Efficacy of Different Treatment Regimens for Antiphospholipid Syndrome-related Recurrent Spontaneous Abortion / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Antiphospholipid syndrome (APS)-related immune factors are considered as an important cause of recurrent spontaneous abortion (RSA). Anticoagulant and anti-inflammatory treatments are believed to effectively improve adverse pregnancy outcomes by affecting the abnormal autoimmune response of the maternal-fetal interface. The aim of this study was to observe the clinical characteristics and treatment outcomes of anticoagulant regimens and anti-inflammatory plus anticoagulation regimens for APS-related RSA.</p><p><b>METHODS</b>APS-related RSA cases from September 2011 to September 2016 at Peking University Third Hospital were retrospectively analyzed. The patients were assigned to study group (anti-inflammation plus anticoagulation) and control group (simple anticoagulation). The incidence of repeat abortion, the incidence of placental dysfunction, the gestational weeks of pregnancy, and the mean weight of the fetus were observed.</p><p><b>RESULTS</b>The pregnancy and neonatal outcome indicators of the repeat pregnancy loss rate (11.11% vs. 22.70%), placental dysfunction-related diseases (6.35% vs. 15.60%), the mean birth weight of infants born after 24 weeks gestation (3152.41 ± 844.67 g vs. 2765.76 ± 816.40 g), full-term delivery weight (3456.28 ± 419.79 g vs. 3076.18 ± 518.79 g), the proportions of low birth weight infants (12.70% vs. 21.98%), and small for gestational age (6.35% vs. 14.18%) differed significantly between the study and control groups (all P< 0.05). The incidence of preterm delivery, term delivery, and stillbirth was not significantly different between the two groups, and there was no significant difference between the study and control groups in gestational age at birth (37.6 ± 3.3 weeks vs. 36.9 ± 3.2 weeks; P > 0.05).</p><p><b>CONCLUSION</b>The anti-inflammatory and anticoagulation regimen is more effective than the simple anticoagulation regimen in the treatment of APS recurrent abortion.</p>

Establishment of a high metastatic potential human hepatocellular carcinoma orthotopic transplantation model with palliative liver resection in nude mice / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To construct a high metastatic potential human hepatocellular carcinoma (HCC) orthotopic transplantation model with palliative liver resection in nude mice.</p><p><b>METHODS</b>A human HCC orthotopic nude mice model was established by administering a single inoculation of the highly metastatic MHCC97H tumor tissue (size 2 mm * 2 mm * 2 mm) into the left liver lobe. At day 14 post-inoculation, a random group of the mice received palliative liver resection; the unresected mice served as controls. Changes in expression levels of 113 genes with metastasis-related functions were evaluated in the residual HCC tissues. At day 35 post-resection, a random group of the mice were sacrificed by cervical dislocation and a comprehensive metastases examination was performed. The remaining mice were used to observe life span. All statistical analyses were performed by the SPSS v17.0 software, and significance was defined as P less than 0.05.</p><p><b>RESULTS</b>The nude mouse model of highly metastatic HCC with palliative liver resection was successfully established. Incidences of intrahepatic and abdominal metastases were higher in the palliative resected group (vs. unresected group: 11.7+/-4.7 vs. 6.3+/-2.8, t = -2.412, P less than 0.05 and 9.8+/-3.4 vs. 5.2+/-2.6, t = -2.641, P less than 0.05 respectively). In addition, the palliative resected group showed significantly enhanced pulmonary metastasis (vs. unresected group: 14.3+/-4.7 vs. 8.7+/-4.7, t = -2.348, P less than 0.05). Differential gene expression levels were found for MTSS1, TGFbl, SMAD2, IL-1b, and MMP7, and were situated in the central position of gene function net of residual HCC. The life-span of the palliative resected group was significantly longer than that of the unresected group (60.8+/-2.7 vs. 51.3+/-1.4 days, x2 = 12.850, P less than 0.01).</p><p><b>CONCLUSION</b>The highly metastatic human HCC nude mouse model with palliative liver resection that was successfully constructed in this study represents a useful investigational tool to assess the biological characteristics of residual cancer and to screen therapeutic strategies.</p>

Molecular pathological changes of human fetal scleral fibroblasts following exposure to extremely low frequency electromagnetic field / 中华实验眼科杂志

