Effects of shRNA-mediated silencing of PSMA7 on cell proliferation and vascular endothelial growth factor expression via the ubiquitin-proteasome pathway in cervical cancer.

This study aims to evaluate the effects of PSMA7 silencing on cervical cancer (CC) cell proliferation and vascular endothelial growth factor (VEGF) expression through the ubiquitin-proteasome pathway. CC tissues (n = 43) and normal tissues (n = 27) were first collected from patients. Human CC cell line (SiHa) and human normal cervical epithelial cells (H8) were obtained and classified into the normal, blank, negative control (NC), PSMA7-shRNA1, and PSMA7-shRNA2 groups, respectively. In situ hybridization was used to detect the expressions of wild-type and mutant p53 proteins. Immunofluorescence assay was carried out to test the activity of 20S proteasomes. Reverse transcription quantitative polymerase chain reaction and Western blot analysis were both performed to determine the expressions of PSMA7, ubiquitin, P27, P53, and VEGF in sample tissues and cells. 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was used to analyze cell proliferation rates, and flow cytometry was used to analyze the cell cycle and the apoptotic rate. Compared with normal tissues, CC tissues showed increased expression levels of PSMA7, ubiquitin, p53, VEGF as well as increased activity of 20S proteasomes but exhibited a decrease in p27 expression. Compared with the blank and NC groups, the PSMA7-shRNA1 and PSMA7-shRNA2 groups all had decreased expression levels of PSMA7, ubiquitin, p53, and VEGF as well as decreased cell proliferation, 20S proteasomes activity, and cell number in the S phase, increased p27 expression, cell apoptosis and cell number in the G0/G1 phase. Our study demonstrated that PSMA7 silencing can suppress CC cell proliferation and VEGF expression in addition to promoting cell apoptosis through inhibiting the UPP signaling pathway.

Epidemiology and genotype distribution of human papillomavirus (HPV) in women of Henan Province, China.

BACKGROUND: Human papillomavirus (HPV) infection is the commonest sexually transmitted infection, which is associated with various clinical conditions. This study aimed to determine the distribution of HPV genotypes in the women of Henan Province, China. METHODS: Cervical samples were collected by liquid-based method and consecutively evaluated cervical cytology and the presence of HPV DNA. Cytological classification was made according to the Bethesda 2001 criteria. HPV DNA was tested with xMAP technology by Luminex200™. RESULTS: In cervical abnormalities, the infection rate of HPV was 84.0%, single type was 71.0%, multiple type was 13.0%, high risk HPV was 78.0% and low risk HPV was 8.0%. The most common genotypes found were HPV16, 52, 58, 33, 18, 6 and 39. The most common HPV genotypes were HPV16, 52, 6, 58 and 33 in NILM, HPV16, 52, 18, 58 and 6 in ASCUS, HPV52, 16, 58, 6 and 39 in LSIL, HPV16, 33, 58, 18 and 51 in HSIL, and HPV16, 18, 33, 58 and 52 in ICC, respectively. The prevalence of single HPV and multiple HPV was 64.8% and 13.3%, respectively. Age-specific prevalence of multiple HPV exhibited a "U" shaped curve. CONCLUSIONS: Single HPV genotype infection was predominantly detected in different groups of cervical lesions in Henan Province, and HPV16, 52, 58, 33, 18 and 6 were the priority HPV types.

[Correlation of toll-like receptor 3 and tumor necrosis factor-alpha with idiopathic fetal growth restriction.].

