismo , Ratones Endogámicos C57BLRESUMEN
Macrophage M2 (MP2)-based cell therapy is a novel medicinal treatment for animals with Experimental Autoimmune Encephalomyelitis (EAE) as an experimental model of multiple sclerosis (MS). This systematic review and meta-analysis study was designed to assess the overall therapeutic effects of MP2 cell therapy on Clinical Score and motor impairment in EAE-induced animals. All experiments on MP2 cell therapy in animals with EAE were gathered (by October 2, 2022) from English (PubMed, Scopus, WoS, Science Direct, and ISC) and Persian (MagIran and SID) databases. The searching strategy was designed using "Experimental Autoimmune Encephalomyelitis," "Multiple Sclerosis," and "Macrophage M2" keywords. Following primary and secondary screenings, eligible papers were selected based on the PRISMA 2020 guideline, and the study quality was assessed using the Animal Research: Reporting of In Vivo Experiments (ARRIVE) checklist. The difference in means of Clinical Score (score 0-5) as the effect size (ES) was analyzed based on the random effect model (CMA software, v.2). Subgrouping (EAE phases of Onset, Peak, and Recovery) was applied, and I2 index was used to assess the heterogeneity index. Publication bias and sensitivity indices were also evaluated. P < 0.05 was considered significant, and the confidence interval (CI) was determined 95%. Among 22 gathered papers, medium to high quality studies were selected for meta-analysis. Difference in means, P value, and I2 for Onset, Peak, and Recovery phases were 0.082 (CI95%: -0.323-0.159, P value: 0.504, I2 : 67.961%), -0.606 (CI95%: -1.518 to -0.305, P value: 0.192, I2 : 96.070%), and -1.103 (CI95%: -1.390 to -0.816, P value: 0.000, I2 : 30.880%), respectively and Overall Effect was found -0.509 (CI95%: -0.689 to -0.328, P value < 0.001). Also, P value (two-tailed) indices for publication bias were 0.366 and 0.583 for Egger's regression intercept and Begg rank correlation, respectively. The P value for sensitivity was detected 0.003. Cell therapy procedure using MP2 can potentially alleviate the Clinical Scores Index and correct the motor defects in Recovery phase of EAE animals. In healthy mice, the brain and myelin surrounding neurons are in a healthy and physiological state (1). To evaluate MS in humans, it is necessary to model this type of disease in animals using EAE procedure through subcutaneous injection of CFA, MOG35-55 , MT, and Pert. Thus, inflammation and autoimmunity occur, which finally lead to myelin destruction and motor symptoms (2). By aspiration of progenitor cells available in bone marrow, the MP2 can be isolated and cultured. By activation of these types of cells, a rich collection of MP2 can be prepared for the cell-therapy process (3). After injection through the tail vein or intra-peritoneal procedure, these cells can be located in CNS through crossing from the BBB. They begin their anti-inflammatory activities and help repair the damaged myelin (4). Eventually, the clinical symptoms can be modified considerably, and the animal motor function improves (5). CFA, complete Freund's adjuvant; MOG35-55 , myelin oligodendrocyte glycoprotein; MT, Mycobacterium tuberculosis; Pert, pertussis; EAE, Experimental Autoimmune Encephalomyelitis; BM, bone marrow; MP2, macrophage M2; and BBB, blood brain barrier.