Expression Of HGF In The Guinea Pig's Retinal Pigment Epithelial Cells And The Ca²⁺ Study In Experimental Myopia

Objective: To determine the expression of hepatocyte growth factor (HGF) and to study the calcium factor (Ca²⁺) changes in cultured guinea pig anterior and posterior retinal pigment epithelial cells in lens-induced myopia. Some conclusions were valuable for revealing the pathological mechanism of myopia.Methods: 10 two-week-old guinea pigs were lens-induced monocularly, a spherical lenses of -10.00D was wear to unilateral eye, another eye in each animal was untreated as self-control (SC). Before and after experiment, refraction was measured using retinoscopy and the oculus uterque axis oculi length determined by A-scan ultrasonography. The retinal pigment epithelial cells of each group were cultured and passaged 1 generation in vitro, The cultured cells were determined to be pigment epithelial cells using immunocytochemistry method. Immunocytochemistry was used to detect the expression of HGF in the retinal pigment epithelial cells and to study by light microscope. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was used to detect the expression of HGFmRNA in the retinal pigment epithelial cells and to study by agarose gel image analytical system. Cultured RPE cells were analyzed by using the fluorescence Ca²⁺ dye fluo-3/AM and laser scanning confocal microscope (LSCM). These data was statistically analysed by SPSS 12.0 system.Results:1 Eyes became myopic -6.70±1.93D and elongated 1.53±0.31mm 45 days after insertion of the lenses (P<0.05).2 Cultured cells began to adherence grow after 2-3 days, and grew to confluent after 1 week. The cultured cells were determined to be retinal pigment epithelium cells using immunocytochemistry methods.3 The immunocytochemistry showed that expression of HGF was detected in all anterior and posterior retinal pigment epithelial cells. The expression of HGF located in cytoplasm. The expressions of HGF in model eyes were higher than control eyes (P<0.05). The expression of HGF in anterior eyes was almost the same with the expression in posterior (P>0.05). There was significant difference of the expression of HGF between model eyes and control eyes (P<0.05).4 Reverse Transcription-Polymerase Chain Reaction (RT-PCR) showed that the expression of HGFmRNA in the retinal pigment epithelial cells in model eyes were higher than control eyes (P<0.05). The expression of HGFmRNA in anterior eyes was almost the same with the expression in posterior (P>0.05). There was significant difference of the expression of HGFmRNA between model eyes and control eyes (P<0.05).5 The fluorescence Ca²⁺ dye fluo-3/AM and laser scanning confocal microscope (LSCM) showed that normal fluorescence in RPE cells was strong and distributed throughout the cells. The model eyes appeared to be more fluorescent than the control eyes. There was no obvious difference bettwen anterior part and posterior pole of retinal pigment epithelial cells. There was significant difference of the fluorescence intensity between model eyes and control eyes (P<0.05).Conclusions: lenses can induce axial growth and lead to myopia in young guinea pigs. Expression of HGF was detected in guinea pigs retinal pigment epithelial cells. It showed that the HGF played an important role in the course of development of myopia. There was fluorescence in RPE cells. The fluorescence intensity in RPE cells of model eyes was stronger than in control eyes. It showed that Ca²⁺ intensity increased during the process of development of myopia.