OBJECTIVE: To investigate the expression and the significance of toll-like receptor 3 (TLR-3) in placenta, tumor necrosis factor-alpha(TNF-alpha) in maternal and cord blood of idiopathic fetal growth restriction (IFGR), and their correlation with the pathogenesis of symmetric and asymmetric IFGR. METHODS: From April 2008 to April 2009, 42 primiparae of singleton pregnancy and their IFGR babies, who delivered at term through cesarean section, in the Third Affiliated Hospital of Zhengzhou University were enrolled. All subjectects were divided into symmetric IFGR group (n = 20) and asymmetric IFGR group (n = 22). Another 42 non-IFGR pairs were randomly selected as the control group. The polink-2 plus polymerized horseradish peroxidase (HRP) immunohistochemical method and the enzyme linked immunosorbent assay (ELISA) were applied to detect TLR-3 and TNF-alpha levels. RESULTS: (1) The expression of TLR-3 protein were observed in all maternal placenta of the three groups. TLR-3 essentially expressed in syncytiotrophoblasts and hofbouer cells in the symmetric IFGR and control group, but expressed mostly in hofbouer cells and less in syncytiotrophoblasts in the asymmetric IFGR group. (2) The expression of TLR-3 in the syncytiotrophoblasts of the symmetric and asymmetric IFGR group was significantly lower than in the control group (111 +/- 14 and 118 +/- 11 vs. 156 +/- 9, P < 0.01). The number of TLR-3 positive in Hofbourer cell in the symmetric IFGR group was lower than the control group (8.9 +/- 2.8 vs 17.5 +/- 2.8, P < 0.01), but the number in the asymmetric IFGR group was higher (23.8 +/- 3.7) compared with the control group (P < 0.01). (3) The TNF-alpha levels in the maternal and cord blood of the symmetric and the asymmetric group were higher than that of the control group [maternal: (90 +/- 10) microg/L and (86 +/- 11) microg/L vs. (73 +/- 9) microg/L; cord blood: (92 +/- 12) microg/L and (96 +/- 8) microg/L vs. (79 +/- 9) microg/L; P < 0.01]. (4) Neither symmetric nor the asymmetric IFGR group showed any correlations between the maternal and cord blood levels of TNF-alpha (P > 0.05). (5) Significant correlation was found between the TNF-alpha level of the cord blood and TLR-3 expression in the placenta in both the symmetric and asymmetric IFGR group (P < 0.05), but no relationship was found between the maternal blood TNF-alpha level and TLR-3 expression in the placenta (P > 0.05). CONCLUSIONS: The variantions of TLR-3 expression in placenta and the increased expression of TNF-alpha in cord blood are associated with the genesis IFGR. The reduced expression of TLR-3 may related to symmetric IFGR, while the increased TLR-3 level in hofbouer cells may lead to asymmetric IFGR.

[Expression of transforming growth factor-beta 1, vascular cell adhesion molecule-1 and endothelium-selectin in placenta of patients with pre-eclampsia].

OBJECTIVE: To study the change and significance of the expression of transforming growth factor-beta 1 (TGF-beta1), vascular cell adhesion molecule-1 (VCAM-1) and endothelium-selectin (E-selectin) in placenta of patients with pre-eclampsia. METHODS: Twenty normal pregnant women (control group) and 40 women with pre-eclampsia (pre-eclampsia group, including 16 women with mild pre-eclampsia and 24 women with severe pre-eclampsia) were selected. The cellular distribution of TGF-beta1, VCAM-1 and E-selectin in placenta in both groups was determined by immunohistochemistry, and the mean density was measured by computer image analysis system. RESULTS: (1) The level of TGF-beta1 in placental villous syncytiotrophoblast of pre-eclampsia group (70.7 +/- 0.5) was significantly higher than that of control group (70.3 +/- 0.6), while the level of VCAM-1 and E-selectin in pre-eclampsia group (VCAM-1: 82.5 +/- 0.5, E-selectin: 53.5 +/- 0.5) was significantly lower than that of control group (VCAM-1: 82.8 +/- 0.3, E-selectin: 53.8 +/- 0.4) (P < 0.05). However, there were no significant differences in women with mild pre-eclampsia (TGF-beta1: 70.6 +/- 0.6, VCAM-1: 82.4 +/- 0.6, E-selectin: 53.4 +/- 0.5) and severe ones (TGF-beta1: 70.8 +/- 0.4, VCAM-1: 82.6 +/- 0.5, E-selectin: 53.6 +/- 0.5) (P > 0.05); (2) The level of E-selectin in placental villous capillary endothelial cells of pre-eclampsia group (63.0 +/- 0.5) was significantly higher than that of control group (62.6 +/- 0.4) (P < 0.05), while there was no significant difference in women with mild pre-eclampsia (63.2 +/- 0.4) and severe pre-eclampsia (62.9 +/- 0.5) (P > 0.05). CONCLUSION: TGF-beta1, VCAM-1 and E-selectin are not only related to placenta shallow bed of pre-eclampsia, but also participate in pathogenic process of vascular endothelial damage of pre-eclampsia.