BackgroundThe effects of extremely low frequency electromagnetic fields (ELF-EMFs) on public health have attracted wide attentions.The association of the thermal effect of ELF-EMFs with cancer and ocular tissue damage has been of concern.However,the pathological changes of scleral tissue after exposure to ELF-EMFs as well as the relationship between these changes and myopia are still poorly understood.ObjectiveThe present study was to investigate the molecular pathological changes of human fetal scleral fibroblasts (HFSFs) after exposure to ELF-EMFs in vitro and to explore the possible mechanism in the occurrence and development of myopia.MethodsHFSFs were cultured and passaged and then exposed to 50 Hz electromagnetic fields,and HFSFs that did not receive the irradiation of ELF-EMFs were used as the control group.The expression of collagen type Ⅰ (COL1A1 ) mRNA and matrix metalloproteinase-2 (MMP-2) mRNA in cultured HFSFs were detected by real-time qualitative polymerase chain reaction (real-time PCR) under different magnetic field intensites (0,0.1,0.2,0.5,1.0 mT) and different exposure time (0,6,12,24,36,48 hours).Cell proliferation assay of HFSFs was detected by the cell counting kit 8 ( CCK8 ) assay.The expression levels of COL1 A1 and MMP-2 proteins in HFSFs were further confirmed by immunofluorescence staining.Results The expression of COL1A1 mRNA was significantly down-regulated under the exposure of 0.2 mT ELF-EMFs for 6 hours,in comparison with the control group;moreover,it decreased in parallel with the increased of flux density (0.099±0.008 vs.0.050±0.004) (P =0.009 ).The expression of MMP-2mRNA was up-regulated conspicuously after exposure to 0.1 mT ELF-EMFs for 24 hours,and it increased with exposure time in comparison with the control group ( 0.009 ±0.001 vs.0.018±0.003 ) ( P =0.038 ).Proliferation of HFSFs (A450) was inhibited following the exposure to 0.2 mT ELF-EMFs for 24 hours in comparison with the control group (P =0.009 ).The expression of COL1 A1 in the experimental group was decreased,compared with the control group,but the expression of MMP-2 was increased.ConclusionsELF-EMFs inhibit the proliferation of HFSFs and expression of COL1 A1 in HFSFs,which might be one of the reasons for the development of myopia.

Effects of MIM-B gene on invasive and metastatic potentials of human hepatocellular carcinoma MHCC97H cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effects of lentivirus mediated siRNA targeting human metastasis suppressor 1 (MTSS1, MIM-B gene) gene on the invasive and metastatic potentials of hepatocellular carcinoma (HCC) MHCC97H cells.</p><p><b>METHODS</b>The siRNA targeting MTSS1 was cloned into one lentivirus work vector. The work vector and three package plasmids were co-transfected into 293T cells with the help of lipefeetamine 2000. Lentivirus was collected in 72 hours and was added to the cultured MHCC97H cells. The total cell MIM-B mRNA and MIM-B protein were extracted and underwent real-time PCR and western-blot test respectively. Boden chamber assay was used to evaluate the invasive potential of MHCC97H cells. Gelatin zymography was used to detect matrix metalloproteinase-2 (MMP2) activity. Metastatic human HCC nude mice models were established by orthotopic implantation with a high metastatic potential human HCC cell line MHCC97H. Twenty-four nude mice bearing orthotopic xenografts were randomized into black control group, Lenti-GFP group and intervention group (Lenti-MTSS1 group) 14 days after orthotopic implantation (8 per group). The ultrasound-guided multi-point injection was performed on mice with borate buffered saline, Lenti-GFP and Lenti-MTSS1 respectively. Mice were sacrificed on day 35 for the examination of pulmonary metastasis. The SPSS 13.0 soft ware was applied to data analysis.</p><p><b>RESULTS</b>The small interfering RNA targeting MTSS1 was constructed successfully with a transfection efficiency of 97.0%, which produced a marked inhibition of invasive ability of MHCC97H cells through Matrigel, being 37.9+/-4.4, 37.4+/-5.3 and 26.6+/-4.6 in the black control group, Lenti-GFP group and Lenti-MTSS1 group (F = 26.695, P value is less than 0.01), respectively. MIM-B expression and MMP2 activity of intervention group were also significantly down-regulated as compared to the control group. The results of in vivo studies showed that the numbers of lung metastatic nodules were 6.5+/-2.6, 6.4+/-2.7 and 3.8+/-1.3 in the black control group, Lenti-GFP group and intervention group respectively with significant statistical difference (F = 3.637, P value is less than 0.05), accorded with tumor tissue MIM-B mRNA expression of 0.39+/-0.19, 0.38+/-0.10 and 0.16+/-0.11 respectively (F = 11.644, P value is less than 0.01) when comparison was made between control group and therapy group.</p><p><b>CONCLUSION</b>Small interfering RNA mediated by lentivirus inhibited MIM-B expression and resulted in inhibition of the invasive and metastatic potentials of MHCC97H cells, which may attributed, in part, the down regulation of MMP2 activity, and thus may provide a new molecular targeted therapy for HCC patients in the future.</p>

