Is intrauterine hematoma associated with adverse pregnancy and obstetric outcomes of ART singletons? A systematic review and meta-analysis.

The objective of our meta-analysis was to estimate the effect of intrauterine hematoma (IUH) on obstetric and pregnancy outcomes of assisted reproductive technology (ART) pregnancies. Four electronic databases were searched up to December 2021 to find studies reporting relevant outcomes of ART pregnancies with IUH. Dichotomous data were expressed as odds ratios (OR) with 95% confidence intervals (CI). Continuous data were expressed as weighted mean difference (WMD) with 95% CI. A total of six observational studies were included in this meta-analysis. Our data suggested that IUH in pregnancies achieved by ART are not associated with increased risks of miscarriage, low birth weight, placenta previa, or premature rupture of membranes. Similar birthweight was noted between the two groups. However, IUH was associated with significantly shorter gestational age at delivery (GA) as well as higher risks of preterm birth. Subgroup analyses have found that the presence of retroplacental haematoma was associated with an increased risk of miscarriage. IUH may be associated with decreased GA and an increased risk of preterm birth. Therefore, Women diagnosed with IUH should be offered increased surveillance during the course of their pregnancy.

Characteristics and establishment of cell lines from human gastrointestinal stromal tumors.

OBJECTIVE: To establish the cell line from specimens of resectable human gastrointestinal stromal tumors (GIST) and to verify the characteristics of cell biology in vitro. METHODS: The tissues from biopsies of human GIST were cultured in RPMI 1640 media supplemented with 10% fetal bovine serum. After growing to 90% confluence, the cells were detached for subculture and their characteristics, including morphology, growth kinetics, karyotype analysis, immunohistochemical analysis and tumorigenicity in nude mice were determined. RESULTS: GIST named GIST-H1 was successfully established. The cell line was passaged for more than 60 times 1 year. The characteristics demonstrated: The population doubling time calculated in the log phase of growth was 47.5 h. The cloning efficiency in the soft agar averaged 24.8%. Electronmicroscopically, there were rich ribosomes and mitochondrion in the cytoplasm. Immunohistochemical analysis showed CD117(+), SMA(+), dog-1(+), CD34(-), and S-100(-). Karyotype analysis illustrated aneuploidy with the modal chromosomal number 60-98. The GIST cells transplanted in nude mice had high tumorigenicity. CONCLUSION: The immortalized GIST cells are developed in vitro and have specific characteristics of GIST.

HSP70 and mucin 5B: novel protein targets of N,N'-dinitrosopiperazine-induced nasopharyngeal tumorigenesis.

N,N'-Dinitrosopiperazine (DNP) induces nasopharyngeal carcinoma (NPC) and shows organ specificity to the nasopharyngeal epithelium. To investigate its mechanism, the rat NPC model was induced using DNP. Rat NPC and normal nasopharyngeal cells were obtained from the NPC model using laser capture. The total proteins from these cell samples were separated with two-dimension polyacrylamide gel electrophoresis techniques, and highly expressed proteins (> five-fold) were analyzed using matrix-assisted laser desorption/ionization time of flight and bioinformatics. The results showed that HSP70 and mucin 5B expression increased not only in rat NPC but also in atypical hyperplasia nasopharyngeal tissues, a precancer stage of NPC. High-expression of heat shock protein 70 (HSP70) and mucin 5B was further supported by western blot analysis. The immunofluorescence and western-blotting studies further showed that DNP induced the expression of HSP70 and mucin 5B in a dosage-dependent manner in normal nasopharyngeal epithelia cells. Our data indicate that DNP triggers over-expression of HSP70 and mucin 5B, and is involved in nasopharyngeal tumorigenesis. HSP70 and mucin 5B may be important targets in nasopharyngeal tumorigenesis induced by DNP.

[Activation of anti-tumor cytotoxic T lymphocytes by fusion of human dendritic cells and melanoma cells].

