Reproductive traits associated with invasiveness in Conyza sumatrensis

Conyza sumatrensis （Retz.） E. Walker, a member of Asteraceae, is a highly invasive species. However, its reproduction biology remains poorly known. To understand the role of reproductive traits in successful invasion of the species, we studied several traits of its reproductive system： the miniature capitulum and gynomonoecious sexual system, the biology and phenology of capitula and florets, pollen/ovule ratio, the mating system （selfcompatibility）, flower visitors, physical traits and dispersal potential of achenes, germination potential of achenes from manually pollinated capitula, and the association of these traits with invasiveness. Our study showed that the reproductive traits of autonomous seed production, versatile mating system of self- and cross-pollination, and generalized pollination system might contribute to the species＇ successful invasive capability. The invasiveness was further enhanced by the high and rapid production of achenes, as well as the high percentage, rapid germination rate and high dispersal capability of achenes. It was concluded that in annual or winter-annual weeds, autonomous seed production contributed significantly to the invasiveness of the species.

RhoA and RhoC -siRNA inhibit the proliferation and invasiveness activity of human gastric carcinoma by Rho/PI3K/Akt pathway

AIM: To evaluate the effects of adenovirus-mediated gene transfer of RhoA siRNA and RhoC siRNA on proliferation and invasion of SGC7901 cells by Rho/ PI3K/Akt pathway. METHODS: Plasmid of RhoA siRNA and RhoC siRNA were constructed and transfected into SGC7901 cells. siRNA and LY294002 (PI3K inhibitor) were designed as the control group. The mRNA and protein expressions of RhoA and RhoC were respectively detected with RT-PCR and western blotting. In order to fi nd out the changes of proliferation and invasion power of SGC7901 cell lines, we analyzed the data by MTT, Boyden chamber and evaluated apoptosis of cell with flow cytometry. We treated BALB /C nude mice with RhoA and RhoC-siRNA, and tumor control rate (%) in nude mice was calculated. RESULTS: RhoA and RhoC siRNA transfections specifically down-regulated the corresponding mRNA and protein levels in SGC7901 Cells.The experiment of permeated artificial basal membrane showed that the invasion power of SGC7901 cell lines are on the decline after treatment of Ad-RhoA and RhoC-siRNA (12.64 ± 3.27 vs 87.38 ± 17.38, P < 0.05). The values of 490 nm wavelength light absorption were different in the five groups. The number of alive cells in the group of RhoA and RhoC-siRNA was lower than others in the 6th d (0.71 ± 0.01 vs 3.82 ± 0,11 P < 0.05). The apoptosis rate of transfected RhoA and RhoC-siRNA group with FACS were 19.07% ± 1.78 and there were significant differences between treated and control groups (19.07 ± 1.78% vs 1.23 ± 0.11%, P < 0.01). The tumor transplantation experiment in BALB/C nude mice showed intratumoralinjection of RhoA or RhoC siRNA can inhibit tumor growth. CONCLUSION: RhoA and RhoC siRNA gene therapy mediated by adenovirus may be useful for inhibiting growth and invasion of SGC7901 through a PI3K/Akt pathway. These results provide a novel therapeutic target in preventing gastric cancer cell invasion and metastasis.

Suppression of matrix metalloproteinase-2 via RNA interference inhibits pancreatic carcinoma cell invasiveness and adhesion

