Difference in cell biological behavior and ultraviolet radiation effect between double-head and unilateral pterygium fibroblasts cultured in vitro

Objective

To explore the difference in the proliferation, migration and ultraviolet radiation effect between double-head and unilateral pterygium fibroblasts (HPFs) cultured in vitro.

Methods

Pterygium tissue was obtained from patients who underwent pterygium excision with conjunctival transposition from March 2017 to March 2018 in the Second Affiliated Hospital of Guangzhou Medical University.Nineteen cases with bilateral pterygium and 19 cases with unilateral pterygium were selected for this research.Twelve cases of normal conjunctival tissue were obtained from donor eyes.The fibroblasts were divided into HPFs-nasal (HPFs-N), HPFs-temporal (HPFs-T), unilateral HPFs and human conjunctiva fibroblasts (HCFs), which was taken from the nasal side of the bilateral pterygium, the temporal side of the bilateral pterygium, the unilateral pterygium and the normal conjunctiva, respectively.Human pterygium and normal conjunctival fibroblasts were isolated and cultured by tissue culture method.The fibroblasts were divided into an ultraviolet irradiation group and a normal light illumination group.The cell growth curve was detected by methyl thiazolyl tetrazolium (MTT) assay.The cell scratch healing rate was detected by the cell scratch test after 48 hours.The expression of α-smooth muscle actin αwas detected by immunofluorescence staining, and the number of positive cells in each group was compared.The fibroblasts cultured in vitro were irradiated with ultraviolet irradiation, and the cell scratch healing rate, growth curve and expression of α-SMA were observed.The study protocol was approved by the Ethics Committee of the Second Affiliated Hospital of Guangzhou Medical University.Written informed consent was obtained from each subject.

Results

The pterygium and conjunctival fibroblasts were spindle shaped, and the growth rate was gradually increased from day 1 to day 7.The growth rate of HPFs-N was the fastest, and the growth rate of HCFs was the slowest.The growth rate of the four types of fibroblasts was significantly increased after exposure to ultraviolet.There were significant differences in the cell scratch healing rates and α-SMA positive cell expression rates among the four fibroblast types under different lighting conditions (F_group=158.064, P<0.05; F_{cell type}=326.582, P<0.05.F_group=4.731, P<0.05; F_{cell type}=172.813, P<0.05), of which the scratches healing rate in the HPFs-N cells after 48 hours under the normal light conditions was (79.67±0.86)%, which was significantly higher than HPFs-T ([54.04±0.33]%), unilateral HPFs ([64.12±0.21]%) and HCFs ([58.86±0.41]%), the α-SMA positive cell expression rate in the HPFs-N cells after 48 hours under the normal light conditions was (28.87±1.02)%, which was significantly higher than that in HPFs-T ([13.67±0.23]%), unilateral HPFs ([20.35±1.72]%) and HCFs ([5.12±0.45]%) (all at P<0.05); the cell scratch healing rates and α-SMA positive cell expression rates were significantly increased in the ultraviolet irradiation group than those in the normal light illumination group (all at P<0.05).

Conclusions

The nasal side of double-head pterygium fibroblasts is more proliferous and more migratory than that of the unilateral pterygium, the expression of α-SMA is also increased, which can be further enhanced by ultraviolet irradiation.