OBJECTIVE: To determine the effect of activation of specific anti-tumor cytotoxic T lymphocytes (CTL) and the ability of cross-presentation in vitro by fusion of HLA-A2+ human dendritic cells (DCs) with HLA-A2- melanoma cells. METHODS: The HLA-A2+ human dendritic cells and HLA-A2- melanoma cells were fused by PEG and were cultivated in complete RPMI1640 media containing FCS (10%) and GM-CSF for 24-48 h, and then co-cultured fusion cells with Melan-A specific T cells. HLA-A2- melanoma cells were negative control,While T2 cells and DC+Pts were positive control. The activation of anti-tumor CTL elicited by the fusion cells was detected by intracellular cytokine staining. RESULTS: The immature DC could express CD80, CD83, CD86, HLA-DR, and HLA-ABC,but the mature DC induced by TNF-alpha, PGE-2, and CD40L further highly expressed above molecules. The rate of specific CTL cells primed by the fusion cells was 16.72%+/-4.26%, negative control was 0.21%+/-1.84%,and positive control was 28.60%+/-5.67%. The CTL from vaccine by fusing DC and LAR6 induced lysis of HLA-A2+ LAR1 cells. CONCLUSION: The HLA-A2 restricted specific anti-tumor CTL can be induced in vitro by fusion of HLA-A2+ human dendritic cells with HLA-A2- melanoma cells.

Induction of apoptosis by epigallocatechin-3-gallate via mitochondrial signal transduction pathway.

PURPOSE: To elucidate the apoptosis induction of Epigallocatechin-3-gallate (EGCG) on nasopharyngeal carcinoma (NPC) cells via mitochondrial signal transduction pathway regulated by EB-virus-encoded latent membrane protein 1 (LMP1). METHODS: The survival rates of pTet-on-LMP1 HNE2 cells after the EGCG treatment were determined by MTT assay. Induction of apoptosis in pTet-on-LMP1 HNE2 cells after the EGCG treatment was analyzed by agarose gel electrophoresis. The activity of caspase-9 was determined by ApoAlert Caspase-9 Fluorescent Assay kit after the EGCG treatment. The protein expressions of cytochrome c and Bcl-2 were analyzed by Western blotting after the EGCG treatment. RESULTS: EGCG inhibited the survival rates of pTet-on-LMP1 HNE2 cells and induced apoptosis of pTet-on-LMP1 HNE2 cells. EGCG raised the activities of caspase-9, enhanced the releasing of cytochrome c from the mitochondria, and suppressed the protein expression of Bcl-2. These are the key targets on the mitochondrial signal transduction pathway in apoptosis. CONCLUSIONS: EGCG inhibited the survival rate of NPC cells and induced apoptosis of NPC cells via the mitochondrial signal transduction pathway. This study suggests that the interference effect of EGCG on targets of the mitochondrial signal transduction pathway plays an important role in the anticancer function.

[Establishment and characterization of a melanoma cell line HME1].

OBJECTIVE: To establish a melanoma cell line and identify its characteristics in vitro. METHODS: The tissues from biopsies of melanoma were performed primary culture. After growing to 90% confluence, the cells were detached and transferred to another flask for subculture, and then we identified their characteristics including growth kinetic, morphology and tumorigenecity. RESULTS: This cell line was cultivated for more than 90 times. Its characteristics were as follows:The pathological morphology of the tumor transplanted in nude mice were similar to that of the original lesion;The growth curves of the 20th passage were determined, and the population doubling time calculated was 56.9 h; The cloning efficiency in soft agar was 19.1%; Karyo-type analysis showed aneuploidy with the modal chromosomal number 85-102; Electronmicroscopical observation showed that there were rich microvilli on the surface of the cells, abundant ribosomes and melanoid grain; SP immunohistochemical staining showed that the cells expressed HMB-45. CONCLUSION: The Melanoma cells were immortalized after being cultured in vitro and a new melanoma cell line was established.

Epstein-Barr virus encoded latent membrane protein 1 induces TRAF1 expression to promote anti-apoptosis activity via NF-kappaB signaling pathway in nasopharyngeal carcinoma.