AIM:To investigate the inhibitory effects of RNA interference (RNAi) on expression of matrix metalloproteinase-2 (MMP-2) gene and invasiveness and adhesion of human pancreatic cancer cell line,BxPC-3.METHODS:RNAi was performed using the vector (pGPU6)-based small interference RNA (siRNA) plasmid gene silence system to specifically knock down MMP-2 expression in pancreatic cancer cell line,BxPC-3. Four groups of different specific target sequence in coding region of MMP-2 and one non-specific sequence were chosen to construct four experimental siRNA plasmids of pGPU6-1,pGPU6-2,pGPU6-3 and pGPU6-4,and one negative control siRNA plasmid of pGPU6 (-). MMP-2 expression was measured by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Cell proliferation and apoptosis were examined by methyl thiazolyl tetrazolium (MTT) and flow cytometry,respectively. The abilities of adhesion and invasion were detected by cell adhesion assay and cell invasion assay using Transwell chambers.RESULTS:The expression of MMP-2 was inhibited and the inhibitory effects of different sequence varied. pGPU6-1 group had the most efficient inhibitory effect,followed by pGPU6-2 and pGPU6-3 groups.Invasiveness and adhesion were more significantly reduced in pGPU6-1,pGPU6-2 and pGPU6-3 groups as compared with pGPU6 (-) and blank control groups. However,no difference concerning cell proliferation and apoptosis was observed after transfection between experiment groups and control groups.CONCLUSION:RNAi against MMP-2 successfully inhibited the mRNA and protein expression of MMP-2 in the pancreatic cancer cell line,BxPC-3,leading to a potent suppression of tumor cell adhesion and invasion without affecting cell proliferation and apoptosis. These findings suggest that the RNAi approach towards MMP-2 may be an effective therapeutic strategy for the clinical management of pancreatic tumor.

Fascin promotes the motility and invasiveness of pancreatic cancer cells

AIM:To explore the role of actin-bundling protein, fascin during the progression of pancreatic cancer. METHODS:The plasmid expressing human fascin-1 was stably transfected into the pancreatic cancer cell line MIA PaCa-2. The proliferation, cell cycle, motility, scattering, invasiveness and organization of the actin filament system in fascin-transfected MIA PaCa-2 cells and control non-transfected cells were determined. RESULTS:Heterogeneous overexpression of fascin markedly enhanced the motility, scattering, and inva-siveness of MIA PaCa-2 cells. However, overexpression of fascin had minimal effect on MIA PaCa-2 cell pro-liferation and cell cycle. In addition, cell morphology and organization of the actin filament system were distinctly altered in fascin overexpressed cells. When transplanted into BALB/c-nu mice, fascin-transfected pancreatic cancer cells developed solid tumors at a slightly slower rate, but these tumors displayed more aggressive behavior in comparison with control tumors. CONCLUSION: Fascin promotes pancreatic cancer cell migration, invasion and scattering, thus contributes to the aggressive behavior of pancreatic cancer cells.

Effects of LY294002 on the invasiveness of human gastric cancer in vivo in nude mice

AIM: To investigate the effects of class phosphatidy-linositol 3-kinase (PI3K) inhibitor LY294002 on the invasiveness and related mechanisms of implanted tumors of SGC7901 human gastric carcinoma cells in nude mice. METHODS: Nude mice were randomly divided into model control groups and LY294002 treatment groups. On days 5, 10 and 15 after treatment, the inhibitory rate of tumor growth, pathological changes in tumor specimens, expression levels of matrix metalloproteinase (MMP)-2, MMP-9, CD34 [representing microvessel density (MVD)] and vascular endothelial growth factor (VEGF), as well as apoptosis indexes in tumor samples were observed.RESULTS: In this study, we showed that treatingthe tumors with LY294002 could significantly inhibit carcinoma growth by 11.3%, 29.4% and 36.7%, after 5, 10 and 15 d, respectively, compared to the control group. Hematoxylin & eosin staining indicated that the rate of inhibition increased progressively (23.51% ± 3.11%, 43.20% ± 3.27% and 63.28% ± 2.10% at 5, 10 and 15 d, respectively) along with apoptosis. The expression of MMP-2 was also downregulated (from 71.4% ± 1.6% to 47.9% ± 0.7%, 31.9% ± 0.9% and 7.9% ± 0.7%). The same effects were observed in MMP-9 protein expression (from 49.4% ± 1.5% to 36.9% ± 0.4%, 23.5% ± 0.9% and 7.7% ± 0.6%), the mean MVD (from 51.2% ± 3.1% to 41.9% ± 1.5%, 30.9% ± 1.7% and 14.9% ± 0.8%), and the expression of VEGF (from 47.2% ± 3.1% to 25.9% ± 0.5%, 18.6% ± 1.2% and 5.1% ± 0.9%) by immunohistochemical staining.CONCLUSION: The classPI3K inhibitor LY294002could inhibit the invasiveness of gastric cancer cells by downregulating the expression of MMP-2, MMP-9, and VEGF, and reducing MVD.

