Evaluation of co-therapy with melatonin and methylprednisolone in Experimental Autoimmune Encephalomyelitis (EAE) [Dataset]
The dataset includes the raw data from the clinical score and flow cytometry analyzes carried out in the work titled " Melatonin synergistically potentiates the effect of methylprednisolone on reducing neuroinflammation in the experimental autoimmune encephalomyelitis mouse model of multiple sc...
| Autores: | , , , , , , , |
|---|---|
| Formato: | conjunto de datos |
| Fecha de publicación: | 2024 |
| País: | España |
| Recursos: | Universidad de Sevilla (US) |
| Repositorio: | idUS. Depósito de Investigación de la Universidad de Sevilla |
| OAI Identifier: | oai:idus.us.es:11441/161856 |
| Acesso em linha: | https://hdl.handle.net/11441/161856 https://doi.org/10.12795/11441/161856 |
| Access Level: | acceso abierto |
| Palavra-chave: | melatonina esclerosis múltiple metilprednisolona EAE melatonin multiple sclerosis methylprednisolone |
| Resumo: | The dataset includes the raw data from the clinical score and flow cytometry analyzes carried out in the work titled " Melatonin synergistically potentiates the effect of methylprednisolone on reducing neuroinflammation in the experimental autoimmune encephalomyelitis mouse model of multiple sclerosis". This study shows the protective synergistic effect of co-treatment with melatonin and methylprednisolone on reducing the severity of EAE by decreasing CD4+ lymphocytes, B cells, macrophages and dendritic cells in the CNS, as well as modulating the population of infiltrated T and B cells toward regulatory phenotypes to the detriment of pro-inflammatory effector functions. In addition, treatment with melatonin from the clinical onset of EAE improves the natural course of the EAE and the response to a subsequent treatment with methylprednisolone in a later relapse of the disease. Eight-week-old female C57BL/6N mice were immunized immunization with 100 μg of MOG35–55 (Cambridge Research Biochemicals, Cleveland) emulsified in CFA (Sigma) containing 50 μg of heat-killed Mycobacterium tuberculosis (H37Ra, ATCC 25177) by subcutaneous injection in both hind legs and two doses of intraperitoneal pertussis toxin (200 ng/day) (List Labs, California) on days 0 and 2 post-induction. Animals were randomly divided to receive melatonin at a concentration of 80 mg/kg and/or methylprednisolone at a concentration of 40 or 160 mg/kg in different treatment regimens. Mice were sacrificed at the peak of the disease (day 15 after induction) and after perfusion, the CNS was collected, homogenized and enzymatically dissociated with 1.87 mg/ml of collagenase IV (Worthington) and 0.25 mg/ml of DNase I (AppliChem) for 35 min at 37°C to obtain a suspension of single cells. Subsequently, a 37%:70% discontinous percoll gradient was carried out to isolate CNS-infiltrating mononuclear cells. To assess the profile of infiltrated immune cells in the CNS, cells were stained for the following antibodies against surface markers: CD45, CD4, CD8α, CD19, CD11b, CD11c, CD44, CD62L, B220, CD138, PD-1 (CD279), CTLA-4 (CD152), FAS (CD95), and CD25. To identify Treg and analyze intracellular production of TNF, IFN-γ and IL-10, after surface staining, cells were fixed and permeabilized using the The dataset includes the raw data from the clinical score and flow cytometry analyzes carried out in the work titled " Melatonin synergistically potentiates the effect of methylprednisolone on reducing neuroinflammation in the experimental autoimmune encephalomyelitis mouse model of multiple sclerosis". This study shows the protective synergistic effect of co-treatment with melatonin and methylprednisolone on reducing the severity of EAE by decreasing CD4+ lymphocytes, B cells, macrophages and dendritic cells in the CNS, as well as modulating the population of infiltrated T and B cells toward regulatory phenotypes to the detriment of pro-inflammatory effector functions. In addition, treatment with melatonin from the clinical onset of EAE improves the natural course of the EAE and the response to a subsequent treatment with methylprednisolone in a later relapse of the disease. Eight-week-old female C57BL/6N mice were immunized immunization with 100 μg of MOG35–55 (Cambridge Research Biochemicals, Cleveland) emulsified in CFA (Sigma) containing 50 μg of heat-killed Mycobacterium tuberculosis (H37Ra, ATCC 25177) by subcutaneous injection in both hind legs and two doses of intraperitoneal pertussis toxin (200 ng/day) (List Labs, California) on days 0 and 2 post-induction. Animals were randomly divided to receive melatonin at a concentration of 80 mg/kg and/or methylprednisolone at a concentration of 40 or 160 mg/kg in different treatment regimens. Mice were sacrificed at the peak of the disease (day 15 after induction) and after perfusion, the CNS was collected, homogenized and enzymatically dissociated with 1.87 mg/ml of collagenase IV (Worthington) and 0.25 mg/ml of DNase I (AppliChem) for 35 min at 37°C to obtain a suspension of single cells. Subsequently, a 37%:70% discontinous percoll gradient was carried out to isolate CNS-infiltrating mononuclear cells. To assess the profile of infiltrated immune cells in the CNS, cells were stained for the following antibodies against surface markers: CD45, CD4, CD8α, CD19, CD11b, CD11c, CD44, CD62L, B220, CD138, PD-1 (CD279), CTLA-4 (CD152), FAS (CD95), and CD25. To identify Treg and analyze intracellular production of TNF, IFN-γ and IL-10, after surface staining, cells were fixed and permeabilized using the FoxP3/transcription factor staining buffer set (eBioscience) and stained with anti-FoxP3 or anti-TNF, -IFN-γ and -IL-10, respectively. The intracellular production of cytokines was carried out in cells cultured at 2.5 × 106 cells/ml with RPMI 1640 supplemented with 5% fetal bovine serum, 1% L-glutamine and 1% of penicillin/streptomycin incubated in the presence or absence of Phorbol-Myristate-Acetate and Ionomycin (PMAI, Sigma) with brefeldin A (eBioscience) for 5 h. Dead cells were excluded from analyses using the LIVE/DEAD® Fixable Dead Cell Stain Kit (Invitrogen). FACS analysis was performed using a Cytek Aurora spectral cytometer, and data were analyzed using FlowJo software (Treestar). |
|---|