Study On Genetic Variation Detection And Genetic Laws Of AZF Region In Infertility Male With Spermatogenesis Impairment

Y chromosome microdeletion is the most common genetic cause of spermatogenetic malfunction in male infertility.The widely reported microdeletions are include three types of AZF region deletions(AZFa,AZFb and AZFc)or the joint deletion(AZFa +b,AZFa+c,AZFb+c and AZFa+b+c).The microdeletion of AZF region was related to changes in various sperms production,including support cell syndrome(SCOS),maturation arrest,and low sperm production.Along with the progress of the treatment technologies,it is an urgent need to understand gene function and genetic mechanism of AZF area on Y chromosome.That is facilitate to recover the etiology of spermatogenic failure for infertility male and timely to use the new technologies such as gene editing,stem cell transplantation for a effective treatment,who have their own offspring.The Y chromosome has a specific sequence structure,which is rich in palindromic sequences and duplicated copies,which is the structural basis for sequence rearrangement of the deletion or duplication and the maintenance genes.Due to the complexity of the Y chromosome structure,the polymerase chain reaction(PCR)method(STS-PCR),which is currently recommended to used for detection.We examined 1866 samples by STS-PCR analysis,and found that the incidence of AZF deletion was 10.39% in the azoospermia group,7.33% in the oligospermia group,and no deletion in patients with normal sperm concentration.There is also no deletion in sperm donors with normal semen.It is also found that there is a high incidence of chromosome karyotype abnormality in sterile men with Y chromosome microdeletion,there may be a correlation.Through the pedigree analysis of the results from father and son,it was found that AZFc deletion and partial deletion of AZFb could be transmitted vertically to the children.The above results suggest,the Y chromosome microdeletion test should be carried out in patients with infertile male,especially before treated with assisted reproductive techniques.Genetic testing and genetic counseling should be following to develop individualized treatment plans.Because it is challenge for the STS-PCR technique to detect of partial deletion and copy number variation,and it is not possible to specify the specific range and size of the missing or repeated detection.We hope to establish a new detection technology that can achieve these targets.Therefore,the newly established detection method is achieved by combining the target area probe capture and next-generation sequencing(NGS)technology.By comparing with the PCR method,we found that the NGS method we established was a high-throughput,high-resolution innovative technology to detect the deletion or variation of the Y chromosome AZF.Its sensitivity and specificity meet the requirements of clinical testing.This method can detect the AZF deletion pattern provided by PCR method,and can also detect the new copy number deletion,repetition and other structural abnormalities,and provide the specific scope and size.Through family analysis,we found that there are genetic mechanisms such as new mutations,vertical transmission,and enlarged deletion.The results suggest that the new NGS detection technology can replace the original PCR technology.We should pay more attention to the study on the physiological function of Y chromosome deletion or duplication.For some structural abnormalities of deletion or duplication,genetic factors that may cause other synergistic effects can lead to the accumulation effect of harmful mutations,resulting in sterility and infertility.We further analyze the embryonic development and clinical outcome assisted reproductive technology(ART)treatment with ejaculated sperm for patients with AZFc or AZFb.And we found patients with Y chromosome AZFc area/AZFb deletions can have their own children.In the treatment of ICSI,the average sperm concentration of AZF group was significantly lower than that in the control group,with statistical difference(P<0.05).There were no statistically significant differences in Fertilized oocyte rate,2PN Cleaved embryos rate,high quality embryo rate,blastocyst formation rate,clinical pregnancy rate and birth rate,etc.(P >0.05).To sum up,the newly established NGS method can replace the STS-PCR method to detect microdeletion on Y chromosome.It can detect other abnormality of copy number variation,such as partial deletion or duplication,and provide abnormal range and size.Family genetic analysis suggests that defects can occur in patients with new mutations,vertical transmission and to expand,and copy number changes(absent or repeated)will increase the risk of expanding deletions.Therefore,the test should be used in the routine of sterile male and sperm bank.The Y chromosome microdeletions can procreate their own offspring through assisted reproductive technology,but it should be informed by genetic counseling that there is a risk of genetic deletion or enlargement of male offspring.For now,the present data suggest that AZF deletions patients have no statistical difference in the clinical outcomes of assisted reproductive treatment.