Association of Expression of Leucine-rich Repeats and Immunoglobulin-like Domains 2 Gene with Invasiveness of Pituitary Adenoma

The Leucine-rich repeats and immunoglobulin-like domains-2（LRIG2） gene expression in pituitary adenoma and its correlation with tumor invasiveness were studied.The expression of LRIG2 mRNA and protein in human pituitary adenoma obtained surgically was detected by RT-PCR（39 cases） and immunohistochemical staining（30 cases）.It was found that LRIG2 was mostly localized at the nucleus of the pituitary adenoma cells.Its expression was significantly higher in the invasive cases than in the non-invasive cases.LRIG2 protein was positive in 14 cases out of 21 cases of invasive adenoma,but only 2 cases were positive in 9 cases of non-invasive adenoma.The positive expression rate of LRIG2 mRNA was 91.3% in invasive cases（total 23 cases） and 62.5% in non-invasive cases（total 16 cases）,respectively.LRIG2 gene is overexpressed in invasive pituitary adenoma.It may play an important role in pituitary adenoma invasiveness and further studies are necessary to elucidate the mechanism under this phenomenon.

Identification of CDK6 and RHOU in Serum Exosome as Biomarkers for the Invasiveness of Non-functioning Pituitary Adenoma

Objective To explore circulating biomarkers for screening the invasiveness of non-functioning pituitary adenomas(NF-PAs).Methods The exosomal RNAs were extracted from serum of patients with invasive NF-PA(INF-PA)or noninvasive NF-PA(NNF-PA).Droplet digital PCR was adapted to detect the mRNA expression of candidate genes related to tumor progression or invasion,such as cyclin dependent kinase 6(CDK6),ras homolog family member U(RHOU),and spire type actin nucleation factor 2(SPIRE2).Student’s t-test was used to analyze the statistical difference in the mRNA expression of candidate genes between the two groups.Receiver operating characteristic(ROC)curve was used to establish a model for predicting the invasiveness of NF-PAs.The accuracy,sensitivity,specificity and precision of the model were then obtained to evaluate the diagnostic performance.Results CDK6(0.2600±0.0912 vs.0.1789±0.0628,t=3.431,P=0.0013)and RHOU mRNA expressions(0.2696±0.1118 vs.0.1788±0.0857,t=2.946,P=0.0052)were upregulated in INF-PAs patients’serum exosomes as compared to NNF-PAs.For CDK6,the area under the ROC curve(AUC)was 0.772(95%CI:0.600-0.943,P=0.005),the accuracy,sensitivity,specificity and precision were 77.27%,83.33%,75.00%and 55.56%to predict the invasiveness of NF-PAs.For RHOU,the AUC was 0.757(95%CI:0.599-0.915,P=0.007),the accuracy,sensitivity,specificity and precision were 72.73%,83.33%,68.75%and 50.00%.In addition,the mRNA levels of CDK6 and RHOU in serum exosomes were significantly positively correlated(r=0.935,P<0.001).After combination of the cut-off scores of the two genes,the accuracy,sensitivity,specificity and precision were 81.82%,83.33%,81.25%and 62.50%.Conclusions CDK6 and RHOU mRNA in serum exosomes can be used as markers for predicting invasiveness of NF-PAs.Combination of the two genes performs better in distinguishing INF-PAs from NNF-PAs.These results indicate CDK6 and RHOU play important roles in the invasiveness of NF-PAs,and the established diagnostic method is valuable for directing the clinical screening and postoperative treatment.

