Observation of spermatogenic cells for infertile patients with Y-chromosomal microdeletion / 中华男科学杂志

<p><b>OBJECTIVE</b>To assess the spermatogenic function of the infertile patients with Y-chromosomal microdeletion.</p><p><b>METHODS</b>Thirty-five 23-44 years old patients with microdeletions of Y chromosome were included in this study. Three semen analyses confirmed that 26 cases were non-obstructive azoospermia and 9 oligospermia with sperm count < 1 x 10(6)/ml. They were divided into 3 groups by the locus of deletion, 5 cases of AZFa + b + c deletion in group 1, 4 cases of AZFb + c and 3 cases of AZFb deletion in group 2, and 23 cases of AZFc deletion in group 3. Semen was collected and centrifuged, the supernatant removed and the centrifugate applied on the clean slides after dilution. Following Wright's-Giemsa staining, the slides were viewed under the microscope. Testis histopathological biopsy was performed for 6 of the cases.</p><p><b>RESULTS</b>In group 1, no spermatogenic cells were observed but only Sertoli cells in 1 case, with a consistency between the result of spermatogenic cell test and that of testis biopsy. In group 2, spermatogenic cell tests revealed spermatocytes in 6 cases, 2 were proved by testis biopsy with sperm maturation arrest in the primary spermatocyte stage, and spermatogenic cells of all developmental stages were seen in 1 AZFb deletion patient with the same sperm maturation arrest as the above two. In group 3, only primary spermatocytes were detected by spermatogenic cell test in 5 oligospermia patients but no spermatogenic cells in the 15 azoospermia cases, and biopsy revealed 2 cases of Sertoli cell-only syndrome.</p><p><b>CONCLUSION</b>The spermatogenic cell test can effectively assess the spermatogenic function of AZF deletion patients. Non-invasive and easily accepted by patients, it is highly recommendable for the evaluation of spermatogenesis of patients with Y-chromosomal microdeletion.</p>

AZF microdeletions are not related with recurrent spontaneous abortion / 中华男科学杂志

<p><b>OBJECTIVE</b>To study the relationship between azoospermia factor (AZF) microdeletions of the Y-chromosome and recurrent spontaneous abortion.</p><p><b>METHODS</b>We collected 26 chorionic villus samples from abortive male embryos and 51 blood samples from the husbands whose wives had recurrent spontaneous abortion, extracted genomic DNA from the samples and detected 12 STSs in the AZFa, AZFb and AZFc regions of Yq11.2 by amplification multiplex PCR.</p><p><b>RESULT</b>AZF microdeletions were found neither in the chorionic villus samples nor in the blood samples.</p><p><b>CONCLUSION</b>There is no relationship between AZF microdeletions and recurrent spontaneous abortion.</p>

Evaluation of CD4+ CD25+ regulatory T cells in the peripheral blood of recurrent spontaneous abortion patients / 中华男科学杂志

<p><b>OBJECTIVE</b>To explore the role of CD4+ CD25+ regulatory T cells (CD4+ CD25+ Tr) in the pathogenesis of recurrent spontaneous abortion.</p><p><b>METHODS</b>Peripheral blood samples were collected from 29 women with unexplainable recurrent spontaneous abortion (the URSA group) and another 20 with normal pregnancy (the control group). The percentage of CD4+ CD25+ Tr in the peripheral blood was measured by flow cytometry.</p><p><b>RESULTS</b>The rate of CD4+ CD25bright Tr in the URSA patients ([1.98 +/- 0.96]%) was significantly lower than that in the control group ([3.21 +/- 1.25]%, P < 0.05), while the percentages of CD4+ CD25+ and CD4+ CD25dim and the ratio of CD4+ CD25bright/CD4+ were not significantly different between the two groups (P > 0.05).</p><p><b>CONCLUSION</b>URSA might be associated with the decreased percentage of CD4+ CD25bright Tr, which plays an important role in fetomaternal immunologic tolerance.</p>

