Glucocorticoids are the first-line treatment for Pemphigus vulgaris (PV), but its serious side effects can be life-threatening for PV patients. Tacrolimus (FK506) has been reported to have an adjuvant treatment effect against PV. However, the mechanism underlying the inhibitory effect of FK506 on PV-IgG-induced acantholysis is unclear. The objective of this study was to explore the effect of FK506 on desmoglein (Dsg) expression and cell adhesion in an immortalized human keratinocyte cell line (HaCaT cells) stimulated with PV sera. A cell culture model of PV was established by stimulating HaCaT cells with 5% PV sera with or without FK506 and clobetasol propionate (CP) treatment. The effects of PV sera on intercellular junctions and protein levels of p38 mitogen-activated protein kinase (p38MAPK), heat shock protein 27 (HSP27), and Dsg were assayed using western blot analysis, immunofluorescence staining, and a keratinocyte dissociation assay. PV sera-induced downregulation of Dsg3 was observed in HaCaT cells and was blocked by FK506 and/or CP. Immunofluorescence staining revealed that linear deposits of Dsg3 on the surface of HaCaT cells in the PV sera group disappeared and were replaced by granular and agglomerated fluorescent particles on the cell surface; however, this effect was reversed by FK506 and/or CP treatment. Furthermore, cell dissociation assays showed that FK506 alone or in combination with CP increased cell adhesion in HaCaT cells and ameliorated loss of cell adhesion induced by PV sera. Additionally, FK506 noticeably decreased the PV serum-induced phosphorylation of HSP 27, but had no effect on p38MAPK phosphorylation. FK506 reverses PV-IgG induced-Dsg depletion and desmosomal dissociation in HaCaT cells, and this effect may be obtained by inhibiting HSP27 phosphorylation.

中文翻译：

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他克莫司通过抑制热休克蛋白 27 磷酸化逆转寻常型天疱疮血清诱导的 HaCaT 细胞中桥粒芯糖蛋白的消耗

糖皮质激素是寻常型天疱疮 (PV) 的一线治疗方法，但其严重的副作用可能会危及 PV 患者的生命。据报道，他克莫司 (FK506) 对真性红细胞增多症具有辅助治疗作用。然而，FK506 对 PV-IgG 诱导的棘层松解症的抑制作用的机制尚不清楚。本研究的目的是探讨 FK506 对 PV 血清刺激的永生化人角质形成细胞系（HaCaT 细胞）中桥粒芯糖蛋白 (Dsg) 表达和细胞粘附的影响。通过用 5% PV 血清刺激 HaCaT 细胞，并加或不加 FK506 和丙酸氯倍他索 (CP) 处理，建立 PV 细胞培养模型。使用蛋白质印迹分析、免疫荧光染色和角质形成细胞解离测定来测定 PV 血清对细胞间连接以及 p38 丝裂原激活蛋白激酶 (p38MAPK)、热休克蛋白 27 (HSP27) 和 Dsg 的蛋白质水平的影响。在 HaCaT 细胞中观察到 PV 血清诱导的 Dsg3 下调，并被 FK506 和/或 CP 阻断。免疫荧光染色显示PV血清组HaCaT细胞表面Dsg3的线状沉积消失，取而代之的是细胞表面颗粒状、团聚的荧光颗粒；然而，这种效应被 FK506 和/或 CP 治疗逆转。此外，细胞解离测定表明，单独使用 FK506 或与 CP 组合使用可增加 HaCaT 细胞中的细胞粘附，并改善 PV 血清诱导的细胞粘附丧失。此外，FK506 显着降低 PV 血清诱导的 HSP 27 磷酸化，但对 p38MAPK 磷酸化没有影响。FK506 可逆转 HaCaT 细胞中 PV-IgG 诱导的 Dsg 耗竭和桥粒解离，这种作用可能通过抑制 HSP27 磷酸化来获得。