Reproductive effects of cadmium on sperm function and early embryonic development in vitro.

Cadmium is a major environmental toxicant that is released into the atmosphere, water and soil in the form of cadmium oxide, cadmium chloride, or cadmium sulfide via industrial activities, such as the manufacturing of batteries and pigments, metal smelting and refining and municipal waste incineration. In the present study, we investigated the effects of cadmium exposure on sperm quality parameters, fertilization capacity and early embryonic development. Our study showed that in vitro incubation of human or mouse sperms with cadmium for a long time (up to 24 hours) could significantly decreased sperm motility in a concentration- and time-dependent manner. Exposure to cadmium in the environment for a short term (30 min) did not affect sperm motility but significantly reduced in vitro fertilization rate. We also evaluated the effects of cadmium at concentrations of 0.625 µg/ml, and 1.25 µg/ml on early embryonic development in vitro and observed that the blastocyst formation rate dramatically decreased with increasing cadmium concentration. This finding emphasizes the hazardous effects of cadmium on sperm quality as well as on natural embryo development and raises greater concerns regarding cadmium pollution.

Another functional frame-shift polymorphism of DEFB126 (rs11467497) associated with male infertility.

DEFB126 rs140685149 mutation was shown to cause sperm dysfunction and subfertility. Indel rs11467497 is another 4-nucleotide frame-shift mutation (151bp upstream of rs140685149) that leads to the premature termination of translation and the expression of peptide truncated at the carboxyl terminus. In the present study, we performed a comprehensive association study to check the contribution of rs140685149 and rs11467497 to male infertility. Our results confirmed the previous findings that there was no association between rs140685149 and sperm motility. In contrast, we found a significant association of another indel rs11467497 with male infertility. Moreover, rs11467497 was shown to be associated with higher number of round cells in the infertile males with low sperm motility. Surprisingly, the two mutations commonly existed in the sperm donors (n = 672), suggesting a potential application of the two indels in the screening for eligible sperm donors. Western blotting assays showed the sperms with rs140685149 2-nt deletion tended to have unstable DEFB126 protein in contrast of no DEFB126 protein expressed in the sperms with rs11467497 4-nt deletion, suggesting a more severe consequence caused by rs11467497 mutation. In conclusion, our study presented a significant contribution of another functional frame-shift polymorphism of DEFB126 (rs11467497) to male infertility.

ICSI treatment of severe male infertility can achieve prospective embryo quality compared with IVF of fertile donor sperm on sibling oocytes.

Azoospermia, cryptozoospermia and necrospermia can markedly decrease the ability of males to achieve pregnancy in fertile females. However, patients with these severe conditions still have the option to be treated by intracytoplasmic sperm injection (ICSI) to become biological fathers. This study analyzed the fertilization ability and the developmental viabilities of the derived embryos after ICSI treatment of the sperm from these patients compared with in vitro fertilization (IVF) treatment of the proven-fertile donor sperm on sibling oocytes as a control. On the day of oocyte retrieval, the number of sperm suitable for ICSI collected from two ejaculates or testicular sperm extraction was lower than the oocytes, and therefore, excess sibling oocytes were treated by IVF with donor sperm. From 72 couples (73 cycles), 1117 metaphase II oocytes were divided into 512 for ICSI and 605 for IVF. Compared with the control, husbands' sperm produced a lower fertilization rate in nonobstructive azoospermia (65.4% vs 83.2%; P< 0.001), crytozoospermia (68.8% vs 75.5%; P< 0.05) and necrospermia (65.0% vs 85.2%; P< 0.05). The zygotes derived in nonobstructive azoospermia had a lower cleavage rate (96.4% vs 99.4%; P< 0.05), but the rate of resultant good-quality embryos was not different. Analysis of the rates of cleaved and good-quality embryos in crytozoospermia and necrospermia did not exhibit a significant difference from the control. In conclusion, although the sperm from severe male infertility reduced the fertilization ability, the derived embryos had potential developmental viabilities that might be predictive for the expected clinical outcomes.

Reproductive effects of two neonicotinoid insecticides on mouse sperm function and early embryonic development in vitro.

