The Study Of Influence Of Amphiregulin On Biologic Paramenters In Myopia Guinea Pigs And Clinical Genetics

Objective: The investigation of clinical epidemiology of high myopia,we analyzed the distribution of refraction and ocular biometric parameters,screenedthe frequence of gene mutation for people with high myopia and explored the effect of intravitreal injection of Amphiregulin(AREG)antibody on refraction,axial length in lens-induced myopic guinea pigs.Method: The study included three portions.Experiment 1: The clinical investigation of gene epidemics in high myopia,after gain the informed consents by patients,200 cases of myopia patients from May 2016 to December 2017 were carried out including visual acuity,refraction,intraocular pressure,axial length,corneal curvature,eye fundus examination.According to inclusion and exclusion criteria,170 patients with high myopia were selected,and fill out the questionnaire about myopia of family members.Drewing 5-6ml blood to detect gene mutation frequency,to observe the genes which related with myopia.Experiment 2: To investigate the effect of intravitreal injection of Amphiregulin(AREG)antibody on refraction,axial length and retinal AREG mRNA and protein expression in lens induced myopic guinea pigs.Sixty two-week-old pigmented guinea pigs were randomly divided into five groups: normal group(without any treated),myopia group(lens induced myopia plus injection of Ringer’s solution 3ul on right eyes),low-dosage group,mediumdosage group,high-dosage group(lens induced myopia plus injection of AREG antibody respectivily 5ug,10 ug 15ug on right eyes),twelve animals in each group.Except the normal group,right eyes of other four groups,were covered with-10.00 D lens,while the rest eyes were not treated.The parameter of refraction,axial length were measured by Ribbon retinoscope and A/B scan of ophthalmology respectively each week.The injection of Ringer’s solution and AREG antibody were once a week and lasted for 3 weeks.The expression of mRNA and protein of AREG were explored by quantitative real-time PCR and immunofluorescence staining.PAS staining and measure the thickness of sclera on posterior pole.Experiment 3: To investigate the effect of intravitreal injection of AREG protein on refraction,axial length and retinal AREG mRNA and protein expression in normal guinea pigs.Forty-eight two-week-old pigmented guinea pigs were randomly divided into four groups: normal group(without any treated),low-dosage group,medium-dosage group,high-dosage group(normal plus injection of AREG protein respectivily 50 ng,100ng,200 ng on right eyes and equivalent linger on left eyes),twelve animals in each group.The parameter of refraction,axial length were measured by Ribbon retinoscope and A/B scan of ophthalmology respectively each week.The injection of Ringer’s solution and AREG protein were once a week and lasted for 4 weeks.The expression of mRNA and protein of AREG were explored by quantitative real-time PCR and immunofluorescence staining.PAS staining and measure the thickness of sclera on posterior pole.Results: 1.There were more common changes in high myopia,serious decrease of vision(0.05±0.04)and refraction(-8.02±2.23)D,increase of axial length(26.65±1.31)mm and corneal curvature(43.11±1.47)D,cornea anterior protrude,anterior chamber deepened(3.76±0.27)mm,intraocular pressure was normal(15.88±2.34)mmHg.2.In high myopia group,increase of axial length is the main type,acquired and scattered.With the growth of age,progressively deepened of myopia,the decrease of refreaction and increase of axial length.The probability of developing high myopia and retinopathy is increased.3.AREG gene have increased frequency of mutation in highly myopic people.The frequency of gene mutation in patients with axial high myopia is higher than that of patients with refractive high myopia.The mutation frequency of the gene increased in high myopia group.The mutation frequency is high.AREG is mainly in China and East Asia.The mutation frequency of high myopia is race specific.4.In retina of myopia guinea pigs,compared with normal guinea pigs,the expression of AREG protein and mRNA is increased.5.AREG widely distribute in retina,such as equator,midpoint between equator and posterior pole,and posterior pole.Lens induced myopia guinea pigs which intravitreal injection of AREG antibody,will in a dose-dependent inhabite the increase of axial length and the decrease of refraction.The thinning of sclera in posterior pole was inhabited,compared with myopia group,the thickness of sclera on posterior pole is increased in three myopia group with intravitreal injection of AREG antibody,especially in low-dosage and mediumdosage(p<0.05 6.Intravitreal injection of AREG protein in normal guinea pigs can not promote the decrease of refraction and increase of axial length.The expression of AREG protein of retina in the posterior pole was increased,but has little influence on AREG mRNA and can not promote the thinning of sclera in posterior pole.Conclusion:1.The increase of axial length of high myopia is the main type,the excessive growth of the axial length is harmful to vision and fundus oculi.2.In high-myopia populations,there is an increase in the frequency of AREG genetic variation,and the axial length induced myipia is the main type.3.Intravitreal injection of AREG antibody in lens induced myopia of guinea pigs can inhabite the progression of myopia,but intravitreal injection of AREG protein in normal guinea pigs can not induced myopia,we believe AREG plays a certain role in the development of myopia,but it is not absolute.