UROKINASE-TYPE PLASMINOGEN ACTIVATOR,ITS RECEPTOR AND INHIBITOR EXPRESSION IN HEPATOCELLULAR CARCINOMA RELATION TO CANCER INVASIVENESS AND PROGNOSIS

Objective: To study the relevance of uPA, uPAR and PAI 1 to hepatocellular carcinoma (HCC) Methods: The expression at protein level of uPA, uPAR and PAI 1 was determined in 48 cases of HCC and 12 cases of benign tumors of liver (as control) by immunohistochemistry Results: When compared to cancer adjacent liver tissue and the control, positive rate of immune staining for uPA, uPAR and PAI 1 on cell membrane were significantly higher in HCC cells ( P <0 05) Positive staining of uPA and uPAR was seen in 16 of 22 and 19 of 22 cases of HCC with invasion, respectively ( P <0 01 and P <0 001) In 8 of 8 cases with cancer embolus, and in 6 of 6 cases with lymph node metastasis was the expression of positive uPAR Compared with 2 of 17 cases without recurrence, uPAR was positive in 15 of 17 recurrent cases ( P <0 01) In 36 cases who survived, 17 was positive uPAR and 15 positive PAI 1, while in 12 cases who died 2 years after surgery, 12 were positive for uPAR and 9 positive PAI 1, respectively ( P <0 01 and P <0 05) In 15 positive cases for all three parameters, 11 had cancer invasion and 7 died within 2 years, while in negative cases, 2 had invasion and none died within 2 years ( P <0 05) Conclusion: Expression of uPA, uPAR and PAI 1 is increased in HCC, uPA and uPAR may contribute significantly to HCC invasion and metastasis uPAR and PAI 1 are associated with poor prognosis of HCC

Inhibition of invasiveness and expression of epidermal growth factor receptor in human colorectal carcinoma cells induced by retinoic acid

Human amniotic basement membrane (HABM) model and agarose drop explant method were used to in-vestigate the effects of retinoic acid(RA) on the invasive-ness alld adhesiveness to the basement membrane, and the migration of a highly invassive human colorectal cancer cell line CCL229. Results showed that 5 ×106 MRA markedly reduced the in vitro invasiveness and adhesiveness to the HABM, and the migration of the CCL229 cells. In addi-tion, to elucidate the relation between expression of epider-mal growth factor receptor (EGFR) and the invasiveness of the colorectal carcinoma cells, two well-differentiated, but with different invasiveness colorectal cancer cell lines were compared at mRNA level for expressioll of EGFR by using EGFR cDNA probe labeled with digoxigenin (DIG). Expression of EGFR was showll to be markedly higher in the highly invassive CCL229 cells than that in the low in- vasive CX-1 cells. Furthermore, expression of EGFR in RA treated CCL229 cells gradually decreased with time,the level being the lowest on day 6 of the RA treatment.

The Expression of Integrinβ1 and FAK in Pituitary Adenomas and Their Correlation with Invasiveness

The expression and possible role of integrin-focal adhesion kinase signal pathway in invasive pituitary adenomas were explored. Forty-nine human pituitary adenomas were detected for the expression of integrinβ1 （INTβ1） and focal adhesion kinase （FAK） by immunohistochemistry, and their correlation with the invasiveness of pituitary adenomas as well as between themselves was ana- lyzed. The results showed that INTβ1 was expressed in 46 cases （93.9%） and FAK in 36 cases （73.5%）, respectively, and their expression levels were highly correlated with tumor invasiveness, but not with the tumor types. It was suggested that the integrin-focal adhesion kinase signal pathway plays a role in the invasiveness of pituitary adenomas.

