Supplementary Material for: The P2X7 Receptor Is Overexpressed in the Lesional Skin of Subjects Affected by Hidradenitis Suppurativa: A Preliminary Study

Background: P2X receptors (P2XRs) are plasma membrane channels involved in the modulation of immune responses. The role of the P2X7 receptor (P2X7R) has never been investigated in hidradenitis suppurativa (HS), which is a recurrent skin disease characterized by inflammatory nodules, scarring, and suppuration. Objective: Our aim was to investigate by immunohistochemistry (IHC) P2X7R, NLRP3 (NOD-like receptor family, pyrin domain-containing 3), and interleukin-1β (IL-1β) expression in HS lesions compared to healthy control (HC) skin. Method: The intensity of IHC immunostaining was semi-quantitatively graded for keratinocytes, neutrophils, lymphocytes, and monocytes. Statistical significance was assessed by the Mann-Whitney U test, Cohen’s κ coefficient, and χ2 test. Results: A total of 59 samples, 31 from HS and 28 from HC, were collected and analysed. In skin keratinocytes, lymphocytes, and monocytes, but not in neutrophils, P2X7R and NLRP3 protein expression was significantly increased in HS versus the HC group. IL-1β protein expression was also higher in HS versus the HC group both in skin keratinocytes and in the inflammatory infiltrate. Cohen’s κ correlation coefficients for the expression of P2X7R versus NLRP3 or IL-1β in skin keratinocytes were significant (κ = 0.43 and 0.34, respectively). The same association between P2X7R and NLRP3 or IL-1β was confirmed by χ2 tests. Conclusion: P2X7R, NLRP3, and IL-1β are overexpressed, and therefore the entire P2X7R/NLRP3/IL-1β pro-inflammatory axis is likely overactive in the skin of HS patients. This observation might provide clues to the pathogenesis of this disease and suggest novel therapies and markers of disease activity.背景：P2X受体（P2XRs）是参与免疫应答调控的质膜离子通道。此前尚无针对P2X7受体（P2X7R）在化脓性汗腺炎（hidradenitis suppurativa, HS）中作用的研究——HS是一种以炎性结节、瘢痕形成及化脓为特征的复发性皮肤疾病。 目的：本研究旨在通过免疫组化（immunohistochemistry, IHC）技术，对比化脓性汗腺炎皮损与健康对照（healthy control, HC）皮肤中P2X7R、NLRP3（核苷酸结合寡聚化结构域样受体家族，含pyrin结构域3）以及白细胞介素-1β（IL-1β）的表达情况。 方法：对角质形成细胞、中性粒细胞、淋巴细胞与单核细胞的免疫组化染色强度进行半定量分级。统计学显著性分析采用曼-惠特尼U检验、科恩κ系数以及卡方（χ²）检验。 结果：本研究共收集并分析59份皮肤样本，其中化脓性汗腺炎组31份，健康对照组28份。在皮肤角质形成细胞、淋巴细胞及单核细胞中（中性粒细胞未观察到显著变化），化脓性汗腺炎组的P2X7R与NLRP3蛋白表达水平较健康对照组显著升高。相较于健康对照组，化脓性汗腺炎组的IL-1β蛋白在皮肤角质形成细胞及炎症浸润区域的表达水平同样更高。皮肤角质形成细胞中，P2X7R与NLRP3或IL-1β的表达的科恩κ相关系数具有统计学意义（分别为κ=0.43和0.34）。P2X7R与NLRP3或IL-1β之间的上述关联亦通过卡方检验得到验证。 结论：化脓性汗腺炎患者皮肤中P2X7R、NLRP3及IL-1β均呈过表达，整条P2X7R/NLRP3/IL-1β促炎轴大概率处于过度激活状态。该发现可为阐明该病的发病机制提供线索，并为开发新型治疗手段及疾病活动度标志物提供思路。