OBJECTIVES: To identify whether Epstein-Barr virus (EBV) encoded latent membrane protein 1 (LMP1) can induce tumor necrosis factor receptor-associated factor 1 (TRAF1) expression and promote its anti-apoptosis activity via the NF-kappaB signaling pathway, and assess that LMP1 suppresses apoptosis in nasopharyngeal carcinoma (NPC). METHODS: A stable transfected cell line HNE2-LMP1 was established by introducing LMP1 cDNA into HNE2 cells. Transactivation of TRAF1 was determined by luciferase reporter assay, while expression of TRAF1 mRNA was detected by RT-PCR and expression of TRAF1 protein and caspase 3 by Western blot analysis. Apoptosis activity was observed through fluorescence staining. RESULTS: LMP1 induced TRAF1 expression in NPC cells and caused a decrease in apoptosis. This induction could be blocked by antisense LMP1. Moreover, LMP1-mediated induction of a TRAF1 promoter-driven reporter gene was significantly impaired when the kappaB site kappaB1 or kappaB5 was disrupted, whereas mutation of kappaB3 had only a minor effect on LMP1 dependent up-regulation of the reporter gene. CONCLUSION: LMP1 induces TRAF1 expression and promotes its anti-apoptosis activity via the NF-kappaB signaling pathway, which may be one of the mechanisms that LMP1 uses to suppress apoptosis in NPC cells.

Epstein-Barr virus induces human nasopharyngeal epithelial cells to escape from the replicative senescence.

OBJECTIVE: To observe the biological changes of primary human nasopharyngeal epithelial cells in the early stage of immortalization. METHODS: The morphological changes of nasopharyngeal epithelial cells were observed by phase contrast microscopy, and the activity profile of senescence-associated beta-galactosidase (SA-beta-Gal) was detected by SA-beta-Gal staining. The expression of p16(INK4a) protein was tested by immunochemical assay, and the life span in vitro of nasopharyngeal epithelial cells was calculated as population doublings. In addition, the expression of Epstein-Barr (EB) virus latent membrane protein 1 (LMP1) was also detected by immunofluorescence staining. RESULTS: Morphologically, cells treated with EB virus and 12-o-tetradecanoyl-phorbol-13-acetate (TPA) formed multi-layer foci, and their cellular life span in vitro was extended (about 155 days of culture). A low percentage of cells (about 4.8%) expressed SA-beta-Gal activity at late primary culture, and did not always express p16(INK4a) protein in the progression of culture. CONCLUSIONS: Nasopharyngeal epithelial cells treated with EB virus in cooperation with TPA can pass through the stage of senescence and enter the early stage of immortalization. Some changes of phenotype occur in these cells. Our results provide data for further studying the mechanism of immortalization and the establishment of a human nasopharyngeal epithelial cell line.

[Matrix metalloproteinase 9 expression is induced by Epstein-Barr virus LMP1 via NF-kappa B or AP-1 signaling pathway in nasopharyngeal carcinoma cells].

OBJECTIVE: To clarify if Epstein-Barr virus encoded LMP1 induces matrix metalloproteinase 9 expression via NF-kappa B or AP-1 signaling pathway, which gives evidence to the elucidation of the mechanism of LMP1- mediated carcinogenesis. METHODS: To determine whether LMP1 or its mutants contribute to MMP9 production via NF-kappa B or AP-1 transcription factor, MMP9-chloramphenicol acetyl transferase (CAT), NF-kappa B mut 9-CAT, AP-1 mut MMP9-CAT were transfected into human nasopharyngeal carcinoma cells stably expressing LMP1 (HNE2-LMP1) or its mutants, [HNE2-LMP1 (1-185), HNE2-LMP1 (1-231), HNE2-LMP1 delta 187-351] by electroporation technic. The difference of MMP9 reporter activity among those cell lines was detected by CAT assay and expression of MMP9 was determined in nasopharyngeal carcinoma cells stably expressing LMP1 or its mutants by zymographic analysis. In the meantime, efforts were made to demonstrate if LMP1 regulates NF-kappa B or AP-1 activation using reporter gene analysis. RESULTS: In contrast with vector-transfected cells, MMP9 CAT activity in HNE2-LMP1, HNE2-LMP1 (1-185), HNE2-LMP1(1-231), HNE2-LMP1 delta 187-351 increased 7.2, 1.3, 3.3, 4.0 times respectively. Zymographic analysis demonstrated that the 92 kDa MMP9 expression was induced in HNE2-LMP1, HNE2-LMP1(1-231) and HNE2-LMP1 delta 187-351 cells, whereas it was negative in HNE2-pSG5 and HNE2-LMP1 (1-185) cells. As compared to the HNE2 cells, NF-kappa B or AP-1 reporter activity in HNE2-LMP1 cells were increased 13.8, 8.4 fold respectively. Moreover, In contrast with MMP9 CAT-transfected cells, MMP9 CAT activity in NF-kappa B mut MMP9-CAT or AP-1 mut MMP9-CAT transfected HNE2-LMP1, HNE2-LMP1 (1-185), HNE2-LMP1(1-231) and HNE2-LMP1 delta 187-351 cells were significantly decreased by 18.1% or 16.3%, 35.0% or 33.3%, 29.1% or 26.1% from the original level. However, there was no difference in NF-kappa B mut MMP9-CAT or AP-1 mut MMP9-CAT transfected HNE2-pSG5, HNE2-LMP1 (1-185) cells. CONCLUSION: In nasophargyngeal carcinoma, Epstein-Barr virus-encoded LMP1 induces MMP9 transcription and enzymatic activity via an NF-kappa B or AP-1 signaling pathway, which may contribute to invasiveness and metastasis.