Assessment for the minimal invasiveness of laparoscopic liver resection by interleukin-6 and thrombospondin-1

BACKGROUND Laparoscopic surgery has been introduced as a minimally invasive technique for the treatment of various field.However,there are few reports that have scientifically investigated the minimally invasive nature of laparoscopic liver resection(LLR).AIM To investigate whether LLR is scientifically less invasive than open liver resection.METHODS During December 2011 to April 2015,blood samples were obtained from 30 patients who treated with laparoscopic(n=10,33%)or open(n=20,67%)partial liver resection for liver tumor.The levels of serum interleukin-6(IL-6)and plasma thrombospondin-1(TSP-1)were measured using ELISA kit at four time points including preoperative,immediate after operation,postoperative day 1(POD1)and POD3.Then,we investigated the impact of the operative approaches during partial hepatectomy on the clinical time course including IL-6 and TSP-1.RESULTS Serum level of IL-6 on POD1 in laparoscopic hepatectomy was significantly lower than those in open hepatectomy(8.7 vs 30.3 pg/mL,respectively)(P=0.003).Plasma level of TSP-1 on POD3 in laparoscopic hepatectomy was significantly higher than those in open hepatectomy(1704.0 vs 548.3 ng/mL,respectively)(P=0.009),and have already recovered to preoperative level in laparoscopic approach.In patients with higher IL-6 Levels on POD1,plasma level of TSP-1 on POD3 was significantly lower than those in patients with lower IL-6 Levels on POD1.Multivariate analysis showed that open approach was the only independent factor related to higher level of IL-6 on POD1[odds ratio(OR),7.48;95%confidence interval(CI):1.28-63.3;P=0.02].Furthermore,the higher level of serum IL-6 on POD1 was significantly associated with lower level of plasm TSP-1 on POD3(OR,5.32;95%CI:1.08-32.2;P=0.04)in multivariate analysis.CONCLUSION In partial hepatectomy,laparoscopic approach might be minimally invasive surgery with less IL-6 production compared to open approach.

Effect of ERBB2 expression on invasiveness of glioma TJ905 cells

Objective:To investigate the influence and possible mechanism of ERBB2 expression on the invasiveness of glioma cells.Methods:Glioma TJ905 cells were separated and cultured.ERBB2shRNA and overexpressing vectors were constructed,which were then transfected.The ERBB2expression was up-regulated or down-regulated.Changes of invasiveness of TJ905 cells were detected by Transwell assay,and the expressions of matrix metalloprotease(MMP)-2 and MMP-9were measured by Western bolt.Results:ERBB2 shRNA transfection vector could effectively inhibit expression of ERBB2:while ERBB2 overexpressing vector transfection could significantly improve the expression of ERBB2 in TJ905 cells.Transwell assay showed that when ERBB2expression was down-regulated,the invasiveness of TJ905 cells was notably decreased;when ERBB2 expression was up-regulated,the invasiveness of TJ905 cells was markedly increased.Meanwhile.Western blot indicated that down-regulating ERBB2 inhibited the expression of MMP-2 and MMP-9,while up-regulating ERBB2 enhanced their expressions.Conclusions:ERBB2 expression is closely related in the invasiveness of glioma TJ905 cells.

Tumor necrosis factor-alpha mediates hyperthermia-induced glioma invasiveness decreases

Background： Thermothempy has already been proved effective for the treatment of various tumors, including glioma. This study was performed to determine whether tumor necrosis factor-alpha was involved in the regulation of this biological process. Methods： RT-PCR and immunocytochemistry were used to investigate the levels of tumor necrosis factor-alpha mRNA and heat shock factor-1 protein, respectively, in glioma cells. Radioimmunoassay was used to dynamically monitor contents of TNF-α in nutrient fluid for C6 cells after hyperthermia treatment. Crystal violet staining method was used to detect glioma invasiveness. Results： The most obvious increase of heat shock factor-1 protein and tumor necrosis factor -alpha mRNA in C6 cells were observed at 30 min and 60 min after hyperthermia, respectively. In addition, the radioactivity of tumor necrosis factor-alpha in C6 cells＇ culture fluid also reached peak at 120 min of hyperthermia. The glioma invasiveness decreases and the concentration of tumor necrosis factor-alpha reached the maximum at 120 min of hyperthermia. Conclusion： Our results showed that the hyperthermia-mediated glioma invasiveness decreases was due to accelerated release of tumor necrosis factor-alpha,which could cause the decreases of glioma invasiveness by promoting the release heat shock factor-1 from neurospongioma cells .

