Fertility in a i(Xq) Klinefelter patient: importance of XIST expression level determined by qRT-PCR in ruling out Klinefelter cryptic mosaicism as cause of oligozoospermia

Abstract

The presence of an isochromosome Xq in Klinefelter syndrome (KS) is an apparently rare condition. In all cases reported so far, patients showed the classic phenotype. We here describe a case of isochromosome Xq [47,X,i(Xq),Y] in a non-mosaic KS patient. The patient exhibited a normal androgenized phenotype, normal testes and normal cognitive abilities. Semen analysis revealed a medium oligozoospermia (5 x 10(6) spermatozoa/ml). After the patient underwent intracytoplasmic sperm injection, he generated two cytogenetically healthy normal females. Fluorescence in situ hybridization analysis showed the presence of a dicentric Xq chromosome that did not show the presence of residual Xp arm up to the 57,820,478 bp position (Xp 1.1) of X chromosome sequence. Preferential inactivation of Xq isochromosome was demonstrated by bromodeoxyuridine replication analysis and transcriptional silencing by DNA methylation at the HUMARA locus. Furthermore, we demonstrated by quantitative RT-PCR an active XIST RNA expression in blood lymphocytes from Klinefelter patients, comparable to that observed in control females and over 30,000-fold greater than in control males. In conclusion, this qRT-PCR approach could be useful for screening of prepuberty males and for diagnosis or exclusion of cryptic Klinefelter mosaics.

Severe XIST hypomethylation clearly distinguishes (SRY+) 46,XX-maleness from Klinefelter syndrome

“…As proposed and indicated earlier (34), KS men in this study undergo appropriate X chromosome inactivation. The striking epigenetic similarities of the X chromosomes of KS patients and women in terms of methylation status and variation suggest that insufficiency and apoptosis of Leydig cell in KS men (7) could be due to the 'female' expression pattern in these cells that could compromise their function.…”

The endocannabinoid system and pivotal role of the CB 2 receptor in mouse spermatogenesis

“…Total RNA was extracted, analyzed by a 2100 Bionalyzer (Agilent) RNA Integrity Number (R.I.N.) Ͼ7.0 and retrotranscribed as described (39). qRT-PCR analysis was performed essentially as described by an iCycler-iQ5 (Bio-Rad) in a 25-l reaction mixture containing 10 -50 ng of cDNA Optimized primers for SYBR-green analysis (GenBank accession nos.…”

Protective role of palmitoylethanolamide in contact allergic dermatitis

“…Total RNA was extracted from either tissue or cells in RNA later, analysed by a 2100 Bioanalyzer (Agilent Technologies, Waldbronn, Germany) (RNA integrity number > 7.0) and retro-transcribed as previously described (44). RNA from CTRL and inflamed ears was obtained from small specimens (5 mg wet tissue weight) of ear skin coming from the experiments carried out both during a previous study (38) and in the present study.…”