Digital Analysis Of The Morphology Of Oocyte Spindle And Chromosome Based On In Situ Optical Imaging

The combination of optical imaging and image processing is an important technique to understand the conformation of biological structure.The spatial conformation of biological structure is of great significance to understand its physiological function and development mechanism.In this paper,two-photon fluorescence imaging technology is used to obtain the biological samples in situ and stereo based on the immunofluorescence method.Furthermore,the conformational information of biological structure is quantified by 3D image segmentation,3D reconstruction and digital modeling.The spatial conformation of the spindle and chromosome in mouse oocytes are studied by this method.The spatial conformation of chromosome is closely related to the chromosomal density,the gene transcription activity and the nucleus function.The spatial conformation of the spindle is related to the arrangement of chromosomes and cell division.Its integrity plays an important role in whether cells can reproduce normally.The conformational study of chromosomes and spindles in oocytes is important for assessing oocyte quality and analyzing reproductive development problems(eg,birth defects).Based on the in vitro culture technique,the effects of methotrexate on the chromosomes and spindle of oocytes are studied.Methotrexate,a drug that inhibits cell proliferation,is widely used in the treatment of diseases such as cancer,rheumatoid arthritis and ectopic pregnancy.Many studies have shown that methotrexate has strong side effects on rapidly dividing somatic cells.However,its side effects on female germ cells are rarely reported.Compared with the control group(84%),it is found that the percentage of germinal vesicle break up in the methotrexate group decreased to 73%(p<0.05).The rate of the first polar body extrusion(53%)in the methotrexate group is also lower than that in the control group(63%;p<0.05).The abnormal chromosome arrangement and abnormal spindle morphology in the methotrexate group are 60% and 49%,respectively,which are significantly higher than those in the control group(24%,17%,p<0.05).These results suggest that methotrexate has an effect on oocyte maturation.Subsequently,the effects of superovulation on the quality of oocytes are studied.As a kind of non-natural ovulation,super ovulation is widely used in clinical assisted reproductive and basic science research.However,the effect of this kind of ovulation on oocyte maturation is still not clear.Our results showed that the chromosome space arrangement is abnormal,but it does not affect the geometry of the chromosome.These results may suggest that the hormone superovulation interferes with the movement and migration of chromosomes.This thesis established a methodology to characterize the spatial conformation of spindle and chromosome in oocytes.The results of this study are of great significance to understand the reproductive development.