[Hemeoxygenase expression and activity in human placenta of pregnancy induced hypertension].

OBJECTIVE: To investigate the expression of heme oxygenase-1 (HO-1) and heme oxygenase-2 (HO-2) in placenta tissue of pregnancy induced hypertension (PIH), and the relationship between HO protein expression and enzymatic activity. METHODS: Protein expression was analyzed qualitatively and semi-quantitatively by Western blotting in placental tissue of PIH (PIH group, n = 30) and normal late pregnant women (control group, n = 30). The levels of HO enzymatic activity in placental tissue were measured with the double wavelength scanning by spectrophotometer. RESULTS: (1) Western blotting analysis demonstrated that the relative protein levels of HO-1 and HO-2 in placental samples of control group were 0.7 +/- 0.4 and 0.8 +/- 0.4 respectively, there were no significant difference between HO-1 and HO-2 protein levels (P > 0.05). (2) The relative levels of HO-1 were 0.6 +/- 0.4 in PIH group, there was no significant difference from those in the control group (P > 0.05); the relative levels of HO-2 protein were 0.6 +/- 0.3 in PIH group, that were obviously lower than those in the control group (P < 0.01). The levels of HO enzymatic activity of PIH group were (0.3 +/- 0.2) nmol x mg(-1) x h(-1), significantly lower than that of the control group [(0.5 +/- 0.3) nmol x mg(-1) x h(-1)] (P < 0.01). The levels of HO activity correlated with HO-2 protein levels. CONCLUSIONS: The expression levels of HO-2 protein were decreased and HO enzymatic activity reduced in PIH group. HO may play a role in the pathophysiology of poor placental perfusion and tissue damage in placenta of PIH.

[Expression and tissue localization of hemeoxygenase in human placenta].

OBJECTIVE: The purpose of this study was to investigate the expression and localization of the two known isoforms of hemeoxygenase (HO) in normal human first trimester placenta and third trimester placenta. METHODS: Reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry were resorted to demonstrate the expression and localization of HO-1 and HO-2 in normal placenta tissue, obtained from 6 approximately 10 week gestation women (20 cases) and the third trimester woman (20 cases). RESULTS: Compared with glyceraldehydes-3-phosphate dehydrogenase (GAPDH), the expression of HO-1 was lower, there was no significant difference between the first trimester (0.31 +/- 0.19) and third trimester (0.28 +/- 0.14) (P > 0.05); the expression of HO-2 was higher, it is significantly higher at third trimester (1.12 +/- 0.58) compared with first trimester placenta (0.70 +/- 0.48) (P < 0.05). The result of immunohistochemistry demonstrated that HO-1 was predominantly localized in villous stroma cell and trophoblast; HO-2 predominantly localized in trophoblast as well as capillaries, with weak staining of villous stroma. The staining score were not normally distributed. The median staining scorse of HO-1 in trophoblast, villous stroma and capillaries at first trimester were 9.0, 2.6 and 2.8, respectively, at third trimester were 8.7, 2.0 and 1.4, there was no difference between the two groups (P > 0.05). The median staining score of HO-2 in capillaries at first trimester was 5.8, significantly lower than that of the third trimester (9.3) (P < 0.05). There was no significant difference between the staining score of HO-2 in trophoblast (10.5, 8.0) and villous stroma (3.6, 2.4) between the first trimester and the third trimester (P > 0.05). CONCLUSIONS: HO-1 and HO-2 as endogenous system may regulate feto-placental circulation, indicated their different roles in placental vascular development and regulation. They may offer protection against cyto-toxic damage in the placenta, and influence immunological function.