Department of Ophthalmology, University Hospital Basel, Basel, Switzerland.
Institute of Molecular and Clinical Ophthalmology Basel, Basel, Switzerland.
PLoS One. 2021 Apr 1;16(4):e0249344. doi: 10.1371/journal.pone.0249344. eCollection 2021.
To examine the effect of prolactin (PRL) on human corneal stromal fibroblasts (CSFs), derived from healthy individuals and from keratoconus (KC) patients, in vitro, specifically assessing physiological and elevated PRL concentrations as apparent during pregnancy.
Eye bank corneas of 3 female and 3 male healthy individuals as well as the corneal buttons of 3 female and 3 male KC patients were utilized for this study. The endothelium of the cornea was removed with sterile surgical scalpels, the probes were washed repeatedly with Dulbecco's PBS and corneoscleral rims were trimmed off. Subsequently the corneal stroma was digested with collagenase type I and the harvested CSFs were cultured. We then examined (1) cell proliferation, (2) cell viability and (3) cytokine release of CSFs upon exposure to prolactin in vitro.
With respect to viability and proliferation our experiments did not show significant differences between CSFs exposed to different PRL concentrations. Our data show a significantly lower IL-8 concentration in normal CSFs exposed to 10ng/ml PRL compared to 0ng/ml and 1000ng/ml at 5 hours post exposition. Moreover, we can report significantly lower secretion of IL-8, IL-6, HGF, VEGF and FGFb in KC CSFs compared to normal CSFs, independent of PRL exposure, as determined by cytokine ELISA.
Our data in part points towards corneal cytokine secretion as a possible link between altered stromal PRL concentrations and KC progression. However, in our small dataset a significant influence of PRL concentration on cytokine secretion can only be described for IL-8 in normal CSFs. Further our results contribute to existing reports on the importance of cytokines in KC development, with an emphasis on significantly lower cytokine secretion in KC CSFs compared to normal controls.
在体外研究催乳素（PRL）对来自健康个体和圆锥角膜（KC）患者的人角膜基质成纤维细胞（CSF）的影响，特别评估妊娠期间出现的生理和升高的 PRL 浓度。
本研究使用了 3 名女性和 3 名男性健康个体的眼库角膜以及 3 名女性和 3 名男性 KC 患者的角膜纽扣。使用无菌手术刀片去除角膜的内皮，探针用 Dulbecco's PBS 反复清洗，并修剪角膜缘。然后用胶原酶 I 消化角膜基质，并培养收获的 CSF。我们随后研究了（1）CSF 在体外暴露于催乳素后的细胞增殖，（2）细胞活力和（3）细胞因子释放。
就活力和增殖而言，我们的实验没有显示出暴露于不同 PRL 浓度的 CSF 之间存在显著差异。我们的数据显示，与 0ng/ml 和 1000ng/ml 相比，在暴露于 10ng/ml PRL 5 小时后，正常 CSF 中的 IL-8 浓度显著降低。此外，我们可以报告 KC CSF 中的 IL-8、IL-6、HGF、VEGF 和 FGFb 分泌明显低于正常 CSF，无论是否暴露于 PRL，这是通过细胞因子 ELISA 确定的。
我们的数据部分表明，角膜细胞因子分泌可能是基质 PRL 浓度改变与 KC 进展之间的联系。然而，在我们的小数据集，PRL 浓度对细胞因子分泌的显著影响仅在正常 CSF 中描述为 IL-8。此外，我们的结果为 KC 发病机制中细胞因子的重要性提供了支持，特别强调了 KC CSF 中的细胞因子分泌明显低于正常对照。
