Study Of The Effects Of Continuous Artificial Light On The Maturation And Quality Of Mouse Oocytes

Light is one of the environmental factors that need to be considered in the production process of animal husbandry.The intensity and frequency of light have an important impact on the growth and reproduction performance of female mammals.Oocyte maturation is a key factor affecting the reproductive performance of female animals.However,the relationship between light duration and maturation/quality of female animal oocytes is still unclear.This study intends to use mice as the research object to initially explore the effects of different durations of light on oocytes maturation and quality.The main research contents and results are as follows:1.Effect of different duration of light on the maturation of oocytesIn order to explore the effect of long light exposure on the maturation of oocytes,the mice were divided into a control group(12 h light:12 h dark)and a continuous artificial light treatment group(24 h light),the treatment time was 0,1,3,5 and 7 weeks respectively.The vaginal smear method was used to identify the estrous cycle in mice,and the results showed that the estrous cycle in mice was significantly disordered after 5 weeks of light treatment.After 7 weeks of light treatment,the results of in vitro maturation culture of oocytes showed that the oocyte polar body extrusion rate was significantly lower than that of the control group(control group:72.6±2.3%,n=146 vs 7 weeks:61.3±1.5%,n=149;P<0.05).Further,immunofluorescence was used to observe the spindle morphology and chromosome arrangement of oocytes in the control group and the 7-week light treatment group.The results showed that 7-week light treatment significantly increased the spindle and chromosome abnormality rate(spindle:40.3±2.7%,n=52 vs 15.7±2.6%,n=70,P<0.01;chromosome:41.5±1.6%,n=52vs 22.7±3.7%,n=70,P<0.01).Finally,the chromosome aneuploidy of oocytes was detected by chromosome spreading.The results showed that 7 weeks of light treatment significantly increased the aneuploidy rate of oocytes(60.1±5.1%,n=33 vs 28.3±3.0%,n=42,P<0.01).According to the above results,the following experiments were uniformly treated with 7 weeks of light exposure.2.Effects of different duration of light on the quality of oocytesIn order to explore the effect of light treatment on the quality of mouse oocytes,this part mainly carried out the following studies:1)The ROS detection kit was used to detect the level of reactive oxygen species(ROS)in oocytes.The results showed that light treatment significantly increased the level of ROS(43.9±1.6,n=42 vs 25.8±1.1,n=53,P<0.0001);the expression of antioxidant enzyme-related genes further was detected using q PCR,and the results showed that the expression of related antioxidant enzymes Cat,Gpx1,and Sod2 were significantly down-regulated(Cat:0.833±0.058 vs 1.217±0.056,P<0.01;Gpx1:0.705±0.076 vs 1.462±0.210,P<0.05;Sod2:0.780±0.047 vs 1.152±0.009,P<0.01),indicating that light treatment induced oxidative stress.2)Immunofluorescence staining was used to detect the number of mitochondria,and the results showed that the number of mitochondria was decreased significantly(31.4±0.9,n=35 vs 52.4±1.2,n=53,P<0.0001);using chemiluminescence to detect ATP levels,the results showed that light treatment significantly reduced ATP levels(0.72±0.01,n=120 vs 1.00±0.02,n=120,P<0.0001),indicating that light treatment impaired mitochondrial function.3)Immunofluorescence staining was used to detect the levels of oocyte apoptosis and autophagy.The results showed that the levels of oocyte apoptosis and autophagy were significantly increased(apoptosis:1.378±0.054 vs 1.000±0.080,P<0.05;autophagy:6.15±0.4,n=73 vs 4.94±0.4,n=39,P<0.05);q PCR was used to detect the expression of apoptosis and autophagy-related genes.The results showed that light treatment significantly down-regulated the expression of apoptosis gene Bcl2(0.407±0.087 vs 1.118±0.119,P<0.01),and up-regulated the expression of the autophagy gene Beclin 1(1.385±0.149 vs 1.009±0.041,P<0.05),the above results indicate that light treatment induced production of apoptosis and the autophagy.4)Immunofluorescence staining was used to detect the epigenetic modification level.The results showed that light treatment resulted in oocyte acetylation level(14.4±1.8,n=41 vs 19.9±2.0,n=47,P<0.05),DNA methylation level(5m C:17.5±1.1,n=91 vs 22.8±1.1,n=76,P<0.001),RNA methylation level(m~6A:11.3±0.5,n=99 vs 7.24±0.3,n=95,P<0.0001),and H3K4me2histone methylation level(32.9±2.6,n=47 vs 45.7±2.8,n=46,P<0.01)were decreased significantly;while histone H3K27me2 methylation level was increased significantly(45.3±2.5,n=48 vs 29.3±1.9,n=48,P<0.0001),indicating that light treatment caused changes in the epigenetic modification level of oocytes.In summary,our results indicate that continuous artificial light results in abnormal maturation and reduced quality of mouse oocytes.This study will help us better understand the relationship between light and livestock reproductive performance,and provide a reference for the optimization of light-related parameters in livestock production.