Abnormal and deregulated skin wound healing associated with prolonged inflammation may result in dermal fibrosis. Since the current therapeutic strategies revealed unsatisfactory, the investigation of alternative approaches such as those based on the use of specific probiotic strains could provide promising therapeutic options. In this study, we aimed to evaluate whether the lysate from S. thermophilus could antagonize the fibrogenic effects of TGF-β1 in normal human dermal fibroblasts (NHDF). NHDF were exposed to TGF-β1 to establish a fibrotic phenotype. Proliferation rate and cell number were measured using the IncuCyte® Live Cell Imager system and the trypan blue dye exclusion test. Phenoconversion markers (α-SMA and fibronectin) and collagen I levels were assessed by western blot and immunofluorescence. The mRNA levels of TGF-β1 were evaluated by RT-PCR. The Smad2/3 phosphorylation level as well as β-catenin and PPARγ expression, were assessed by western blot. The cell contractility function and migration of NHDF were studied using collagen gel retraction assay, and scratch wound healing assay, respectively. The effects of S. thermophilus lysate, alone or combined with TGF-β1, were evaluated on all of the above-listed parameters and markers associated with TGF-β1-induced fibrotic phenotype. Exposure to the S. thermophilus lysate significantly reduced the key mediators and events involved in the abnormal activation of myofibroblasts by TGF-β1 within the fibrotic profile. The S. thermophilus treatment significantly reduced cell proliferation, migration, and myo-differentiation. In addition, the treatment with probiotic lysate reduced the α-SMA, fibronectin, collagen-I expression levels, and affected the collagen contraction ability of activated dermal fibroblasts. Moreover, the probiotic targeted the TGF-β1 signaling, reducing Smad2/3 activation, TGF-β1 mRNA level, and β-catenin expression through the upregulation of PPARγ. This is the first report showing that S. thermophilus lysate had a remarkable anti-fibrotic effect in TGF-β1-activated NHDF by inhibiting Smad signaling. Notably, the probiotic was able to reduce β-catenin and increase PPARγ levels. The findings support our point that S. thermophilus may help prevent or treat hypertrophic scarring and keloids.

中文翻译：

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益生菌嗜热链球菌对抗 TGF-β1 诱导的正常人真皮成纤维细胞纤维化反应的功效

与长期炎症相关的异常和失调的皮肤伤口愈合可能导致真皮纤维化。由于目前的治疗策略并不令人满意，因此研究替代方法（例如基于使用特定益生菌菌株的方法）可以提供有希望的治疗选择。在这项研究中，我们旨在评估来自嗜热链球菌的裂解物是否可以拮抗 TGF-β1 在正常人真皮成纤维细胞 (NHDF) 中的纤维化作用。NHDF 暴露于 TGF-β1 以建立纤维化表型。使用 IncuCyte® Live Cell Imager 系统和台盼蓝染料排斥试验测量增殖率和细胞数量。通过蛋白质印迹和免疫荧光评估表型转化标记物（α-SMA 和纤连蛋白）和胶原 I 水平。通过 RT-PCR 评估 TGF-β1 的 mRNA 水平。通过蛋白质印迹评估 Smad2/3 磷酸化水平以及 β-连环蛋白和 PPARγ 表达。分别使用胶原凝胶回缩试验和划痕伤口愈合试验研究了 NHDF 的细胞收缩功能和迁移。单独或与 TGF-β1 联合使用嗜热链球菌裂解物对上述所有与 TGF-β1 诱导的纤维化表型相关的参数和标志物的影响进行了评估。暴露于嗜热链球菌裂解物显着减少了纤维化特征中 TGF-β1 异常激活肌成纤维细胞所涉及的关键介质和事件。嗜热链球菌处理显着降低了细胞增殖、迁移和肌分化。此外，益生菌裂解物处理降低了 α-SMA，纤维连接蛋白、胶原蛋白-I 的表达水平，并影响活化的真皮成纤维细胞的胶原蛋白收缩能力。此外，益生菌靶向 TGF-β1 信号，通过上调 PPARγ 减少 Smad2/3 激活、TGF-β1 mRNA 水平和 β-catenin 表达。这是第一份报告显示嗜热链球菌裂解物通过抑制 Smad 信号传导在 TGF-β1 激活的 NHDF 中具有显着的抗纤维化作用。值得注意的是，益生菌能够减少 β-连环蛋白并增加 PPARγ 水平。这些发现支持我们的观点，即嗜热链球菌可能有助于预防或治疗增生性疤痕和瘢痕疙瘩。和 β-catenin 表达通过上调 PPARγ。这是第一份报告显示嗜热链球菌裂解物通过抑制 Smad 信号传导在 TGF-β1 激活的 NHDF 中具有显着的抗纤维化作用。值得注意的是，益生菌能够减少 β-连环蛋白并增加 PPARγ 水平。这些发现支持我们的观点，即嗜热链球菌可能有助于预防或治疗增生性疤痕和瘢痕疙瘩。和 β-catenin 表达通过上调 PPARγ。这是第一份报告显示嗜热链球菌裂解物通过抑制 Smad 信号传导在 TGF-β1 激活的 NHDF 中具有显着的抗纤维化作用。值得注意的是，益生菌能够减少 β-连环蛋白并增加 PPARγ 水平。这些发现支持我们的观点，即嗜热链球菌可能有助于预防或治疗增生性疤痕和瘢痕疙瘩。