Effect of postoperative adjuvant transarterial chemoembolization on late recurrence of hepatocellular carcinoma after radical resection / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To identify the effect of postoperative adjuvant transarterial chemoembolization (TACE) on late recurrence of hepatocellular carcinoma (HCC) patients after radical resection.</p><p><b>METHODS</b>From year 2001 to 2007, 2436 HCC patients underwent radical resection were retrospectively selected. Patients underwent resection only were classified into control group, while those received adjuvant TACE within 2 months after operation were classified into intervention group. Patients were further stratified into those with tumor<or=5 cm and presenting low or high risk factors for recurrence, as well as tumor>5 cm and presenting low or high risk factors for recurrence. Patients with single tumor and without microscopic tumor thrombus were defined as low risk for recurrence; otherwise they were defined as high risk. The effect of adjuvant TACE on late (>2 years) recurrence was evaluated.</p><p><b>RESULTS</b>Recurrence rates of tumor<or=5 cm and presenting low, high risk factors for recurrence, as well as tumor>5 cm and presenting low, high risk factors for recurrence at 2-year after resection were 20.38%, 33.06%, 30.54% and 50.82%, respectively in the control group, compared with 25.41%, 39.61%, 40.55% and 51.57%, respectively in the intervention group; there were no significant differences between intervention group and control group in each stratum. For patients recurred or died within the first 2 years after resection, the median survival of tumor>5 cm and presenting high risk factors for recurrence was 24 months in the intervention group and 12 months in the Control group. Only in this subgroup, the survival curve of the intervention group was significantly higher than that compared to the control group. For patients who remained recurrence free and survived within the first 2 years after resection, there were no significant differences in the recurrence curves between the intervention group and control group in each stratum; while cumulative survival rates in the subgroup of patients with tumor size is less than or equal to 5 cm and presenting low risk factors for recurrence were 93.95%, 91.50% and 88.42% respectively in the control group, compared with 91.70%, 81.32% and 78.19% respectively in the intervention group at 3-, 4- and 5-year after resection (P=0.0062); for other subgroups, there were no significant differences in the survival curves between the intervention group and the control group in each stratum. Cox regression model suggested adjuvant TACE was not an independent risk factor for late recurrence; however, it might have negative effect on survival [hazard ratio (HR)=1.50, P=0.062] for those patients (especially patients with tumor is less than or equal to 5 cm and presenting low risk factors for recurrence).</p><p><b>CONCLUSIONS</b>The value of adjuvant TACE was mainly due to its therapeutic actions on residual tumor or early recurrence. It had no effect on postponing or eliminating late recurrence; moreover, it could be a risk rather than a benefit in patients at low risk for recurrence (especially those with tumor is less than or equal to 5 cm and presenting low risk factors for recurrence).</p>

Therapeutic efficacy and prognostic factors of sorafenib treatment in patients with unresectable primary hepatocellular carcinoma / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To evaluate the efficacy and analyze the prognostic factors of sorafenib treatment in patient with unresectable primary hepatocellular carcinoma (HCC).</p><p><b>METHODS</b>During the period from December 2005 to March 2009, 50 patients with unresectable primary HCC of Child-Pugh status A were treated with sorafenib (400 mg, Bid). The tumor response was evaluated with CT or MRI imaging every 6 - 8 weeks according to the RECIST criteria. The overall survival (OS) and time to progression (TTP) were defined as the time from administration of sorafenib to the death or the last follow up and were evaluated by Kaplan-Meier method.</p><p><b>RESULTS</b>There was no PR or CR, but 28 patients (56.0%) achieved stable disease. The median follow up time was 15 months with a median OS of 14 months and median TTP of 4 months. The common adverse events were dermal reaction (68.0%, 34/50), diarrhea (52.0%, 26/50), hypertension (4.0%, 2/50), hair loss (14.0%, 7/50), myelosuppression (16.0%, 8/50), and liver dysfunction (20.0%, 10/50). However, most of the drug-related adverse events were grade I-II and reversible. The patients with lower tumor burden and without distant metastasis had better prognosis.</p><p><b>CONCLUSION</b>Soafenib is effective for unresectable primary HCC with tolerable toxicity. Tumor stage is a predominant prognostic factor.</p>

Analysis of differential gene expression in rat hepatic stellate cells activated by culture or hepatocellular carcinoma cell induction / 中华肿瘤杂志

