Hereditary risk in icsi with sperm from non-mosaic Klinefelter syndrome patients

To verify the actual risk of hereditary Klinefelter Syndrome (KS) or incidental aneuploidy in ICSI treatment of KS patients, which has been warned in previous cytogenetic studies using testicular cells from KS patients. Cytogenetic analysis in KS patients, their testicular cells and delivered babies. 1) 45 babies who were born by ICSI treatment of KS patients in our hospital underwent chromosomal analysis using amniocentesis or peripheral blood samples. 2) Fluorescent in-situ hybridization (FISH) analysis for sex chromosome constitution was carried out in morphologically normal meiotic cells or mature sperm which were identified and isolated from testicular cell suspension in 5 or 10 KS patients, respectively. 3) Using blood samples of 4 KS patients and their parents and 18 KS patients, the origin of the extra X chromosome was determined with X-chromosome haplotype markers, according to the method by Shrivastava et al. (2014). DNA was extracted from blood samples of KS patients and their parents with consent. Multiplexed PCR amplifications of the 12 X-STR loci were conducted using an Investigator Argus X-12 QS Kit (Quigen). Electrophoresis was run on an ABI PRISM 3100 Genetic Analyzer for the PCR products. The data obtained was analyzed with GeneMapper ID software. 1) No chromosomal abnormality was found in 45 KS patient’s babies examined. 2) In the most of 5 KS patient’s testes examined, spermatogonia showed XY and XXY mosaicism. However, the sex chromosome constitution of all primary spermatocytes and spermatids was normal, suggesting the possibility that XXY spermatogonia can not enter meiosis. In one case, there were no XXY spermatogonia. 3) X-chromosomal STR DNA profiles were compared among KS patient and their parents. In 3 of the 4 KS patients, both two X chromosomes were maternal origin, showing that an extra X chromosome was left in an oocyte as a result of chromosomal non-disjunction at the 1st or 2nd meiotic division. In one patient, X-chromosomes were inherited from parents, suggesting that fertilization of XY-sperm is the cause of KS. In addition, it was surmised that 12 cases in 18 patients showed maternal origin and 6 in 18 patients were paternally. Although the sample number applied for X-chromosomal STR DNA profiling is not enough, the present data may indicate that contribution of XX oocyte to the production of XXY embryos is greater than XY sperm. Namely, a XX oocyte penetration by a Y sperm is the main cause of KS. All data indicates that the risk of KS baby resulting from ICSI treatment of KS patient couples may be a lot lower than expected from the previous studies. Cytogenetic analysis with smear of testicular cell mixture that was used in the studies may overestimate chromosomal abnormality. This finding encourages the opinion that we should strongly recommend vigorous treatment with ART for KS patients.