Direct multiplex-PCR from whole blood for rapid detection of Y chromosome microdeletions / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To establish a rapid and simple method to detect Y chromosome microdeletions directly using whole blood as starting material for multiplex-PCR.</p><p><b>METHODS</b>Using a self-prepared pHpH-Bufferq, multiplex-PCR amplification was directly carried out from the anticoagulant whole blood sample without DNA extraction step. Twelve sequence tagged sites (STS), namely SY84, SY86, SY127, SY134, SY124, SY132, SY152, SY157, SY239, SY242, SY254 and SY255, in AZFa, AZFb, and AZFc gene regions were detected in 5 different tubes. In order to ensure the validity of the experiments, sex-determining region Y (SRY) and X-linked or Y-linked zinc finger gene (ZFX/Y) were used as internal controls. Furthermore, conventional PCR using genomic DNA extracted from each blood sample was performed in parallel for evaluating the accuracy of the experiments.</p><p><b>RESULTS</b>A total of 156 male samples were detected, and a normal male sample and a female sample were used as a positive and a negative control respectively. The results showed that 144 samples had no deletion; one sample was AZF-deleted; one sample was AZFb-deleted; seven samples were AZFc-deleted; one sample was both AZFb- and AZFc- deleted; and two samples were all AZFa-, AZFb- and AZFc- deleted. The observed results from two kinds of starting material (whole blood and purified DNA) are completely consistent.</p><p><b>CONCLUSION</b>In our method, PCR amplification was directly carried out from whole blood without any DNA extraction step. So it has the advantages of low cost, simple process, time-saving operation and less cross-contamination. The whole process can be completed within 2 hours. Thus the efficiency of clinical detection is improved greatly.</p>

Prenatal diagnosis of two pregnancies with risk of chromosomal disorders / 中华男科学杂志

<p><b>OBJECTIVE</b>To report the prenatal diagnosis of 2 cases of pregnancy with the risk of chromosomal disorders. In Case 1, the pregnant woman had a daughter with testicular regression syndrome and a segmental duplication of Ypter --> Yp11.2 and a deletion of Yq11.23 --> Yqter. In Case 2, both the pregnant woman and her husband were carriers of chromosomal balanced translocation.</p><p><b>METHODS</b>Two samples of amniotic fluid were obtained at the 19th week of gestation for fetal karyotype analysis. For Case 1, FISH with a probe of Xp/Yp subtelomere was performed on the metaphase of the amniotic fluid, genomic DNA of the amniotic fluid extracted and multiplex PCR conducted for AZF regions. Both the pregnant women underwent sonography to confirm the karyotypic diagnosis.</p><p><b>RESULTS</b>Cytogenetic, FISH and multiplex PCR analysis of the cultured amniotic fluid cells from Case 1 showed a normal male karyotype, and ultrasound scan of the fetus showed normal male external genitalia and normal development. Cytogenetic analysis of the cultured amniotic fluid cells from Case 2 revealed a karyotype of balanced translocation with t(13 ; 14) from the father, and no abnormality of the fetus was found by ultrasound scan.</p><p><b>CONCLUSION</b>It is helpful to perform cytogenetical and molecular prenatal diagnosis in combination with ultrasound scan for the fetus with the risk of chromosomal disorders and subsequently for genetic counseling.</p>

Clinical, molecular and cytogenetic studies on 4 patients with 46, XX (SRY positive) male syndrome / 中华男科学杂志

<p><b>OBJECTIVE</b>To analyze the clinical, molecular and cytogenetic features of 46, XX (SRY positive) male syndrome.</p><p><b>METHODS</b>The clinical features of 4 patients with 46, XX (SRY positive) male syndrome were analyzed retrospectively. Karyotyping, FISH, PCR amplification of the SRY gene, and Y-chromosome microdeletion were performed to study their molecular cytogenetic features.</p><p><b>RESULTS</b>The Four patients were all sociopsychologically males of short stature and came to hospital for infertility. Physical examination revealed that their testes were small in volume and soft in texture, but their penes were normal. Semen analyses showed complete azoospermia. Detection of serum sexual hormone suggested hypergonadotropic hypogonadism. All were karyotyped as 46, XX. Molecular analyses revealed the presence of the SRY gene and absence of AZFa, b and c of the Y chromosome. FISH analysis showed that SRY genes were translocated to Xp in 3 of the patients.</p><p><b>CONCLUSION</b>Phenotypically 46, XX (SRY positive) male patients are males generally, for the presence of the SRY gene in the whole genome and azoospermia due to the deletion of AZF. The clinical characteristics of the patient include testis dysgenesis, infertility and short stature. The long arm of the Y chromosome might contain the gene associated with body height. Extensive molecular and cytogenetic studies on 46, XX male syndrome may help to elucidate its genotype-phenotype relation.</p>