<p><b>OBJECTIVE</b>Hepatic stellate cells (HSC) in hepatocellular carcinoma (HCC) transdifferentiate into extracellular matrix-producing myofibroblasts. Activated HSC can promote invasion and metastasis of HCC. To understand the differences of HSC in normal liver and HCC, we compared the gene expression patterns in HCC cell induction-activated and culture-activated rat HSC.</p><p><b>METHODS</b>HSC were isolated by density centrifugation and exposed to conditioned medium from rat HCC cell line C5F. Expression of 22 012 genes in quiescent HSC, culture-activated HSC and HCC induction-activated HSC was analyzed by cDNA microarray and confirmed by real-time RT-PCR and Western blot.</p><p><b>RESULTS</b>1672 genes were differentially expressed in culture-activated HSC, including proinflammatory factors, cell adhesion molecules, cell surface receptors, signaling transduction molecules and immune factors. 711 genes were differentially expressed in HCC induction-activated HSC. Some of them were identical to those in culture-activated HSC. HCC Induction-activated HSC showed specific gene expression patterns, including Raf1, Rac2, Adam17, Wnt6, MMP-9 and TNF, suggesting that HCC cells can specifically induce HSC activation.</p><p><b>CONCLUSION</b>The gene expression patterns in HCC induction-activated HSC are different from those in culture-activated HSC. HCC induction-activated HSC may play a major role in the invasion and metastasis of HCC. In vivo activation should be considered as the standard for the study of HSC biology. HCC induction-activated HSC should be considered as the standard for HSC biology studies.</p>

Effect of siRNA targeted against survivin on the malignant behaviors of human hepatoma cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study survivin expression in human hepatoma cells and the effects of survivin siRNA on the malignant phenotypes of human hepatocellular cell line HCCLM6.</p><p><b>METHODS</b>Four hepatocellular carcinoma (HCC) cell lines were used. Semi-quantitative RT-PCR and Western blot were used to measure and compare their survivin expressions. The siRNA expression vector pshRNA-survivin targeting the mRNA of survivin and vector pGPU6/GFP/Neo-NC (as a control) were constructed, and then transfected into HCCLM6 cells. FQ-PCR was used to quantify the mRNA levels of survivin. The malignant phenotypes of transfected HCCLM6 cells, including invasive activities and adhesive capabilities, were analyzed.</p><p><b>RESULTS</b>Survivin expression gradually increased with the increase of the invasion and metastasis behaviors of the four HCC cell lines (P<0.05). The expression of survivin was highest in cell line HCCLM6. Survivin mRNA level was decreased by 93.500%+/-3.117% after the pshRNA-survivin transfection. The cell adhesion rates significantly decreased in the cells transfected with pshRNA-survivin (cell adhesion rates were 11.403%+/-1.256% vs 32.545%+/-1.367%, t=20.732, P<0.01). The migrating number of HCCLM6 cells (13.5+/-0.9) transfected with pshRNA-survivin was also significantly decreased (t=14.5, P<0.01) as compared with the control group (32.6+/-1.4).</p><p><b>CONCLUSION</b>The expression of survivin in HCC might have a close relationship to their invasion and metastasis properties. Sequence-specific shRNA can significantly reduce the survivin expression in the HCCLM6 cell line. Suppression of survivin expression in HCCLM6 cells transfected with pshRNA-survivin can reduce their invasive and adhesive capabilities.</p>

Functional localization of metastasis suppressor genes for HCC on human chromosome 8 / 中华肝脏病杂志

<p><b>OBJECTIVE</b>We previously showed that introduction of a normal, neomycin-tagged human chromosome 8 reduced the metastatic capacity of C5F rat liver cancer cell line, which had high metastatic potential without affecting tumorigenicity, suggesting the presence of one or more metastasis suppressor genes encoded on human chromosome 8. We proceeded to define further the region harboring the metastasis suppressor gene(s) and to determine the random loss of human chromosome 8 by PCR amplification of sequence tag site (STS) markers.</p><p><b>METHODS</b>The national Center for Biotechnology Information (NCBI) databases were used as references of the relative genetic distances of the STS markers. C5F genomic DNA and A9/neo8 genomic DNA were used as negative and positive controls for chromosome 8 amplification, respectively. Genomic DNA was isolated and quantified from cultured hybrid clones (A9/C5F-1 and A9/C5F-2 microcell hybrid clones served as metastasis-unsuppressed groups; A9/C5F-4, A9/C5F-8 and A9/C5F-10 microcell hybrid clones served as metastasis suppressed groups). STS-PCR products were separated by electrophoresis through 2% agarose gel.</p><p><b>RESULTS</b>Metastasis-suppressed microcell hybrid clones (A9/C5F-4, A9/C5F-8 and A9/C5F-10) conserved STS markers between D8S542 --> D8S1973 (8p21.1-23.1). In contrast, metastasis-unsuppressed clones (A9/C5F-1 and A9/C5F-2) lacked several markers in this region. In attempts to refine the region retained in the microcell suppressed clones, more densely spaced STS markers in the human chromosome 8p21.1-23.1 were used. We found that the metastasis-suppressed clones retained 18cM region between D8S542 and D8S1973 (8P21.1-23.1), where as the metastasis-unsuppressed clones lacked the region.</p><p><b>CONCLUSION</b>Our results suggest that a metastasis suppressor gene is located within the interval between D8S542 and D8S1973 on human chromosome 8p21.1-23.1.</p>

