Transcriptomic patterns of dermal Schwann cell reactivity in neuropathic itch and pain

神经性瘙痒和疼痛中真皮雪旺细胞反应性的转录组模式

• 批准号: 399376570
• 负责人: Professor Dr. Frank Birklein
• 金额: --
• 依托单位: Neurologische Klinik
• 依托单位国家: 德国
• 项目类别: Research Units
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/399376570?language=en
• 关键词: Transcriptomic; patterns; dermal; Schwann; cell

Neuropathic pruritus leads to significant impairment of life quality. During the first funding period, we recruited 245 patients with polyneuropathies (PNP): 24 reporting itch as only symptom and 58 reporting itch and pain. We defined a PNP subgroup characterized by high pain ratings upon electrical C fiber stimulation and mechanical hyperalgesia. In skin biopsies, we noted a pronounced loss of dermal nerve fibers paralleled by a strong branching of remaining C fibers in patients with itch. By RNA-sequencing (RNA-seq) and quantitative PCR, we found increased expression of IFI44L in skin biopsy samples from patients with itch only. We showed feasibility of subepidermal Schwann cell (SC) type mapping by immunohistochemistry. Utilizing single-nucleus RNA-seq (snRNA-seq), we could characterize SC subtypes and decode their subtype-specific transcriptomic signatures. Notably, IFI44L is a direct target of the microRNA let-7d, which we previously found to be increased in skin from patients with chronic pain, and let-7 represses netrin expression in SCs. Given that let-7 family members are strongly expressed in SCs, we postulate that SC expression of let-7-IFI44L-netrin pathway members might mediate itch and pain in a subgroup of PNP patients. Thus, we will investigate SC morphology and protein expression in itch and pain, aiming at a deeper understanding of SC related pathways in the skin of well-defined subgroups of patients. PNP patients with either pain or itch, will be recruited and carefully assessed by clinical measurements. For comparison, we will include patients with brachioradial pruritus and receive skin samples of patients with psoriasis from collaborating project #5. We will then identify and characterize groups of PNP patients with itch, pain and high pain sensitivity upon C fiber stimulation aiming at understanding their psychophysical characteristics and at selecting homogenous samples to be used in high-throughput transcriptomic technologies. Next, oligonucleotide-tagged antibody multiplex snRNA-seq will allow us to analyze cell type-specific gene expression from cryo-preserved skin biopsy samples obtained from large clinical cohorts. By bioinformatic analysis, we will be able to decode the transcriptomic profile of cutaneous SCs between different clinical entities and define molecular traits of homeostatic and reactive SCs. Multiplex RNA in situ mapping will enable us to map back levels of SC subtype reactivity to the underlying skin microenvironment. In summary, work with high-quality frozen skin biopsy samples from well-characterized patients with neuropathic pain and itch in combination with single-cell and spatial transcriptomic tools will allow us to identify novel cell-type specific targets and biomarkers for future interventional studies. We hypothesize that identification of novel SC characteristics relative to itch or pain will help better understand their role in relation to axon pathology and the clinical phenotype.

神经性瘙痒会导致生活质量显着受损。在第一个资助期间，我们招募了 245 名多发性神经病 (PNP) 患者：24 名报告瘙痒为唯一症状，58 名报告瘙痒和疼痛。我们定义了一个 PNP 亚组，其特征是在电 C 纤维刺激和机械痛觉过敏时具有高疼痛等级。在皮肤活检中，我们注意到瘙痒患者的真皮神经纤维明显丧失，同时剩余的 C 纤维也有强烈的分支。通过 RNA 测序 (RNA-seq) 和定量 PCR，我们发现仅瘙痒患者的皮肤活检样本中 IFI44L 的表达增加。我们展示了通过免疫组织化学进行表皮下施万细胞（SC）类型作图的可行性。利用单核 RNA 序列 (snRNA-seq)，我们可以表征 SC 亚型并解码其亚型特异性转录组特征。值得注意的是，IFI44L 是 microRNA let-7d 的直接靶标，我们之前发现慢性疼痛患者皮肤中的 microRNA let-7d 增加，而 let-7 抑制 SC 中的 netrin 表达。鉴于let-7家族成员在SC中强烈表达，我们假设let-7-IFI44L-netrin途径成员的SC表达可能介导PNP患者亚组的瘙痒和疼痛。因此，我们将研究瘙痒和疼痛时的 SC 形态和蛋白表达，旨在更深入地了解明确亚组患者皮肤中 SC 相关通路。将招募患有疼痛或瘙痒的 PNP 患者并通过临床测量进行仔细评估。为了进行比较，我们将纳入肱桡瘙痒症患者，并从合作项目#5 中接收银屑病患者的皮肤样本。然后，我们将识别和表征在 C 纤维刺激后出现瘙痒、疼痛和高疼痛敏感性的 PNP 患者群体，旨在了解他们的心理物理特征并选择用于高通量转录组技术的同质样本。接下来，寡核苷酸标记的抗体多重 snRNA-seq 将使我们能够分析从大型临床队列获得的冷冻保存的皮肤活检样本中的细胞类型特异性基因表达。通过生物信息学分析，我们将能够解码不同临床实体之间皮肤 SC 的转录组学特征，并定义稳态和反应性 SC 的分子特征。多重 RNA 原位作图将使我们能够将 SC 亚型反应水平映射回底层皮肤微环境。总之，使用来自具有明确特征的神经性疼痛和瘙痒患者的高质量冷冻皮肤活检样本，结合单细胞和空间转录组学工具，将使我们能够为未来的介入研究确定新的细胞类型特异性靶点和生物标志物。我们假设识别与瘙痒或疼痛相关的新 SC 特征将有助于更好地理解它们在轴突病理学和临床表型中的作用。