Summary： This paper aimed to study the ability of baicalein to block human cytomegalovirus （HCMV） infection in extravillous cytotrophoblasts （EVT） and its effect on the vasoactive intestinal peptide （VIP） expre...Summary： This paper aimed to study the ability of baicalein to block human cytomegalovirus （HCMV） infection in extravillous cytotrophoblasts （EVT） and its effect on the vasoactive intestinal peptide （VIP） expression in HCMV-infected EVT in vitro. A human trophoblast cell line （HPT-8） was chosen in this study. HCMV with 100 TCIDs0was added into culture medium to infect HPT-8 cells, and then HCMV pp65 antigen was assayed by immunofluorescence staining. The infection status was determined by vi- rus titration. Real-time quantitative polymerase chain reaction （qRT-PCR） was used to detect virus DNA load in the infected cells. The expression of VIP mRNA and protein in the infected cells was measured by qRT-PCR, immunocytochemistry and Western blotting. Concentration of VIP secreted in supernatants was determined by ELISA. Red-stained HCMV pp65 antigens were found in infected HPT-8 cells 48 h after infection. HCMV replicated in large quantity in infected HPT-8 cells 4 days after infection, reaching a peak at day 6 post-infection. After treatment with baicalein, virus DNA load in in- fected HPT-8 cells was decreased （P〈0.05）, and the levels of VIP mRNA and protein, and the concen- tration were raised to the normal （P〉0.05）. Our study suggested that baicalein exerts a positive effect on the VIP expression in HCMV-infected EVT at maternal-fetal interface.展开更多
The effect of human cytomegalovirus （HCMV） on invasive capability of early pregnant extravillous cytotrophoblasts （EVTs） was investigated in vitro. Primary EVTs were obtained by complex phosphoesterasum digestion ...The effect of human cytomegalovirus （HCMV） on invasive capability of early pregnant extravillous cytotrophoblasts （EVTs） was investigated in vitro. Primary EVTs were obtained by complex phosphoesterasum digestion and gradient centrifugation from villous tissue aseptically taken from healthy pregnant women. Cytokeratin7 （CK7）, vimentin （Vim） and cerbB-2 were immunocytochemically detected to identify source of cells, and HCMVpp65 antigen was assayed to determine the infection state of primary EVTs by immunocytochemical staining. The EVTs were divided into two groups： control group and HCMV group, and the expression of c-erbB-2, matrix metalloproteinase-2 （MMP-2） and MMP-9 proteins was detected in two groups by immunocytochemistry and Western blotting. Enzymic activity changes of MMP-2 and MMP-9 were tested by gelatin zymography in primary EVTs infected with HCMV. The invasion of primary EVTs was detected by cell invasion assay in vitro after they were infected by HCMV. The cell source identification showed that the cells obtained were highly-pure primary EVTs, and primary EVTs could be infected by HCMV. Primary EVTs could express c-erbB-2, MMP-2 and MMP-9 proteins, and as compared with control group, the protein expression was decreased significantly in HCMV groups （P〈0.05）. Primary EVTs could secrete active MMP-2 and MMP-9 in vitro, and the activity of two MMPs was decreased sig- nificantly in HCMV groups （P〈0.05）. The in vitro cell invasion assay showed that the number of primary EVTs permeating Matrigel in HCMV group was decreased （P〈0.05）. We are led to conclude that HCMV can infect primary EVTs and inhibit their invasion capability, suggesting that the im- paired EVT＇s invasion capability might be related to the abnormal expression of c-erbB-2, MMP-2 and MMP-9 proteins.展开更多
The effects of leptin on cytotrophoblast proliferation and invasion activity were investigated. Immunohistochemistry was used to determine the placental expression of leptin in first-trimester pregnancy. By using RT-P...The effects of leptin on cytotrophoblast proliferation and invasion activity were investigated. Immunohistochemistry was used to determine the placental expression of leptin in first-trimester pregnancy. By using RT-PCR and quantitative real-time PCR, the expression of leptin in cytotrophoblast and the effect of leptin on cytotrophoblast secretion were detected. The potential of cell proliferation, invasiveness and migration was assessed by MTT, Transwell invasion assay and migration assay respectively when the cytotrophoblast was cultured with different concentrations of leptin. The results showed that： （1） Leptin was distributed diffusely around cell membrane, in cytoplasma, and on nuclear mem- brane of cytotrophoblast; （2） Leptin mRNA was expressed in cytotrophoblast. Ten ng/mL leptin could promote the secretion of cytotrophoblast significantly （P〈0.01）; （3） After culture with different concentrations of leptin for 24 h or longer, the proliferation of cytotrophoblast was inhibited, while in 24 h leptin could promote cytotrophoblast invasion and migration. Leptin at a concentration of 500 ng/mL could promote cytotrophoblast invasiveness and migration significantly as compared with controls （P〈0.05）. It was suggested that leptin could inhibit cytotrophoblast proliferation, and promote cytotro- phoblast invasion and migration activity.展开更多