Acetamiprid (ACE) and imidacloprid (IMI) are two major members in the family of neonicotinoid pesticides, which are synthesized with a higher selectivity to insects. The present study determined and compared in vitro effects of ACE, IMI and nicotine on mammalian reproduction by using an integrated testing strategy for reproductive toxicology, which covered sperm quality, sperm penetration into oocytes and preimplantation embryonic development. Direct chemical exposure (500 µM or 5 mM) on spermatozoa during capacitation was performed, and in vitro fertilization (IVF) process, zygotes and 2-cell embryos were respectively incubated with chemical-supplemented medium until blastocyst formation to evaluate the reproductive toxicity of these chemicals and monitor the stages mainly affected. Generally, treatment of 500 µM or 5 mM chemicals for 30 min did not change sperm motility and DNA integrity significantly but the fertilization ability in in vitro fertilization (IVF) process, indicating that IVF process could detect and distinguish subtle effect of spermatozoa exposed to different chemicals. Culture experiment in the presence of chemicals in medium showed that fertilization process and zygotes are adversely affected by direct exposure of chemicals (P<0.05), in an order of nicotine>IMI>ACE, whereas developmental progression of 2-cell stage embryos was similar to controls (P>0.05). These findings unveiled the hazardous effects of neonicotinoid pesticides exposure on mammalian sperm fertilization ability as well as embryonic development, raising the concerns that neonicotinoid pesticides may pose reproductive risks on human reproductive health, especially in professional populations.

[Quantitative analysis by real-time elastosonography for the differential diagnosis of azoospermia: preliminary application].

OBJECTIVE: To evaluate the quantitative analysis by real-time elastosonography in the differential diagnosis of obstructive azoospermia (OA) and non-obstructive azoospermia (NOA). METHODS: We evaluated the elastosonographic images of 200 cases of OA, 300 cases of NOA and 100 normal healthy controls, calculated the strain ratio of the testis to the scrotal skin and the median strain ratio among the three groups, and analyzed the best cut-off point for differentiating OA and NOA by the receiver operation characteristic (ROC) curve. RESULTS: The median strain ratio of NOA was 0.49 +/- 0.43, while that of OA was 0.35 +/- 0.31, with significant difference between the two groups (Z = - 19.173, P = 0.000 < 0.017). According to the results of ROC curve analysis, the area under the curve was 0.857 +/- 0.012 and the best cut-off point for differentiating OA and NOA was 0.395 (sensitivity = 84.5%, specificity = 74.5%, accuracy = 80.5%). CONCLUSION: Quantitative analysis by real-time elastosonography is a new valuable technique for the differential diagnosis of azoospermia.

Differential diagnosis of azoospermia and etiologic classification of obstructive azoospermia: role of scrotal and transrectal US.

PURPOSE: To evaluate the usefulness of scrotal and transrectal ultrasonography (US) in the differential diagnosis of azoospermia and the etiologic classification of obstructive azoospermia. MATERIALS AND METHODS: This study was approved by the institutional ethics committee, and all patients provided written informed consent. Between April 2006 and November 2008, 100 infertile men (mean age, 32 years; range, 22-51 years) with azoospermia were evaluated at scrotal and transrectal US, with an emphasis on abnormal US findings of the seminal tract and measurement of testicular volume. On the basis of the results of percutaneous epididymal sperm aspiration, testicular fine-needle aspiration cytology, or testicular biopsy, 97 patients with azoospermia were divided into obstructive or nonobstructive groups. The remaining three patients had Klinefelter syndrome and did not undergo testicular biopsy. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic performance of different indexes for discrimination between obstructive and nonobstructive azoospermia. Cases of obstructive azoospermia were further classified according to obstructive causes. RESULTS: Seminal tract abnormalities were more common in obstructive (92.2% [59 of 64 patients]) than in nonobstructive (2.8% [one of 36 patients]; P < .001) azoospermia. Testicular volume was significantly larger for obstructive (median, 16.0 mL; range, 8.4-27.5 mL) than for nonobstructive (median, 8.6 mL; range, 0.9-21.0 mL; P < .001) azoospermia. The area under the ROC curve for discrimination between the groups through combined assessment of the seminal tract and testicular volume at scrotal and transrectal US was 0.96. Sensitivity, specificity, and accuracy for combined assessment in discriminating between obstructive and nonobstructive azoospermia were 95.3% (61 of 64 patients), 97.2% (35 of 36), and 96.0% (96 of 100), respectively. CONCLUSION: Scrotal US and transrectal US are effective imaging modalities for distinguishing obstructive from nonobstructive azoospermia and can provide meaningful diagnostic information for determining the etiologic classification of obstructive azoospermia.

[Ultrasonographic features of epididymides in obstructive azoospermia].

