Changes in human umbilical vein endothelial cells induced by endothelial nitric oxide synthase traffic inducer.

This study investigated the changes in human umbilical vein endothelial cells (HUVECs) induced by overexpression of endothelial nitric oxide synthase traffic inducer (NOSTRIN) and its role in cellular injury. Recombinant NOSTRIN-expressing and empty vectors were transfected into cultured HUVECs, and factor VIII-related antigen was examined by using immunohistochemical analysis. Growth curves were generated for both transfected and untransfected cells and these indicated that the proliferative ability of cells overexpressing NOSTRIN was significantly decreased. The expression of NOSTRIN and eNOS proteins was detected by using Western blot analysis, endothelial NOS (eNOS) activity was assayed by using spectrophotometry, and NO2 (-)/NO3 (-) levels were measured using nitrate reductase. Immunohistochemical analysis demonstrated that all groups expressed NOSTRIN in the plasma membrane and cytoplasm, and Western blot analysis confirmed that NOSTRIN levels were significantly higher in cells transfected with the NOSTRIN plasmid (P<0.01). The activity of eNOS and the levels of NO2 (-)/NO3 (-) were significantly decreased in NOSTRIN overexpressing cells as compared with empty vector and untransfected cells (P<0.01 and P<0.01, respectively). Morphological and ultrastructural changes were observed under light and electron microscopy, and it was found that NOSTRIN-overexpressing cells were elongated with deformities of the karyotheca, injury to the plasma membrane, increased lipids in the cytoplasm, and shortened microvilli. This study showed that overexpression of NOSTRIN had a significant effect on eNOS activity in HUVECs and resulted in significant cellular damage.

[Testicular CR16 and spermatogenesis].

Spermatogenesis is a complex regulatory process depending on a variety of hormones (such as FSH, LH, T, and 17beta estradiol), cytokines, and genes. Research on gene regulation in spermatogenesis has become a hot spot and revealed some spermato-genesis-related genes, such as AYZ, DAZ, YRRM, NOSTRIN, and so on. Reports are rarely seen on the role of CR16 in male reproduction, and its action mechanism in spermatogenesis is not yet clear. This article updates the role of CR16 in spermatogenesis in the male reproductive system from the perspective of Sertoli cells forming a blood-testis barrier.

[Expression of NOSTRIN in the testis tissue of azoospermia patients].

OBJECTIVE: To determine the expressions of endothelial nitric oxide synthase traffic inducer (NOSTRIN) and endothelial nitric oxide synthase (eNOS) in the testis tissue of azoospermia patients, and investigate their correlation with the pathogenesis of azoospermia. METHODS: We detected the expressions of NOSTRIN and NOSTRIN mRNA by immunohistochemistry and RT-PCR respectively, determined the activity of eNOS by spectrophotometry, and measured the stable metabolic end product NO, NO2- / NO3-, by nitrate reductase assay in the testis tissues of 17 patients with idiopathic azoospermia (the azoospermia group) and 10 normal men (the normal group). RESULTS: NOSTRIN and NOSTRIN mRNA were expressed in the spermatogonia, sertoli cells, stromal cells and vascular endothelial cells, more lowly in the azoospermia than in the normal group (0.312 +/- 0.076 versus 0.793 +/- 0.082, P < 0.01). The activity of eNOS was significantly increased in the idiopathic azoospermia patients ([33.727 +/- 3.58] U/mg) compared with the normal men ([17.69 +/- 3.84] U/mg) (P < 0.01). The level of NO2- / NO3- was significantly higher in the azoospermia than in the normal group ([48.56 +/- 8.49] micromol/L versus [25.37 +/- 9.61] micromol/L, P < 0.01). The expression of NOSTRIN showed a significant negative correlation with the activity of eNOS (r = -0.57, P < 0.01) as well as with the level of NO2- / NO3- (r = -0.61, P < 0.01) in the testis tissue of the idiopathic azoospermia patients. CONCLUSION: The expression of NOSTRIN is decreased, while the activity of eNOS and the level of NO2- / NO3- increased in the testis tissue of azoospermia patients, which may be associated with the pathogenesis of azoospermia.

