Supplementary Material for: The Effects of Mucopolysaccharide Polysulfate on Steroid-Induced Tight Junction Barrier Dysfunction in Human Epidermal Keratinocytes and a 3D Skin Model

Introduction: The long-term use of topical corticosteroids (TCS) is associated with side effects such as skin atrophy and barrier deterioration. Moisturizers, such as mucopolysaccharide polysulfate (MPS), have been reported to prevent relapses in atopic dermatitis (AD) when used in combination with TCS. However, the mechanisms underlying the positive effects of MPS in combination with TCS in AD are poorly understood. In the present study, we investigated the effects of MPS in combination with clobetasol 17-propionate (CP) on tight junction (TJ) barrier function in human epidermal keratinocytes (HEKa) and 3D skin models.
Methods: The expression of claudin-1, which is crucial for TJ barrier function in keratinocytes, and transepithelial electrical resistance (TEER) was measured in CP-treated human keratinocytes incubated with and without MPS. A TJ permeability assay, using Sulfo-NHS-Biotin as a tracer, was also conducted in a 3D skin model.
Results: CP reduced claudin-1 expression and TEER in human keratinocytes, whereas MPS inhibited these CP-induced effects. Moreover, MPS inhibited the increase in CP-induced TJ permeability in a 3D skin model.
Conclusion: The present study demonstrated that MPS improved TJ barrier impairment induced by CP. The improvement of TJ barrier function may partially be responsible for the delayed relapse of AD induced by the combination of MPS and TCS.

研究背景：外用糖皮质激素（topical corticosteroids, TCS）长期使用会引发皮肤萎缩、皮肤屏障功能受损等不良反应。有研究表明，黏多糖聚硫酸酯（mucopolysaccharide polysulfate, MPS）类保湿剂与外用糖皮质激素联用时，可预防特应性皮炎（atopic dermatitis, AD）复发。然而，二者联合使用改善特应性皮炎的具体作用机制尚不明确。本研究旨在探究黏多糖聚硫酸酯与丙酸氯倍他索17-丙酸酯（clobetasol 17-propionate, CP）联合使用，对人表皮角质形成细胞（human epidermal keratinocytes, HEKa）及三维皮肤模型中紧密连接（tight junction, TJ）屏障功能的影响。
研究方法：分别检测在添加与未添加黏多糖聚硫酸酯的培养基中，经丙酸氯倍他索处理的人表皮角质形成细胞内，紧密连接屏障功能关键蛋白闭合蛋白-1（claudin-1）的表达水平，以及跨上皮电阻（transepithelial electrical resistance, TEER）。此外，以磺基-NHS-生物素（Sulfo-NHS-Biotin）为示踪剂，在三维皮肤模型中开展紧密连接通透性实验。
研究结果：丙酸氯倍他索会降低人表皮角质形成细胞中闭合蛋白-1的表达水平与跨上皮电阻值，而黏多糖聚硫酸酯可抑制上述丙酸氯倍他索诱导的变化。此外，黏多糖聚硫酸酯可抑制三维皮肤模型中丙酸氯倍他索诱导的紧密连接通透性升高。
研究结论：本研究证实，黏多糖聚硫酸酯可改善丙酸氯倍他索诱导的紧密连接屏障功能受损。黏多糖聚硫酸酯与外用糖皮质激素联合使用可延缓特应性皮炎复发，其部分作用机制可能与紧密连接屏障功能的改善有关。