Sperm-egg fusion disorder in a Chinese male patient was associated with a rare ADAM20 variant.

We report here a 28-year-old male with infertility. No abnormality was found in his semen examination. The couple achieved a successful pregnancy under the help of intracytoplasmic sperm injection during which we found that sperm could enter the zona pellucida, but could not fuse with the egg within the short insemination period. We then performed whole-exome sequencing technology on this patient and found a rare variant (c.641A>C:p.D214A) in ADAM20, which encoded a disintegrin and metalloprotease 20 protein. To further verify the pathogenicity of this variant, we analyzed ADAM20 protein expression in spermatozoa by immunostaining analysis, which showed a mis-localization of ADAM20 in the patient's spermatozoa. Therefore, we concluded that mutation in ADAM20 may be associated with sperm-egg fusion disorder in this patient.

Genetic contribution of SUN5 mutations to acephalic spermatozoa in Fujian China.

Acephalic spermatozoa is an extremely rare disease associated with primary infertility. A recent study showed that genetic alterations in the SUN5 gene lead to this disease, and SUN5 mutations could explain the disease in about half of the patients. Therefore, in the present study, to re-visit the genetic contribution of SUN5 mutations to acephalic spermatozoa, we recruited 15 unrelated affected individuals and screened the SUN5 gene for mutations by whole-exome sequencing (WES) and Sanger sequencing. Five of the 15 (33.33%) subjects were found to carry the same homozygous mutation in the SUN5 gene c.381delA (p.V128Sfs*7). Neither homozygous nor compound heterozygous mutations in SUN5 were found in the other 10 patients. The c.381delA mutation resulted in the truncation of the SUN5 protein and decreased the expression and altered the distribution of the outer dense fiber 1 (ODF1) protein. Thus, in our study SUN5 mutations accounted for only one-third of the patients in our cohort, which is lower than the percentage reported previously. Thus, our study suggests that the contribution of SUN5 mutations to acephalic spermatozoa might not be as high as described previously. These results will help in the genetic counseling of patients with acephalic spermatozoa.

A novel mutation in HAUS7 results in severe oligozoospermia in two brothers.

Severe oligozoospermia (SO) is a common disease resulting in male infertility; however, its pathophysiology remains unclear. Here, we report two brothers with SO. Whole-exome sequencing (WES) identified a hemizygous variant in HAUS7 (c.G386T:p.G129V), an X-linked gene. HAUS7 has been reported to play a role in the meiotic maturation and chromosome alignment of germ cells. The two patients inherited this variant from their mother, and this variant was considered to be a highly pathogenic mutation by in silico analysis. Moreover, in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) was carried out in both the proband's wife and the brother's wife, but they failed to become pregnant after the embryo transfers. Therefore, this novel mutation in HAUS7 gene may be associated with severe oligozoospermia.

A Case of Bilateral Testicular Tumors Subsequently Diagnosed as Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency.

21-hydroxylase deficiency (21-OHD) caused congenital adrenal hyperplasia (CAH) is a group of autosomal recessive genetic disorders resulting from mutations in genes involved with cortisol (CO) synthesis in the adrenal glands. Testicular adrenal rest tumors (TARTs) are rarely the presenting symptoms of CAH. Here, we describe a case of simple virilizing CAH with TARTs, in a 15-year-old boy. The patient showed physical signs of precocious puberty. The levels of blood adrenocorticotropic hormone (ACTH), urinary 17-ketone steroids (17-KS), dehydroepiandrosterone sulfate (DHEA-S), and serum progesterone (PRGE) were elevated, whereas those of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and CO were reduced. Computed tomography (CT) of the adrenal glands and magnetic resonance imaging (MRI) of the testes showed a soft tissue density (more pronounced on the right side) and an irregularly swollen mass (more pronounced on the left side), respectively. Pathological examination of a specimen of the mass indicated polygonal/circular eosinophilic cytoplasm, cord-like arrangement of interstitial cells, and lipid pigment in the cytoplasm. Immunohistochemistry results precluded a diagnosis of Leydig cell tumors. DNA sequencing revealed a hackneyed homozygous mutation, I2g, on intron 2 of the CYP21A2 gene. The patient's symptoms improved after a three-month of dexamethasone therapy. Recent radiographic data showed reduced hyperplastic adrenal nodules and testicular tumors. A diagnosis of TART should be considered and prioritized in CAH patients with testicular tumors. Replacement therapy using a sufficient amount of dexamethasone in this case helps combat TART.

Apoptosis induced by aqueous extracts of crocodile bile in human heptacarcinoma SMMC-7721.

In the present study, effects of aqueous extracts from Crocodylus siamensis bile (AE-CB) on SMMC-7721 cell growth, cell cycle, and apoptosis were investigated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, inverted microscopy, fluorescence microscopy, propidium iodide (PI) single- and fluorescein isothiocyanate (FITC)/PI double-staining flow cytometry, and western blotting. Our data have revealed that AE-CB significantly inhibited the growth of SMMC-7721 cell and arrested cell cycle at G0/G1 phase. SMMC-7721 cells showed typical apoptotic morphological changes after treated with AE-CB for 48 h. Cell death assay indicated that SMMC-7721 cells underwent apoptosis in a dose-dependent manner induced by AE-CB. In addition, AE-CB treatment could downregulate the protein level of Bcl-2 and upregulate the Bax, leading to the increase in the ratio of Bax to Bcl-2 in SMMC-7721 cells. Meanwhile, it was observed that the expression of Survivin and c-Myc decreased, but the expression of P53 increased. All these events were associated with increase of reactive oxygen species. The data indicated that mitochondrial pathway might play an important role in bile extract-induced apoptosis in SMMC-7721 cells. These results provide significant insight into the anticarcinogenic action of bile extract on SMMC-7721 cells.

Antiproliferation and apoptosis induced by tamoxifen in human bile duct carcinoma QBC939 cells via upregulated p53 expression.

Tamoxifen (TAM) is a nonsteroidal antiestrogen that has been used in the treatment of breast cancer for over 30years. Recently, it was shown that TAM also has efficacy on gastrointestinal neoplasms such as hepatocarcinoma and pancreatic carcinoma, and that the chemopreventive activities of TAM might be due to its abilities to inhibit cell growth and induce apoptosis. In the present study, we investigated the effects of tamoxifen on growth and apoptosis in the human bile duct carcinoma (BDC) cell line QBC939 using MTT assay, inverted microscopy, fluorescence microscopy, transmission electron microscopy, classic DNA fragmentation agarose gel electrophoresis assay, PI single- and FITC/PI double-staining flow cytometry, and Western blotting. Our data revealed that TAM could significantly inhibit growth and induce apoptosis in QBC939 cells. Increased expression of p53 was observed in TAM-treated cells, indicating that p53 might play an important role in TAM-induced apoptosis in QBC939 cells. These results provide significant insight into the anticarcinogenic action of TAM on BDC.