[Expression of CR16 in the testis of patients with idiopathic azoospermia].

OBJECTIVE: To investigate the expression of corticosteroids and regional expression 16 (CR16) in the testis of patients with idiopathic azoospermia and the role of CR16 in spermatogenesis. METHODS: Immunohistochemistry and RT-PCR were used to detect the expression levels of the CR16 protein and mRNA in the testes of 48 patients with idiopathic azoospermia and 10 healthy men. RESULTS: Immunohistochemistry showed that the CR16 protein expressed in the Sertoli cells and spermatids-binding region in the epithelium of seminiferous tubules. The level of the CR16 protein was markedly lower in the idiopathic azoospermia patients than in the healthy men, and RT-PCR also showed a significantly decreased level of CR16 mRNA in the testis of the patients. CONCLUSION: The expressions of the CR16 protein and mRNA decrease markedly in the testis of patients with idiopathic azoospermia, indicating a correlation with the pathogenesis of azoospermia.

Plasma sFlt-1-to-PlGF ratio is correlated with inflammatory but not with oxidative stress in Chinese preeclamptic women.

PURPOSE: Considerable interest has been focused on angiogenic factors and angiogenic imbalance in the field of pre-eclampsia (PE), owing to its gaining role in the development of PE. This study was addressed to investigate the associations of sFlt-1-to-PlGF plasma ratios with oxidative stress assessed by the level of 8-isoprostane, and inflammation measured by the level of high-sensitive C-reactive protein (hs-CRP), and adipocytokines. METHODS: A total of 83 patients with PE including 47 mild PE (MPE) and 36 severe PE (SPE) and 50 age-matched normotensive subjects in the third trimester of pregnancy were examined. Measurements included body mass index (BMI), systolic and diastolic blood pressure (BP) levels, plasma concentrations of hs-CRP, 8-isoprostane, adiponectin, and leptin. RESULTS: Subjects with PE had higher levels of sFlt-1/PlGF (P < 0.01), hs-CRP (P < 0.01), 8-isoprostane and leptin (both P < 0.01) and lower adiponectin (P < 0.01) than did normotensive control subjects. Significant positive correlations were found between plasma sFlt-1/PlGF and hs-CRP (r = 0.437, P < 0.01) or leptin (r = 0.656, P < 0.01). A weak inverse correlation emerged between sFlt-1/PlGF and adiponectin (r = -0.306, P < 0.01). When a multiple regression analysis was performed, with sFlt-1/PlGF as a dependent variable and all the other parameters as independent variables, sFlt-1/PlGF maintain a significant relationship with leptin (beta = 0.219, P < 0.05) and with hs-CRP (beta = 0.295, P < 0.01) as well as with systolic BP(beta = 0.446, P < 0.05). CONCLUSIONS: In Chinese preeclamptic women, plasma sFlt-1-to-PlGF ratio is correlated with inflammatory and adipocytokines but not with oxidative stress.

Comparison of three methods for the gene analysis of fetal cells from maternal peripheral blood.

BACKGROUND: Although great advances in techniques for noninvasive prenatal diagnosis using fetal cells from maternal peripheral blood have achieved, current technology does not meet the demands required for clinical use. In this study, we aimed to establish reliable methods for the gene analysis of fetal cells from maternal peripheral blood. METHODS: Primed extension preamplification (PEP)-polymerase chain reaction (PCR), multiple primed in situ labeling (PRINS), and nested PCR were individually applied to detect the sex determining region Y (SRY) gene in single fetal cells collected from maternal peripheral blood. RESULTS: The sensitivity and specificity of the detection of the SRY gene by PEP-PCR were 97.39% (149/153) and 99.17% (119/120), respectively. The sensitivity and specificity of PRINS were 97.56% (40/41) and 100% (35/35), respectively. The sensitivity and specificity of nested-PCR were 80.00% (24/30) and 87.50% (14/16), respectively. CONCLUSIONS: PEP-PCR and PRINS are reliable techniques for the gene analysis of single fetal cells from maternal peripheral blood because of their high sensitivity and specificity. PEP-PCR and PRINS can be used as standard methods of noninvasive prenatal diagnosis using single fetal cells from maternal peripheral blood.