Treatment of postoperative recurrence of hepatocellular carcinoma with radiofrequency ablation comparing with repeated surgical resection / 中华外科杂志

<p><b>OBJECTIVE</b>To evaluate the efficacy of radiofrequency ablation for the treatment of postoperative recurrence of hepatocellular carcinoma and whether radiofrequency ablation can be used as first line treatment for recurrent hepatocellular carcinoma.</p><p><b>METHODS</b>There were 213 patients with small recurrent hepatocellular carcinoma (tumor size of 3 cm or less and no more than 3 nodules) who treated in Liver Cancer Institute, Fudan University from January 2000 to December 2005. Among these patients 68 were treated with radiofrequency ablation and 145 were treated with repeated surgical resection. Kaplan-Meier method was used to evaluate the overall survival or disease free survival. Log-rank used to determine the survival difference between groups and COX proportional hazard was used for multivariate analysis to evaluate the risk factors for prognosis. The overall survival or disease free survival was calculated from the time treated with radiofrequency or repeated surgical resection.</p><p><b>RESULTS</b>The 1-, 3-, 5-years overall survival rates were 94.7%, 65.1%, 37.3% and 88.1%, 62.6%, 41.0% in radiofrequency ablation group and surgical repeated resection group, respectively. There was no significant difference between two groups (P = 0.693). However, the disease free survival was better in repeated surgical resection than in radiofrequency ablation, which were 79.4%, 48.1%, 34.4% and 58.0%, 27.8%, 12.4% in repeated surgical resection and radiofrequency ablation, respectively (P = 0.001). The interval between recurrence and initial hepatectomy with more than 2 years was independent factor favor to good prognosis.</p><p><b>CONCLUSIONS</b>Radiofrequency ablation seems to be as effective as repeated surgical resection owing to comparable overall survival and can be considered as alternative therapy for surgical resection treatment of small recurrent hepatocellular carcinoma.</p>

Relationship between epithelial-mesenchymal transition and lung metastasis in hepatocellular carcinoma / 中华外科杂志

<p><b>OBJECTIVE</b>To evaluate the relationship between epithelial mesenchymal transition (EMT) and lung metastasis in hepatocellular carcinoma.</p><p><b>METHODS</b>There were 100 patients who underwent surgical resection of hepatocellular carcinoma between January 2000 and March 2004. They were classified with non-distance metastasis and lung metastasis depend on the close following up till March 2007. Their hepatocellular carcinoma specimens were retrospectively examined for EMT markers (E-cadherin, Vimentin, Fibronectin) with immunochemistry staining in tissue microarray. Univariate and multivariate analysis were used for study the relationship between EMT and lung metastasis.</p><p><b>RESULTS</b>Univariate analysis showed that down regulation of E-cadherin, overexpression of fibronectin, cytosolic expression of vimentin, AFP >or= 400 ng/ml, tumor size more than 10 cm, portal vein involvement, poorly differentiated of tumor had close correlation with lung metastasis. Multivariate analysis indicated that overexpression of fibronectin was independent factor for lung metastasis apart from tumor size more than 10 cm, portal vein involvement and poorly differentiated of tumor.</p><p><b>CONCLUSION</b>The results proposed that EMT has close relation with lung metastasis in hepatocellular carcinoma.</p>

A preliminary study of the killing function in vitro by T lymphocytes activated by dendritic cells loaded with exosomes secreted by hepatic cancer cell lines with high or low metastatic potentials / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the tumor cell killing function of T lymphocytes stimulated by dendritic cells (DC) and to analyze the differences of protein contents of exosomes in each type of cell.</p><p><b>METHODS</b>The exosomes of hepatic cell lines with high (P group) or low (F group) metastatic potentials were isolated by a process of four-step centrifugation and the collected exosomes were observed under an electron microscope (EM). The tumor cell killing experiment was performed by adding T lymphocytes activated by DC loaded with exosomes from corresponding P and F group cells and was studied using 3H-TdR experiments. The proteomic analysis was performed by surface-enhanced laser desorption/ ionization time of flight mass spectrometry (SELDI-TOF-MS ) on the exosomes of P and F group cells.</p><p><b>RESULTS</b>The density distribution and content of exosomes in the P group were not equal to those in the F group observed by EM. The CD80, CD86, MHC-I and MHC-II in the P group were 64.27+5.00, 44.89+10.11, 84.35+19.89 and 59.03+19.37, and those in the F group were 71.53+4.85, 50.01+9.50, 80.68+29.87 and 58.86+21.11, respectively (P>0.05, compared with the control group). The counts per minute value in the P group was 528.40+179.06 and 78.80+24.44 in the F group after being loaded with exosomes (P<0.01, compared with the control group). There were significant differences between the proteins in the exosomes of hepatic cancer cell lines with high or low metastatic potentials.</p><p><b>CONCLUSION</b>Exosomes have potential values of application in immunotherapy and in biotherapy for recurrences and metastases of hepatic carcinomas.</p>

