Robat-Jazi Behrouz, Oraei Mona, Bitarafan Sama, Mesbah-Namin Seyed Alireza, Noori-Zadeh Ali, Mansouri Fatemeh, Parastouei Karim, Anissian Ali, Yekaninejad Mir Saeed, Saboor Yaraghi Ali Akbar
Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran..
Iranian Center of Neurological Research, Neuroscience Institute, Imam Khomeini Hospital Complex, Tehran University of Medical Sciences, Tehran, Iran.
Iran J Allergy Asthma Immunol. 2023 Oct 29;22(5):452-467. doi: 10.18502/ijaai.v22i5.13995.
Previous studies noted an imbalance in T helper (Th) 17 and regulatory T cells (Tregs) in experimental autoimmune encephalomyelitis (EAE), a multiple sclerosis animal model. calcitriol, vitamin D's active form, was found to ameliorate EAE symptoms by favoring Tregss over Th17 cells, suggesting immunomodulatory effects. This study aimed to assess calcitriol's impact on EAE manifestations and cytokine profile in mice. In this study, we recruited twenty-eight C57BL/6 mice and divided them into 4 groups: healthy controls, EAE, EAE with calcitriol treatment, and healthy mice with calcitriol treatment. CD4+ T cells were isolated from splenocytes using magnetic-activated cell sorting. Real-time polymerase chain reaction was employed to quantify the genes associated with Th9 cells (i.e., SPI1 encoding PU.1 and IL9 encoding interleukin [IL]-9). Moreover, the levels of IL-17 and transforming growth factor beta (TGF-β) were evaluated through enzyme-linked immunosorbent assay in the supernatant of CD4+ T cell culture stimulated by anti-CD3 and anti-CD28 antibodies for 72 hours. In the supernatant of CD4+ T cell cultures, the levels of interleukin-17 (IL-17) were significantly increased, while the levels of transforming growth factor beta (TGF-β) were decreased in the EAE Group compared to the healthy control group. Calcitriol treatment reversed these changes and attenuated EAE symptoms, as confirmed in hematoxylin and eosin, and luxol fast blue stains. Notably, calcitriol increased IL9 gene expression in both EAE and healthy mice. This study provides further evidence of the anti-inflammatory effects of calcitriol and its role in attenuating EAE.
先前的研究指出，在实验性自身免疫性脑脊髓炎（EAE，一种多发性硬化症动物模型）中，辅助性T细胞（Th）17和调节性T细胞（Tregs）存在失衡。维生素D的活性形式骨化三醇被发现可通过促进Tregs而非Th17细胞来改善EAE症状，提示其具有免疫调节作用。本研究旨在评估骨化三醇对小鼠EAE表现和细胞因子谱的影响。在本研究中，我们招募了28只C57BL/6小鼠，并将它们分为4组：健康对照组、EAE组、骨化三醇治疗的EAE组和骨化三醇治疗的健康小鼠组。使用磁珠分选法从脾细胞中分离出CD4+ T细胞。采用实时聚合酶链反应来定量与Th9细胞相关的基因（即编码PU.1的SPI1和编码白细胞介素[IL]-9的IL9）。此外，通过酶联免疫吸附测定法评估在抗CD3和抗CD28抗体刺激72小时的CD4+ T细胞培养上清液中IL-17和转化生长因子β（TGF-β）的水平。在CD4+ T细胞培养上清液中，与健康对照组相比，EAE组白细胞介素-17（IL-17）水平显著升高，而转化生长因子β（TGF-β）水平降低。苏木精和伊红染色以及Luxol固蓝染色证实，骨化三醇治疗可逆转这些变化并减轻EAE症状。值得注意的是，骨化三醇增加了EAE小鼠和健康小鼠中IL9基因的表达。本研究为骨化三醇的抗炎作用及其在减轻EAE中的作用提供了进一步的证据。
