In vitro study on human cytomegalovirus affecting early pregnancy villous EVT’s invasion function. Discussion

Discussion

HCMV is the most common pathogen in intrauterine infection. Among the new born babies in Europe and America, HCMV infection rate is 2%, while that in developing countries is even higher. HCMV intrauterine infection may cause miscarriage, stillbirth, fetus developmental retardation, central nervous system damage and other serious results, but its specific pathogenesis remains unknown. HCMVpp65 protein is the major envelope protein in early stage of HCMV reproduction. The definite symbol of HCMV active infection is controlling HCMV gene expression, inhibiting host’s cell metabolism and promoting replication of virus.

In the later stage of blastula implantation, CT is divided into VT and EVT. VT locates in inner layer of villus, being active in karyokinesis and able to fuse and form ST which floats on the surface of villus, functioning as internal secretion, immunity and material exchange; EVT proliferates and migrates, gathering together to be cell column and invading into uterus decidua and superficial muscle layer, forming anchoring villus, which enables placenta to be fixed in vascular system of uterus. During embryo implantation, EVT invades decidua and uterus superficial muscle layer and physiologically remodels uterine spiral arteries, which is the necessary condition of successful pregnancy. EVT invasion function decreases, which may lead to miscarriage, stillbirth, fetus developmental retardation, gestational hypertension and other complications of pregnancy. In order to probe the molecular mechanism of HCMV intrauterine infection’s causing miscarriage, stillbirth, fetus developmental and other abnormal outcomes, early-pregnancy villus of healthy women in normal pregnancy is used for culturing in vitro, and HCMV is added into culture medium. After 48 h, it is observed that villus HCMV infection condition and the c-erbB-2, MMP-2 and MMP-9’s expression levels in villus and characteristics of space distribution change. Immunohistochemistry staining results show that HCMVpp65 antigen signal does not show in control group, but in almost all CT, EVT, ST, interstitial cells and around small blood vessels of villous interstitial substance, that is, after adding HCMV into villous culture medium, 48 h later, villous cells are infected.

c-erbB-2 is part of epidermal growth factor receptor, being a kind of oncogene protein with tyrosine kinase activity and closely linked with embryogenesis, tissue repair and regeneration. It is found that c-erbB-2 highly expresses in EVT which is cultured in vitro, being one of the symbolic proteins of EVT. c-erbB-2’s expression time and space is highly consistent with EVT invasion function in villus. Therefore, c-erbB-2 can be used to analyze EVT invasion function in villus. This experimental result indicates that c-erbB-2 mainly expresses in early-pregnancy ST remote plasma membrane, EVT, especially villous stem cell column remote EVT’s plasma membrane, being consistent with the reports of Mühlhauser et al; HCMV infection brings about the reduce of villous c-erbB-2 expression level, which probably is related with the decrease of EVT invasion function.

MMP is one of the key enzymes which affect EVT migration and invasion function, being characteristic of specificity and timeliness in various trophocytes. Maternal-fetal interface’s physical environment, changes in extracellular matrix components, growth factor, cell factor and hormone together maintain the balance of MMP/tissue inhibitor of metalloproteinase(TIMP) system and control the cytotrophoblast cells’ proliferation, differentiation and invasion. It is found in study that MMP-2 and MMP-9 are the main stroma metalloprotease which mediate EVT invasion. MMP-9 highly expresses in early and medium pregnancy EVT; MMP-2 expresses in all EVT during the whole gestational period and is active in early and medium pregnancy. The changes of expression level and activity are the same as EVT invasion function’s changes. In our study, villous immunohistochemistry staining shows that MMP-2 and MMP-9 protein express largely in early pregnancy villous EVT endochylema, while MMP-9 protein also expresses in high level in early pregnancy villous interstitial substance and VT endochylema; HCMV infection results that villous MMP-2 and MMP-9 expression level both significantly decrease. Villous MMP-2 and MMP-9 expression level reduces, which will lower EVT’s proliferation, differentiation and invasion function. The mechanism of HCMV damaging the balance of MMP/TIMP system still remains unclear.