Osteopontin promotes the malignant phenotypes of human hepatocellular carcinoma cell line SMMC-7721 / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the effects of osteopontin (OPN) on the phenotypes of human hepatocellular carcinoma cell line SMMC-7721.</p><p><b>METHODS</b>Human hepatocellular carcinoma SMMC-7721 cells were transfected with plasmid pcDNA 3.1(-)/OPN and cells transfected with a mock plasmid served as controls. OPN expression was verified by RT-PCR and Western blot, and concentrations of OPN, MMP-2, MMP-9 and uPA were measured by ELISA. A series of functional assays in vitro were used to monitor the changes of SMMC-7721 malignant phenotypes.</p><p><b>RESULTS</b>OPN expression of SMMC-7721 cells was elevated after transfection. Concentrations of OPN, MMP-2 and uPA in the medium of SMMC-7721 cells after transfection were higher than those of the controls [(3.02+/-0.12) ng/ml vs (1.43+/-0.07) ng/ml, (43.04+/-3.06) ng/ml vs (22.15+/-4.34) ng/ml, and (4.78+/-0.70) ng/ml vs (1.61+/-0.34) ng/ml respectively, P less than 0.01], but MMP-9 concentration did not increase [(7.82+/-2.25) ng/ml vs (7.70+/-1.92) ng/ml]. Functional assays in vitro indicated that SMMC-7721 cells after transfection showed higher adhesive, migrant and invasive capabilities than those of the controls (cell adhesion rates were 75.33%+/-10.59% vs 57.34%+/-2.52%; number of outer surface cells in migrant assay was 14.33+/-2.51 vs 6.34+/-1.53; cell number in the invasive assay was 8.23+/-1.53 vs 4.12+/-1.29 respectively, P less than 0.05).</p><p><b>CONCLUSION</b>OPN might enhance the expression of MMP-2 and uPA to promote malignant phenotypes of SMMC-7721 cells.</p>

Different expressions of chemokine receptors in human hepatocellular carcinoma cell lines with different metastatic potentials / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To compare different expression profiles of all known chemokine receptors in human hepatocellular carcinoma (HCC) cell lines with different metastasis potentials.</p><p><b>METHODS</b>Eighteen pairs of chemokine receptor primers were designed using Premier software. Expression profiles of the 18 chemokine receptors on four HCC cell lines of lower to higher potentials of metastasis (SMMC-7721, MHCC97-L, MHCC97-H and HCCLM6) were analyzed by RT-PCR. Expression of CXCR4 was detected by RT-PCR.</p><p><b>RESULTS</b>Expression profiles of chemokine receptors on four HCC cell lines with different metastatic potentials had significant differences (P < 0.01), in which CCR10, CXCR4 and CXCR6 expressions decreased gradually as the metastatic potential of the cell lines increased. The expressions of CCR3, CCR4, CCR10, CCR12 and XCR1 on HCCLM6 were significantly reduced compared with SMMC-7721 (P < 0.01), whereas the expressions of CXCR1 (P = 0.006) and CXCR5 (P = 0.003) exceeded that of SMMC-7721. Except for CXCR2, CXCR6 and XCR1, most of chemokine receptors on MHCC97-H were expressed differently compared with MHCC97-L (P < 0.05), in which expressions of CCR1 (P = 0.002), CCR2 (P = 0.004) and CCR5 (P = 0.046) exceeded MHCC97-L. CXCR4 was detected only on the positive controls and SMMC-7721 when the template of total RNA was reduced one-half in RT-PCR.</p><p><b>CONCLUSION</b>Chemokine receptors are expressed very differently at mRNA level on HCC cell lines with different metastatic potentials. The different profiles of chemokine receptors in tumor microenvironment and the function of CXCR4 in HCC should be further studied. Our findings have important implications in understanding the relationship between chemokine receptors and the metastatic potential of HCC.</p>