The neurotrophic factor Artemin promotes the motility and invasiveness of MIA PaCa-2 pancreatic cancer cells

Objective： The aim of this study was to analyze the capacity of Artemin promoting the motility and invasiveness of MIA PaCa-2 pancreatic cancer （PAC） cells. Methods： The PAC cell line MIA PaCa-2 was cultured in vitro and studied using Transwell chamber analysis. The motility and invasiveness ability affected by different concentrations of Artemin and its receptor GFRa3 were determined. Expression level of matrix metalloproteinase-2 （MMP-2）, epithelial cadherin （E-cadherin） were quantitative analysis using RT-PCR and Western blot in MIA PaCa-2 cells stimulated with Artemin and receptor GFRa3. Results： MIA PaCa-2 PAC cell motility and invasiveness was significantly increased with Artemin and its receptor GFRa3 increasing concentrations than control （P 〈 0.01）. 150 ng/mL was the best of both the role of concentration. MMP-2 was increased significantly （t = 6.35, t = 7.32）, while E-cadherin was significantly lower （t = 4.27, t = 5.61）, after affected by the 150 ng/mL Artemin and GFRα3,respectively. The difference was statistically significant compared with the control group （P 〈 0.01）. Conclusion： Artemin and its receptor GFRa3 can promote PAC cell motility and invasiveness ability and contribute to the aggressive behavior. The mechanism may be related to increased expression of MMP-2 molecule and E-cadherin downregulation expression.

IN VIVO COMPARATIVE OBSERVATION ON THE INVASIVENESS OF VARIOUS ORGANS BY DIFFERENT LEUKEMIA CELLS

Using patho-morphological method and transplantation bio-assay, the in vivo invasiveness of leukemia cells is three transplantable mouse T cell leukemia models was comparatively studied. The results showed that the invasion to the liver was consistent, but that to other organs was obviously different. L615 and L7212 leukemia cells preferred to the bone marrow and spleen than to the peritoneum while L7811 leukemia cells were just the opposite. Transplantation bio-assay demonstrated that leukemia cells were present in the bone marrow of L615 mice as early as 6 hours after leukemic cell inoculation, but no leukemia cells was detected in bone marrow of L7811 mice 2 days after inoculation. In the terminal phase, L615 mice bone marrow became filled with leukemia cells, but L7811 mice bone marrow contained only a few leukemia cells. The difference of invasiveness of leukemia cells among organs is probably related to "homing" receptor. The same type of leukemia cells may possess multiple "homing" receptor.

Vibramycin inhibits the expression of MMP-2 and the invasiveness of PC-3 cells in vitro

Objective To study the inhibition of vibramycin on the expression of matrix metalloproteinase-2 (MMP-2) and the invasiveness of androgen-independent prostatic carcinoma cell line PC - 3 in vitro. Methods Immunohistochemistry stain and transwell chamber were used to investigate the expression of MMP-2 in different concentration of vibramycin treated PC-3 cells and the invasive ability of different concentration of vibramycin treated PC-3 cell. Results The positive rate of MMP-2 inJune 2003 Vol12 No2 PC-3 cells was decreased at a concentration of 5 mg/L of vibramycin and decreased dramatically at the concentration of 10 mg/L. The cells moved throuth the membrane was(82. 0 ± 4.6)/field in the control group, while decreased to(26.1 ±3.6),(7.2 ±2.2) and(3.3± 0.7)/field in 5,10 and 20 mg/L vibramycin treated PC-3 cell respectively. Conclusion Vibramycin can inhibit the invasiveness and metastatasis of PC-3 cells, the mechanism of which is related to the inhibition of MMP-2 in PC-3 cell.10refs.

