Huntington’s disease (HD) is caused by the expression of a mutated variant of Huntingtin (mHtt), which results in the complex pathology characterized by a defective function of the nervous system and altered inflammatory responses. While the neuronal effects of mHtt expression have been extensively studied, its effects on the physiology of immune cells have not been fully described. Mast cells (MCs) are unique tissue-resident immune cells whose activation has been linked to protective responses against parasites and bacteria, but also to deleterious inflammatory allergic reactions and, recently, to neurodegenerative diseases. Bone marrow-derived mast cells (BMMCs) were obtained from wild-type (WT-) and mHtt-expressing (R6/1) mice to evaluate the main activation parameters triggered by the high-affinity IgE receptor (FcεRI) and the Toll-like receptor (TLR) 4. Degranulation was assessed by measuring the secretion of β-hexosaminidase, MAP kinase activation was detected by Western blot, and cytokine production was determined by RT-PCR and ELISA. TLR-4 receptor and Htt vesicular trafficking was analyzed by confocal microscopy. In vivo, MC-deficient mice (c-KitWsh/Wsh) were intraperitonally reconstituted with WT or R6/1 BMMCs and the TLR4-induced production of the tumor necrosis factor (TNF) was determined by ELISA. A survival curve of mice treated with a sub-lethal dose of bacterial lipopolysaccharide (LPS) was constructed. R6/1 BMMCs showed normal β-hexosaminidase release levels in response to FcεRI, but lower cytokine production upon LPS stimulus. Impaired TLR4-induced TNF production was associated to the lack of intracellular dynamin-dependent TLR-4 receptor trafficking to perinuclear regions in BMMCs, a diminished ERK1/2 and ELK-1 phosphorylation, and a decrease in c-fos and TNF mRNA accumulation. R6/1 BMMCs also failed to produce TLR4-induced anti-inflammatory cytokines (like IL-10 and TGF-β). The detected defects were also observed in vivo, in a MCs-dependent model of endotoxemia. R6/1 and c-KitWsh/Wsh mice reconstituted with R6/1 BMMCs showed a decreased TLR4-induced TNF production and lower survival rates to LPS challenge than WT mice. Our data show that mHtt expression causes an impaired production of pro- and anti-inflammatory mediators triggered by TLR-4 receptor in MCs in vitro and in vivo, which could contribute to the aberrant immunophenotype observed in HD.

中文翻译：

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突变的亨廷顿蛋白影响肥大细胞中的Toll样受体4细胞内运输和细胞因子产生

亨廷顿舞蹈病（HD）是由亨廷顿舞蹈症（mHtt）突变变体的表达引起的，这导致了复杂的病理，其特征是神经系统功能缺陷和炎症反应改变。虽然已经广泛研究了mHtt表达的神经元作用，但尚未充分描述其对免疫细胞生理学的作用。肥大细胞（MCs）是独特的组织驻留免疫细胞，其激活与针对寄生虫和细菌的保护性反应有关，但也与有害的炎症性过敏反应有关，最近与神经退行性疾病有关。从野生型（WT-）和表达mHtt的（R6 / 1）小鼠中获得骨髓衍生的肥大细胞（BMMC），以评估由高亲和力IgE受体（FcεRI）和Toll-像受体（TLR）一样。4.通过测量β-己糖胺酶的分泌来评估颗粒形成，通过Western blot检测MAP激酶的活化，并通过RT-PCR和ELISA测定细胞因子的产生。通过共聚焦显微镜分析TLR-4受体和Htt囊泡运输。在体内，用WT或R6 / 1 BMMC腹膜内重建MC缺陷小鼠（c-KitWsh / Wsh），并通过ELISA测定TLR4诱导的肿瘤坏死因子（TNF）的产生。绘制了用亚致死剂量的细菌脂多糖（LPS）处理的小鼠的生存曲线。R6 / 1 BMMC响应FcεRI表现出正常的β-己糖胺酶释放水平，但LPS刺激后细胞因子生成降低。受损的TLR4诱导的TNF产生与缺乏向BMMC的核周区域的细胞内动力依赖性TLR-4受体运输，ERK1 / 2和ELK-1磷酸化减少以及c-fos和TNF mRNA积累减少有关。R6 / 1 BMMC也未能产生TLR4诱导的抗炎细胞因子（如IL-10和TGF-β）。在体内依赖于内毒素血症的MCs模型中，还可以在体内观察到检测到的缺陷。与WT小鼠相比，用R6 / 1 BMMC重组的R6 / 1和c-KitWsh / Wsh小鼠显示TLR4诱导的TNF生成减少，对LPS攻击的存活率降低。