AZF gene expression analysis in peripheral leukocytes and testicular cells from idiopathic infertility.
The aim of this study was to assess the frequency of AZF microdeletions in peripheral leukocytes and testicular cells in Chinese men with idiopathic infertility. Expression in testicular cells was also determined. In this study, we screened 62 idiopathic infertile patients, in whom karyotype, sperm count and hormonal parameters were evaluated. Genomic DNA was extracted from the peripheral leukocytes. Molecular analysis was performed by two multiplex polymerase chain reactions (PCR) using a set of eight sequence tagged sites (STS) from 3 different regions of the Y chromosome. Total cellular RNA was extracted from the testicular tissue using a Trizol-method. Reverse Transcription (RT) reactions were performed to synthesize cDNA. Amplification of DFFRY, RBM and DAZ genes was performed to analyze their expression in testicular cells. In this cohort, we found 12 submicroscopic deletions (12/62, 19.4%). Nine patients (9/33, 27.2%) were detected in the azoospermic group and three (3/29, 10.3%) in the severe oligozoospermic group. RT-PCR analysis from testicular cells gave normal amplifications for SRY and DFFRY mRNA in 62 idiopathic patients; two patients were negative for RBM expression; no RBM and DAZ were detected for a case; 12 patients had no expression in the AZFc region involving the DAZ gene. Of 12 cases, three patients with normal PCR analysis of DAZ gene on genomic DNA showed no RT-PCR amplification for DAZ mRNA. The use of RT-PCR of specific spermatid expressed genes in conjunction with examining microdeletions using peripheral leukocytes is suggested to avoid the transmission of the Y chromosomal microdeletions from a father to a son via testicular sperm aspiration (TESE), intracytoplasmic sperm injection (JCSI).
Song
Ning-Hong
NH
Department of Urology, First Affiliated Hospital, Nanjing Medical University, Nanjing, China.
Yin
Chang-Jun
CJ
Zhang
Wei
W
Zhuo
Zuo-Min
ZM
Ding
Guan-Xiong
GX
Zhang
Jing
J
Hua
Li-Xin
LX
Wu
Hong-Fei
HF
D041322:Chromosomes, Human, Y; D017931:DNA Primers; D015870:Gene Expression; D005787:Gene Frequency; D056426:Genetic Loci; D006801:Humans; D007248:Infertility, Male / Q000235:genetics* / Q000503:physiopathology; D007621:Karyotyping; D007962:Leukocytes / Q000502:physiology*; D008297:Male; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029589:Seminal Plasma Proteins / Q000235:genetics*; D017384:Sequence Deletion; D012729:Sex Chromosome Aberrations; D025064:Sex Chromosome Disorders / Q000235:genetics; D013737:Testis / Q000503:physiopathology*
D017931:DNA Primers; D029589:Seminal Plasma Proteins; D012313:RNA