The investigation of the technology of microcell mediated chromosome transfer for functional localization of metastasis suppressor genes for liver cancer on human chromosomes / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>In order to seek the functional evidence that there could be metastatsis suppressor gene for liver cancer on human chromosomes, the objective of this study is to establish a method of microcell mediated chromosome transfer (MMCT).</p><p><b>METHODS</b>Human chromosome 8 randomly marked with neo gene was introduced into highly metastatic rat liver cancer C5F cell line by treating the single human chromosome donor cells with sequential steps of micronucleation, enucleation and microcell fusion. Double selections of G418 and HAT were applied to screen positive microcell hybrids, which were cloned by single cell isolation. Microcell hybrid clones were confirmed by STS-PCR and WCP-FISH.</p><p><b>RESULTS</b>Microcell hybrids resistant to HAT and G418 were obtained, from which 15 clones were obtained by single-cell isolation cloning. STS-PCR and WCP-FISH proved that human chromosome 8 had been successfully introduced into rat liver cancer cell line C5F. The human chromosome 8 introduced into C5F was found to have random loss of chromosome fragments by STS-PCR and consistent recombination with rat chromosome by WCP-FISH.</p><p><b>CONCLUSION</b>The successfulls introduction of human chromosome into highly metastatic rat liver cancer cell line has established the technical basis for functional localization of metastasis suppressor gene(s) for liver cancer on human chromosomes.</p>

Transcription factor activity profile of human hepatocellular carcinoma cell lines with different metastatic potentials / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To examine the activities of transcription factors (TFs) in human hepatocellular carcinoma (HCC) cell lines with different metastatic potentials, so as to identify the TFs associated with HCC metastasis.</p><p><b>METHODS</b>Transcription factor activity profile of Hep3B, MHCC97L and MHCC97H, three HCC cell lines with different metastatic potentials, were examined using protein/DNA array. Electrophoretic mobility shift assays (EMSA) and Western blot were used to confirm the results obtained by protein/DNA array.</p><p><b>RESULTS</b>From a total of 345 screened TFs, 7 activity differential TFs were found, of which 5 showed increased activity, including p53, hypoxia inducible factor-1 alpha (HIF-1alpha), signal transducer and activator of transcription 3 (Stat3) and Sp1, and 2 showed decreased activity including Rb and Smad3.</p><p><b>CONCLUSION</b>The abnormal functioning of transcription factors is closely associated with HCC metastasis. Our present findings could be of help in expanding our understanding of the mechanism of HCC metastasis and identify new predictive biomarkers and therapeutic targets.</p>

The relationship between lymphangiogenesis and lymphatic metastasis in murine hepatic carcinoma of high and low metastatic potentialities / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the relationship between lymphangiogenesis and lymphatic metastasis in mice bearing hepatic carcinoma and analyze the mechanism of the lymphatic metastasis.</p><p><b>METHODS</b>Hepatic carcinoma cell lines of high and low potentialities of lymphatic metastasis were injected into the footpads of Balb/c mice. Their metastases to lymph nodes were examined. The tumor tissues of each group were stained with 5'-nucleotidase-ALP to observe the lymphoangiogenesis. The total RNA of high and low metastatic potential cell lines were extracted for metastasis gene DNA array. The vascular endothelial cell growth factor C (VEGF-C) and VEGF-D of each cell line were detected using semi-quantitative RT-PCR and were further quantatively analyzed using real time PCR.</p><p><b>RESULTS</b>The para-common iliac a. and renal hilar lymph nodes metastases of the high metastatic potential cells were significantly higher than in the controls (P>0.05). The quantity of lymphatic vessels in the high metastasis group was significantly larger than that of the control group (P<0.05). The expressions of CD44, E-cadherin, HER2/neu, H-Ras and VEGF-C in the high metastasis group were higher than those in the low metastasis group shown by the cDNA micro array experiment but the expressions of nm23A, nm23-E4, p16ink4a, CD61 were lower. The VEGF-C expression was higher and the VEGF-D was lower in the high metastasis group compared to those of the low metastasis group shown by semi-quantitative RT-PCR. The secretion of VEGF-D was significantly lower and the ratio of VEGF-C/VEGF-D was significantly higher in the high metastasis group than the low metastasis group (P<0.05).</p><p><b>CONCLUSIONS</b>The lymphatic metastasis of hepatic carcinoma is related to lymphoangiogenesis. The changes of VEGF-C and VEGF-D expressions might be a cause influencing the lymphoangiogenesis. VEGF-C/VEGF-D might be an effective parameter in affecting lymphatic metastases.</p>