Lower serum folate is associated with development and invasiveness of gastric cancer

AIM: To evaluate the associations of serum folate levelwith development, invasiveness and patient survival of gastric cancer. METHODS: In this nested case-control study, patients with newly diagnosed gastric cancer undergoing gastrectomy were enrolled, and patients receiving chemotherapy prior to surgery, with other concurrent malignancy, or of the aboriginal and alien populations were excluded. In total, 155 gastric cancer patients and 149 healthy controls were enrolled for determination of serum folate levels and their correlation with gastric cancer. Using the median value of serum folate computed among the overall population as the cutoff value, the associations between serum folate and gastric cancer in all cases and different age and gender subgroups were analyzed by multivariate logistic regression analysis. In the patient cohort of gastric cancer, receiver-operating characteristic analyses were performed to calculate the best cutoff values of serum folate, and the associations between serum folate levels and clinicopathological features were further analyzed by multivariate regression analysis. Survival analyses were conducted using the Cox proportional hazards model.RESULTS: The mean serum folate level was significantly lower in gastric cancer patients than that in controls(3.71 ± 0.30 ng/mL vs 8.00 ± 0.54 ng/mL, P < 0.01), and folate levels were consistently lower in gastric cancer patients regardless of age and gender(all P < 0.01). Using the median serum folate value as the cutoff value, low serum folate was significantly associated with gastric cancer risk in the whole population(OR = 19.77, 95%CI: 10.54-37.06, P < 0.001) and all strata(age < 60 years OR = 17.39, 95%CI: 7.28-41.54, age ≥ 60 years(OR = 21.67, 95%CI: 8.27-56.80), males(OR = 17.95, 95%CI: 7.93-40.62), and females(OR = 20.95, 95%CI: 7.66-57.31); all P < 0.001. In the patient cohort of gastric cancer, the respective cutoff values showed that low serum folate levels were significantly associated with serosal invasion(OR = 2.54, 95%CI: 1.23-5.23), lymphatic invasion(OR = 2.23, 95%CI: 1.17-4.26), and liver metastasis(OR =6.67, 95%CI: 1.28-34.91) of gastric cancer(all P < 0.05). Serum folate level below 1.90 ng/mL was associated with poor patient survival(HR = 1.84, 95%CI: 1.04-3.27, P < 0.05) in univariate analysis.CONCLUSION: Lower serum folate levels were significantly associated with gastric cancer development and invasive phenotypes. The role of folate depletion in gastric cancer invasion warrants further study.

Silencing MTA1 by RNAi Reverses Adhesion, Migration and Invasiveness of Cervical Cancer Cells (SiHa) via Altered Expression of p53, and E-cadherin/β-catenin Complex

It has been reported that metastasis-associated gene 1 （Mta1） is overexpressed in many malignant tumors with high metastatic potential. In addition, some studies indicated that MTA1 participated in invasion, metastasis, and survival of cancer cells by regulating cell migration, adhesion and proliferation. But the role of MTA1 is unclear in vitro in the development of cervical cancer cells. This study investigated whether and how MTA1 mediated cell proliferation, migration, invasion and adhesion in cervical cancer. MTA1 expression level was detected by Western blot in two cervical cancer cell lines of different invasion potentials. The effects of MTA1 expression on SiHa cell apoptosis, cycle, proliferation, migration, invasion and adhesion were tested by flow cytometry, MTT, wound-healing assay, Transwell assay and adhesion assay, respectively. The expression levels of p53, E-cadherin, and β-catenin activity were evaluated in untreated and treated cells. The results showed that MTA1 protein expression was significantly higher in SiHa than in HeLa, which was correlated well with the potential of migration and invasion in both cell lines. Furthermore, the cell invasion, migration and adhesion capabilities were decreased after inhibition of MTA1 expression mediated by Mta1-siRNA transfection in SiHa. However, no significant differences were found in cell apoptosis, cycle, and proliferation. In addition, E-cadherin and p53 protein levels were significantly up-regulated, while β-catenin was significantly down-regulated in SiHa transfected with the siRNA. These results demonstrated that MTA1 played an important role in the migration and invasion of cervical cancer cells. It was speculated that the decreased migration and invasion capability by inhibiting the MTA1 expression in the SiHa cell line may be mediated through the altered expression of p53, and E-cadherin/β-catenin complex. MTA1 could serve as a potential therapeutic target in cervical cancer.

