Optical Diagnostic Techniques Group, Department of Theoretical, Atomic and Optical Physics, University of Valladolid, Valladolid, Spain.
Instituto Universitario Fernández Vega, Universidad de Oviedo, Oviedo, Spain.
Ophthalmic Res. 2020;63(2):203-212. doi: 10.1159/000501507. Epub 2019 Nov 6.
To compare the clinical and histological outcomes after intrastromal corneal ring segment (ICRS) implantation with and without plasma rich in growth factors (PRGF) in an experimental animal model.
First, the toxicity of PRGF was tested in hen's keratocyte cultures. Then, an animal model with 18 hens was randomly divided into 2 groups. In the first group, one ICRS was implanted in each eye (ICRS group). In the second group, the ICRS was firstly immersed 30 min in PRGF-Endoret solution, then implanted and, finally, PRGF-Endoret was inoculated into the channel (PRGF-ICRS group). Animals of each group were also separated into 3 groups regarding the time they were sacrificed, and corneal tissue was fixed for histological analysis at 2, 7 and 30 days. Cell death was detected by terminal uridine nick end labelling (TUNEL) assay. Proliferation was labelled by 5-bromo-2-deoxyuridine (BrdU) incorporation and myofibroblast differentiation by alpha-smooth muscle actin (αSMA) immunodetection. Clinical examination, analyzing epithelial wound closure, deposits and stromal haze, was carried out at the different study times.
No toxic effect was observed by PRGF in hen stromal cell cultures. Clinically, in PRGF-ICRS corneas at 7 days, there were more deposits with higher intensity than in ICRS group. Histologically, at day 2 there was less epithelial damage over the segment in the PRGF-ICRS group, corneal oedema around the segment disappeared earlier and, at day 7, there was also double the number of cells around the segment than in the ICRS group displaying different morphologies. The number of TUNEL-positive cells was statistically higher in the PRGF-ICRS group at 7 and 30 days, and the number of BrdU-positive cells was statistically higher at all analyzed times. However, there were no differences in the number of αSMA-positive cells at 30 days between both groups.
The ICRS immersion in PRGF-Endoret prior and after to its corneal implantation, in an experimental animal model, enhances clinical deposits and histological cell turnover without increasing myofibroblast differentiation reducing stromal wound-healing time after surgery.
比较富含生长因子的富血小板纤维蛋白（PRGF）在角膜基质内环段（ICRS）植入物中的临床和组织学结果。
首先，在鸡角膜细胞培养物中测试 PRGF 的毒性。然后，使用 18 只母鸡的动物模型随机分为 2 组。在第一组中，每只眼睛植入一个 ICRS（ICRS 组）。在第二组中，首先将 ICRS 在 PRGF-Endoret 溶液中浸泡 30 分钟，然后植入，最后将 PRGF-Endoret 接种到通道中（PRGF-ICRS 组）。每组动物还根据处死时间分为 3 组，在第 2、7 和 30 天固定角膜组织进行组织学分析。通过末端尿嘧啶核苷尼克末端标记（TUNEL）检测细胞死亡。增殖通过 5-溴-2-脱氧尿苷（BrdU）掺入标记，肌成纤维细胞分化通过α-平滑肌肌动蛋白（αSMA）免疫检测标记。在不同的研究时间进行临床检查，分析上皮伤口闭合、沉积物和基质混浊。
PRGF 在鸡基质细胞培养物中未观察到毒性作用。临床上，在 PRGF-ICRS 角膜中，第 7 天的沉积物密度更高。组织学上，在 PRGF-ICRS 组中，段上的上皮损伤程度较低，段周围的角膜水肿更早消失，第 7 天，段周围的细胞数量也增加了一倍，显示出不同的形态。第 7 天和第 30 天，PRGF-ICRS 组中 TUNEL 阳性细胞的数量明显高于 ICRS 组，所有分析时间的 BrdU 阳性细胞数量也明显高于 ICRS 组。然而，两组在第 30 天的αSMA 阳性细胞数量没有差异。
在实验动物模型中，将 ICRS 浸入 PRGF-Endoret 中，然后将其植入角膜，可增强临床沉积物和组织学细胞更新，同时减少术后基质伤口愈合时间，而不会增加肌成纤维细胞分化。