基金supported by grants from the National Natural Science Foundation of China (Nos. 39970769, 30371488,30672243, and 81200354)the Natural Science Foundation of Hubei Province (No. 2009CDD216)
文摘Summary： This paper aimed to study the ability of baicalein to block human cytomegalovirus （HCMV） infection in extravillous cytotrophoblasts （EVT） and its effect on the vasoactive intestinal peptide （VIP） expression in HCMV-infected EVT in vitro. A human trophoblast cell line （HPT-8） was chosen in this study. HCMV with 100 TCIDs0was added into culture medium to infect HPT-8 cells, and then HCMV pp65 antigen was assayed by immunofluorescence staining. The infection status was determined by vi- rus titration. Real-time quantitative polymerase chain reaction （qRT-PCR） was used to detect virus DNA load in the infected cells. The expression of VIP mRNA and protein in the infected cells was measured by qRT-PCR, immunocytochemistry and Western blotting. Concentration of VIP secreted in supernatants was determined by ELISA. Red-stained HCMV pp65 antigens were found in infected HPT-8 cells 48 h after infection. HCMV replicated in large quantity in infected HPT-8 cells 4 days after infection, reaching a peak at day 6 post-infection. After treatment with baicalein, virus DNA load in in- fected HPT-8 cells was decreased （P〈0.05）, and the levels of VIP mRNA and protein, and the concen- tration were raised to the normal （P〉0.05）. Our study suggested that baicalein exerts a positive effect on the VIP expression in HCMV-infected EVT at maternal-fetal interface.
基金supported by a grant from the National Natural Science Foundation of China(No.30672243)Hubei Provincial Natural Science Foundation(No.2009CDB216)
文摘The effect of human cytomegalovirus （HCMV） on invasive capability of early pregnant extravillous cytotrophoblasts （EVTs） was investigated in vitro. Primary EVTs were obtained by complex phosphoesterasum digestion and gradient centrifugation from villous tissue aseptically taken from healthy pregnant women. Cytokeratin7 （CK7）, vimentin （Vim） and cerbB-2 were immunocytochemically detected to identify source of cells, and HCMVpp65 antigen was assayed to determine the infection state of primary EVTs by immunocytochemical staining. The EVTs were divided into two groups： control group and HCMV group, and the expression of c-erbB-2, matrix metalloproteinase-2 （MMP-2） and MMP-9 proteins was detected in two groups by immunocytochemistry and Western blotting. Enzymic activity changes of MMP-2 and MMP-9 were tested by gelatin zymography in primary EVTs infected with HCMV. The invasion of primary EVTs was detected by cell invasion assay in vitro after they were infected by HCMV. The cell source identification showed that the cells obtained were highly-pure primary EVTs, and primary EVTs could be infected by HCMV. Primary EVTs could express c-erbB-2, MMP-2 and MMP-9 proteins, and as compared with control group, the protein expression was decreased significantly in HCMV groups （P〈0.05）. Primary EVTs could secrete active MMP-2 and MMP-9 in vitro, and the activity of two MMPs was decreased sig- nificantly in HCMV groups （P〈0.05）. The in vitro cell invasion assay showed that the number of primary EVTs permeating Matrigel in HCMV group was decreased （P〈0.05）. We are led to conclude that HCMV can infect primary EVTs and inhibit their invasion capability, suggesting that the im- paired EVT＇s invasion capability might be related to the abnormal expression of c-erbB-2, MMP-2 and MMP-9 proteins.
文摘The effects of leptin on cytotrophoblast proliferation and invasion activity were investigated. Immunohistochemistry was used to determine the placental expression of leptin in first-trimester pregnancy. By using RT-PCR and quantitative real-time PCR, the expression of leptin in cytotrophoblast and the effect of leptin on cytotrophoblast secretion were detected. The potential of cell proliferation, invasiveness and migration was assessed by MTT, Transwell invasion assay and migration assay respectively when the cytotrophoblast was cultured with different concentrations of leptin. The results showed that： （1） Leptin was distributed diffusely around cell membrane, in cytoplasma, and on nuclear mem- brane of cytotrophoblast; （2） Leptin mRNA was expressed in cytotrophoblast. Ten ng/mL leptin could promote the secretion of cytotrophoblast significantly （P〈0.01）; （3） After culture with different concentrations of leptin for 24 h or longer, the proliferation of cytotrophoblast was inhibited, while in 24 h leptin could promote cytotrophoblast invasion and migration. Leptin at a concentration of 500 ng/mL could promote cytotrophoblast invasiveness and migration significantly as compared with controls （P〈0.05）. It was suggested that leptin could inhibit cytotrophoblast proliferation, and promote cytotro- phoblast invasion and migration activity.