Changes in the immune function of dendritic cells (DC) derived from HBV-related hepatocellular carcinoma (HCC) patient's peripheral blood monocytes (PBMC) pulsed with tumor antigen / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To identify the phenotype and immune function of dendritic cells derived from HBV-related HCC patients's peripheral blood monocytes pulsed with soluble tumor antigen, and their relation to immune escape.</p><p><b>METHODS</b>Peripheral blood monocytes were isolated from 18 HBV-related hepatocellular carcinoma (HCC) patients, 11 HBV-related liver cirrhosis patients (LC) and 10 health blood donors; DCs were induced in the completed medium containing GM-CSF and IL-4. The morphology of DCs was studied using a confocal microscope and scanning electronic microscope, and the phenotype of DCs were detected by flow cytometric analysis. The mixed leucocyte reaction test was employed to determine the stimulatory capacity of DCs before and after being pulsed with soluble tumor antigen (prepared from HCCLM6 cell line). IL-12 ELISA kit was used to investigate IL-12 secretion of DCs in the supernate of MLR.</p><p><b>RESULTS</b>The amount of PBMC and DCs was significantly lower in LC and HCC compare to those in the healthy subjects; the expression levels of HLA-DR, CD1a, CD80 and CD86 on DC surfaces were lower in LC and HCC patients than those of the healthy group; the stimulating capacity of DC in MLR and levels of IL-12 in supernate of MLR were also lower in LC and HCC, but were enhanced after tumor antigen pulsed in all three groups, particularly in the LC group; the secretion of IL-12 in MLR supernate was still lower than that of the healthy group.</p><p><b>CONCLUSION</b>The phenotype and function defects of DC derived from PBMC of LC and HCC patients might play a key role in immune escape in HBV infection and HCC. The function of DC of LC patients can be enhanced after the tumor was antigen-pulsed.</p>

Relationship between DLC-1 expressions and metastasis in hepatocellular carcinoma / 中华肝脏病杂志

<p><b>OBJECTIVES</b>To study the relationship between the expression level of DLC-1 mRNA (located in 8p) and the invasion/metastasis of human hepatocellular carcinoma (HCC).</p><p><b>METHODS</b>Fifty-one surgical specimens of human HCC were divided into high-invasive and low invasive groups according to their clinicopathological features. DLC-1 mRNA expression was studied in the 51 HCC specimens as well as 5 different metastasis potential cell lines using real-time quantitative PCR (RQ-PCR).</p><p><b>RESULTS</b>The expression level of DLC-1 mRNA in HCC specimens with high invasiveness was significantly lower than that with low invasiveness (P < 0.05). The expression levels of DLC-1 mRNA were significantly different between non-metastatic (Hep3B and HepG2) and metastatic (MHCC97-H, MHCC97-L and HCCLM3) cell lines (P < 0.05). From MHCC97-L to HCCLM3, with an increase of invasiveness and metastatic potentials, the expression level of DLC-1 decreased correspondingly, and its expression level in HCCLM3 was significantly lower than that in MHCC97-L (P < 0.01).</p><p><b>CONCLUSION</b>The expression of DLC-1 mRNA may play an important role in inhibiting the invasiveness and metastasis of HCC.</p>

Influence of reconstruction of immunological functions of T lymphocytes on mouse hepatocarcinoma metastasis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effectiveness of reconstruction of immunological functions of T cells on the degree of metastases of mouse hepatocarcinoma and the mechanisms of their functioning.</p><p><b>METHODS</b>The T cell model of immunological functions in Balb/c nu/nu mice was established and the effectiveness of the model was evaluated. The mice were divided into 4 groups. The immunological functions of T cells in experiment groups of Balb/c nu/nu mice were reconstructed. Metastases of the cancer in lymph nodes in each group were examined histologically. The formation time and growth rate of the tumors were calculated. The expression of MHCI and II of the tumor cell line and the difference of expression of immune associated gene were detected by Th1-Th2-Th3 gene array.</p><p><b>RESULTS</b>The ratio of CD3, CD4, CD8 and CD4/CD8 in the reconstructed group was higher than that in the control group. The average formation time was 7.7+/-0.6 days in Balb/c nu/nu mice and 11.5+/-1.3 days in Balb/c mice. The extent of metastases of the experiment group was lower than that of the control group (P < 0.05). The expression of MHCI of the high metastasis cell line was lower than that of the low metastasis cell line (P < 0.05). The expressions of Th1/Th2 associated genes in lymphocytes of high metastasis mice were lower than those of the low metastasis mice.</p><p><b>CONCLUSION</b>Reconstruction of the immunological function of T cells can influence the metastasis of mouse hepatocarcinoma. The alteration of MHC molecule and low expression of Th1/Th2 correlated genes in lymphocytes may be a factor influencing the metastasis of liver cancer.</p>