Targeted Silencing of Heparanase Gene by Small Interfering RNA Inhibits Invasiveness and Metastasis of Osteosarcoma Cells

The effects of targeted silencing of heparanase gene by small interfering RNA（siRNA） on invasiveness and metastasis of osteosarcoma cells（MG63 cells） were investigated in the present study.Two complementary oligonucleotide strands were synthesized and inserted into pGenesil-1 vector based on the mRNA sequence of heparanase gene.The expression vector containing short hairpin RNA（pGenesil-shRNA） was constructed successfully.MG63 cells were randomly allocated into 3 groups：blank group,empty vector（pGenesil） transfected group and expression vector（pGenesil-shRNA） transfected group.Under the induction of Lipofectamine 2000,the recombinants were transfected into MG63 cells.Heparanase gene expression level was detected by RT-PCR and Western blotting.Cell prolifera-tion was measured by MTT assay.Cell invasiveness and metastasis were examined by cell adhesion and Transwell-ECM assays.HUVECs migration assay was applied for the detection of angiogenesis.As compared with negative controls,the mRNA and protein expression levels of heparanase were down-regulated by 76.1%（P0.01） and 75.3%（P0.01） respectively in the pGenesil-shRNA transfected group.Meanwhile,the proliferation,adhesiveness,invasiveness and angiogenesis properties of MG63 cells were all significantly inhibited.It was suggested that targeted silencing of heparanase gene by siRNA could dramatically inhibit the invasiveness and metastasis of osteosarcoma cells.

Methanol extract of Codium fragile inhibits tumor necrosis factor-ɑ-induced matrix metalloproteinase-9 and invasiveness of MDA-MB-231 cells by suppressing nuclear factor-κB activation

Objective:To evaluate whether the methanol extract of Codium fragile(MECF) regulates tumor necrosis factor-α(TNF-α)-induced invasion of human breast cancer MDA-MB-231 cells by suppressing matrix metalloproteinase-9(MMP-9).Methods:Reverse transcriptionpolymerase chain reaction(RT-PCR) and western blot analysis were performed to analyze the expression of MMP-9 and nuclear factor-κB(NF-κB) subunits,p65 and p50,and IκB in MDA-MB-231 cells.3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide(MTT) assay was used for cell viability.MMP-9 activity and invasion were measured by gelatin zymography and a matrigel invasion assay,respectively.NF- κB activity was measured by an electrophoretic mobility shift assay and luciferase activity.Results:MECF had no effects on cell viability up to a concentration of 100 μg/mL in human breast cancer MDA-MB-231 cells regardless of the presence of TNF-α.MDA-MB-231 cells that were stimulated with TNF-α showed a marked increase of invasion compared to the untreated control,whereas pretreatment with MECF downregulated the TNF-α-induced invasion of MDA-MB-231 cells.Additionally,zymography,western blot analysis,and reverse transcriptase-polymerase chain reaction(RT-PCR) confirmed that MECF decreased TNF-α-induced MMP-9 expression and activity which is a key regulator for cancer invasion.According to an electrophoretic morbidity shift assay,pretreatment with MECF in MDA-MB-231 cells significantly decreased the TNF-α-induced DNA-binding activity of nuclear factor- κB(NF- κB),which is an important transcription factor for regulating cancer invasion-related genes such as MMP-9.Furthermore,treatment with MECF sustained the expression of p65 and p50 in response to TNF-α in the cytosolic compartment.The luciferase assay demonstrated that MECF attenuated TNF-α-induced NF- κB luciferase activity.Conclusion:MECF exhibited its antiinvasive capability by downregulating TNF-α-induced MMP-9 expression,resulting from the suppression of NF- κB activity in the human breast cancer cell line MDA-MB-231.