OBJECTIVE: To investigate the ultrasonographic features of epididymides in congenital obstructive azoospermia (COA) and acquired obstructive azoospermia (AOA). METHODS: A total of 211 infertile men with obstructive azoospermia were observed by scrotal ultrasonography, and the features of the epididymal ultrasonograms were compared between COA and AOA. RESULTS: COA exhibited significantly higher rates of ectasia in the epididymal head, cord-like changes, abrupt tapering and absence of the epididymal body and tail than AOA (P < 0.05), while AOA showed markedly higher rates of epididymal body and tail duct ectasia and epididymal inflammatory mass than COA (P < 0.01). Tubular ectasia of the epididymal duct in the head, body and tail were markedly higher in the COA (14 [5.9%], 41 [17.2%] and 20 [8.4%] cases in 236 epididymides) than in the AOA (P < 0.05). Retiform ectasia were markedly higher in the AOA (119 [64.0%], 142 [76.3%] and 109 [58.6%] cases in 186 epididymides) than in the COA (P < 0.05), with statistically significant differences between the two groups (P < 0.05). Ultrasonographically, the epididymides of the COA patients were characterized by irregular ectasia of the epididymal tube with decreased and unclear wall echoes (P < 0.05), and those of the AOA patients by regular ectasia with enhanced wall echoes (P < 0.01). CONCLUSION: The ultrasonographic epididymal features of COA are obviously different from those of AOA, which is of important clinical application value for distinguishing the two conditions from each other.

[The detection of mutations in VHL gene from a single cell in a patient with von Hippel-Lindau disease].

OBJECTIVE: To explore a technology for diagnosing VHL mutations from a single cell and provide experimental evidences for the feasibility of applying technology in detecting genetic mutations from a single cell. METHODS: After whole genome amplification (WGA) based on multiple displacement amplication (MDA) for a single cell, we did regular PCR following sequencing and detected the genotypes using the real time PCR based on TaqMan probes. We detected VHL mutations by the different terminal fluorescent changing. RESULTS: The rate of amplification for single cell based on MDA was 90.91%. The rate of contamination was 0. After sequencing, the allele drop out (ADO) rate of heterozygotes was 26.67%(8/30); combined with the different terminal fluorescent changing, the rate of ADO of heterozygotes was 16.67%. CONCLUSION: WGA based on MDA for a single cell followed by regular PCR with sequencing and real time PCR can specially and accurately detect the VHL genotypes of single cells.

[Male infertility update].

Exact etiological factors cannot be found in more than 70% of male infertility. Although the causes of some male infertility are known, the pathogenesis is not yet clear. Therefore, the diagnosis and treatment of male infertility are particularly complicated. Some breakthroughs have been made in the treatment of refractory male infertility since the development of ICSI in 1992. However, traditional treatment should also be taken into account for individual male infertility. The standardization of diagnosis and treatment and the improvement of the success rate and safety of assistant reproductive technology will be the dominant factors in the development of male infertility researches.

[Clinical analysis of serum endothelin in type 2 diabetic patients with vascular complications].

OBJECTIVE: To explore the clinical significance of changes in urine microalbumin (UM) and endothelin (ET) in different courses of type 2 diabetes. METHODS: A clinical analyses of 30 type 2 diabetic patients with vascular complications (group A) was conducted in comparison with type 2 diabetic patients without vascular complications (group B) and patients with impaired glucose tolerance (IGT, group C). With 30 healthy subjects with family history of type 2 diabetes (group D) and 30 healthy subjects without such family history (group E) as controls, UM and ET were determined in all the subjects for a statistical analysis. RESULTS: Significant differences in UM content was noted between group A and the other 4 groups (P<0.05), while the content did not differ significantly between the latter 4 groups (P>0.05). ET content was significantly higher in group A than in the other 4 groups (P<0.01), and was the lowest in group E (P<0.01). CONCLUSIONS: Type 2 diabetic patients sustain extensively impaired endothelium function, which is exacerbated with the progression of the disease courses. Synchronized changes between vascular complications and impaired endothelium function indicates that vascular endothelial injury relates to vascular complications and the progression of diabetes.

[Relationship between serum vWF and PAF in type 2 diabetic patients and diabetic nephropathy].

OBJECTIVE: To observe the changes in urine albumin (UALB), von Willebrand factor (vWF) and platelet-activating factor (PAF) in different courses of type 2 diabetes. METHODS: Levels of UALB, vWF and PAF were determined in 30 type 2 diabetic nephropathy patients(group A), type 2 diabetic patients without DNP(group B, n=30), patients with impaired glucose tolerance (group C, n=30), the first-degree relatives of type 2 diabetic patients with normal glucose tolerance(group D, n=30) and 30 normal glucose tolerance subjects without family history of type 2 diabetes(group E). RESULTS: UALB and PAF contents were significantly higher in group A than in the other groups (P<0.01), while between the latter 4 groups, the contents showed no significant differences (P>0.05). Serum vWF level was significantly higher in group A than those in the other groups (P<0.05). CONCLUSIONS: Renal dysfunction and platelet activation are present in type 2 diabetes with microvascular complications, and the synchronized changes between renal dysfunction and platelet activation indicate the involvement of elevated PAF in vascular injury.