大麻素内酰胺对单核细胞来源的朗格汉斯细胞的调节作用：对免疫稳态和皮肤炎症的影响。

INTRODUCTION: The endocannabinoid system (ECS), named after the chemical compounds found in the cannabis plant, is a regulatory network of neurotransmitters, receptors, and enzymes that plays crucial roles in skin health and disease. Endogenous ligands of the ECS, called endocannabinoids, have proven to be important regulators of immune responses. One of the most prevalent endocannabinoids, arachidonoylethanolamide (also known as anandamide), is known for its anti-inflammatory effects. Langerhans cells (LCs) are the sole antigen-presenting cells present in the human epidermis. They serve as the first line of defense against pathogens and are essential for the skin's specific immune responses and play a critical role in maintaining tissue homeostasis; however, little is known about the effect of endocannabinoids on these cells. Our research aimed to provide the connection between monocyte-derived Langerhans cells (moLCs) and the ECS, shedding light on their collaborative roles in immune homeostasis and inflammation. METHODS: Human monocytes were differentiated into moLCs using established protocols. Anandamide was applied during the differentiation process to test its effect on the viability, marker expression, and cytokine production of the cells, as well as in short term treatments for intracellular calcium measurement. TLR ligands applied after the differentiation protocol were used to activate moLCs. The impact of anandamide on the functionality of moLCs was further assessed using differential gene expression analysis of bulk RNA-Seq data, moLC-T cell cocultures, while ELISpot was employed to determine polarization of T cells activated in the aforementioned cocultures. RESULTS: Anandamide did not significantly affect the viability of moLCs up to 10 µM. When applied during the differentiation process it had only a negligible effect on CD207 expression, the prototypic marker of LCs; however, there was an observed reduction in CD1a expression by moLCs. Anandamide had no significant effects on the maturation status of moLCs, nor did it affect the maturation induced by TLR3 and TLR7/8 agonists. MoLCs differentiated in the presence of anandamide did however show decreased production of CXCL8, IL-6, IL-10 and IL-12 cytokines induced by TLR3 and TLR7/8 activation. Anandamide-treated moLCs showed an increased capability to activate naïve T cells; however, not to the level seen with combined TLR agonism. RNA sequencing analysis of moLCs differentiated with anandamide showed modest changes compared to control cells but did reveal an inhibitory effect on oxidative phosphorylation specifically in activated moLCs. Anandamide also promoted the polarization of naïve T cells towards a Th1 phenotype. DISCUSSION: Our results show that anandamide has nuanced effects on the differentiation, maturation, cytokine secretion, metabolism and function of activated moLCs. Among these changes the decrease in CD1a expression on moLCs holds promise to selectively dampen inflammation induced by CD1a restricted T cells, which have been implicated as drivers of inflammation in common inflammatory skin conditions such as psoriasis, atopic dermatitis and contact dermatitis.

简介：内源性大麻素系统（ECS）以大麻植物中发现的化学物质命名，是一种神经递质、受体和酶的调节网络，在皮肤健康和疾病中起着至关重要的作用。ECS 的内源性配体，称为内源性大麻素，已被证明是免疫反应的重要调节剂。最常见的内源性大麻素之一，花生四烯酸乙醇酰胺（也称为大麻素），以其抗炎作用而闻名。朗格汉斯细胞（LCs）是人类表皮中唯一的抗原呈递细胞。它们是抵御病原体的第一道防线，对于皮肤的特异性免疫反应至关重要，并在维持组织内稳态方面发挥着关键作用；然而，关于内源性大麻素对这些细胞的影响知之甚少。我们的研究旨在提供单核细胞衍生的朗格汉斯细胞（moLCs）与 ECS 之间的联系，阐明它们在免疫稳态和炎症中的协同作用。

方法：使用已建立的方案将人单核细胞分化为 moLCs。在分化过程中应用大麻素，以测试其对细胞活力、标志物表达和细胞因子产生的影响，以及进行短期细胞内钙测量处理。在分化方案后应用 TLR 配体以激活 moLCs。使用批量 RNA-Seq 数据的差异基因表达分析、moLC-T 细胞共培养进一步评估大麻素对 moLC 功能的影响，同时使用 ELISpot 确定在上述共培养中激活的 T 细胞的极化。

结果：大麻素的浓度高达 10µM 时，对 moLCs 的活力没有显著影响。在分化过程中应用时，它对 LC 的典型标志物 CD207 的表达只有轻微影响；然而，moLCs 中 CD1a 的表达减少。大麻素对 moLCs 的成熟状态没有显著影响，也没有影响 TLR3 和 TLR7/8 激动剂诱导的成熟。然而，在大麻素存在下分化的 moLCs 显示出 TLR3 和 TLR7/8 激活诱导的 CXCL8、IL-6、IL-10 和 IL-12 细胞因子产生减少。大麻素处理的 moLCs 显示出激活幼稚 T 细胞的能力增加；然而，与 TLR 激动剂联合使用的水平相比并没有增加。用大麻素分化的 moLCs 的 RNA 测序分析与对照细胞相比显示出适度的变化，但在激活的 moLCs 中确实显示出氧化磷酸化的抑制作用。大麻素还促进了幼稚 T 细胞向 Th1 表型的极化。

讨论：我们的结果表明，大麻素对内源性大麻素系统的分化、成熟、细胞因子分泌、代谢和功能有细微的影响。在这些变化中，moLCs 上 CD1a 表达的减少有望选择性地抑制由 CD1a 限制的 T 细胞诱导的炎症，CD1a 限制的 T 细胞已被认为是常见炎症性皮肤疾病（如银屑病、特应性皮炎和接触性皮炎）中炎症的驱动因素。