[EB virus encoded latent membrane protein 1 modulates the phosphorylation of epidermal growth factor receptor in nasopharyngeal carcinoma cell line].

OBJECTIVE: To elucidate the regulation of the phosphorylation of epidermal growth factor receptor (EGFR) by the EB virus encoded latent membrane protein 1 (LMP1) in nasopharyngeal carcinoma cell line. METHODS: The levels of EGFR expression and phosphorylation in pTet-on LMP1 HNE2 cell, a nasopharyngeal carcinoma (NPC) cell line, in the dynamic expression of LMP1 induced by different concentrations of doxycycline (Dox) were observed. The EGFR dominant negative mutant and LMP1 antisense expression plasmid were transiently transfected into pTet-on LMP1 HNE2 cells by lipofectamine, and the changes in EGFR phosphorylation were observed by immunocoprecitation and Western blot. The changes in EGFR phosphorylation were observed after EGF treatment. RESULTS: In pTet-on LMP1 HNE2 cells, Dox-induced LMP1 upregulated EGFR expression and phosphorylation in a dose-dependent manner. After EGFR dominant negative mutant was transfected into pTet-on LMP1 HNE2 cells, the increase of EGFR phosphorylation was inhibited completely. When LMP1 antisense expression plasmid was transfected into pTet-on LMP1 HNE2 cells, the levels of EGFR phosphorylation were also inhibited significantly. Meanwhile, after EGF had been added into pTet-on LMP1 HNE2 cells, increase of EGFR phosphorylation was induced, but it was completely blocked by EGFR dominant negative mutant and the introduction of LMP1 antisense. CONCLUSION: EB virus encoded LMP1 not only induces the dose-dependent expression of EGFR, but also the dose-dependent phosphorylation of EGFR. The phosporylation of EGFR may play a vital role in the development of nasopharyngeal carcinoma.

The Primary Study on Expression and Function of D-Type Cyclins in Nasopharyngeal Carcinoma Cell Lines.

Previous work of the authors established the protein expression spectra of D-type cyclins, and found that there were differential expression in nasopharyngeal carcinoma (NPC) biopsies. Using Western Blotting, aberrant overexpression of the cyclinD1 protein in NPC cell lines is reported. CyclinD2 and cyclinD3, the other members of D-type cyclins, were also detected in NPC cell lines. A comparison of the expression patterns in the cell lines, positive or negative in the latent membrane protein 1 (LMP1) encoded by Epstein-Barr virus, revealed that the expression of D-Type cyclin proteins might be increased by EBV-LMP1. The abundance of these proteins showed characteristic variation consistent with a cell- cycle-dependent oscillation and the peak levels expressed in G1 as analyzed by double-parameter flow cytometry. These data suggest that cyclinD1 is essential for cell cycle progression in G1 and may play a role in NPC carcinogenesis.

Doxycycline Dependent Expression of Epstein-Barr Virus Latent Membrane Protein 1 with Tet Regulating System in Nasopharygeal Carcinoma Cell Line.

Latent membrane protein 1 (LMP1) of Epstein-Barr virus (EBV) plays a vital role in cell transformation and the nasopharygeal carcinoma (NPC) is highly related to EBV infection. To elucidate whether LMP1 exerts oncogenic effect in NPC and its possible mechanism, a dual-stable LMP1 integrated NPC cell line with Tet-on regulating system were established. Results revealed that expression of LMP-1 could be regulated by Tet system when NPC cell line HNE(2) served as receipt cell, and doxycycline could increase this expression by about 10 fold. Therefore, it is possible that different level expression of LMP-1 results in its different biological functions and various efforts on nasopharyngeal carcinogenesis.