Effect of gonadotropin-releasing hormone analogue on human breast cancer cell line in vitro

AIM: To investigate the effects of gonadotropin-releasing hormone (GnRH) analogue on the growth of breast cancer cell lines MCF-7 and MDA-MB-231 in vitro and to explore the related mechanisms with PI3K/Akt or ERK/MAPK pathways. METHODS: The proliferation of human breast cancer cell lines MCF-7 and MDA-MB-231 treatment with triptorelin was detected by MTT assay and the distribution of the cell cycle was determined by flow cytometry. The phosphorylation of the ERK1/2 and Akt was evaluated by Western blotting. RESULTS: Triptorelin inhibited the proliferation of MCF-7 cells at concentration of 10^-5 mol/L after treated for 192 h or at concentration of 10^-4 mol/L after treated for 168 h and 192 h. Triptorelin inhibited the proliferation of MDA-MB-231 cells at concentration of 10^-4 mol/L after treated for 192 h (P<0.05).Treatment with triptorelin for 192 h at concentration of 10^-4 mol/L had no statistical significance effect on phosphorylation of ERK1/2 and Akt(P>0.05).CONCLUSION: Inhibitory effect of GnRH analogue triptorelin on human breast cancer cells is not just the connection with the down-regulation of pituitary hormone, but also a direct inhibitory effect. The role may not be involved in the activation of ERK/MAPK and PI3K/Akt signaling pathways.     

Possible mechanisms of acquired resistance to 5-FU in human colon cancer

AIM: To explore the mechanisms of resistance to 5-fluorouracil(5-FU) in human colon cancer cells.METHODS: 5-FU-resistant cell lines were established and their IC50 were calculated by detection of cell survival rate.Western blotting was performed to analyze the expression of several proteins,by which the possible mechanisms of acquired resistance to 5-FU were determined in human colon cancer cells.RESULTS: The resistant cells were resistant to 5-FU-induced S phase arrest as well as the expression of DNA damage marker-phosphor-histone H2A.X.Furthermore,data demonstrated that over-expression of Bik,Bcl-Xs,and Bcl-XL proteins were observed in 5-FU-resistant cell lines.However,the DLD1/Bcl-XL cells were only partially resistant to 5-FU-induced apoptosis,but not 5-FU-induced S phase arrest and phosphor-histone H2A.X.CONCLUSION: Over-expression of Bcl-XL protein certainly contributes to acquired 5-FU resistance in human colon caners,but has no effect on 5-FU-induced DNA damage and cell cycle disorder,suggesting that other mechanisms are involved in acquired resistance to 5-FU in human colon cancer.     

Effects of furin inhibitor on metastasis of human breast cancer MCF-7 cells

AIM：To investigate the mechanism underlying breast cancer metastasis and to provide theoretical data for studying the pathogenesis of breast cancer onset and development. METHODS：Human breast cancer MCF-7 cells were treated with different concentrations of furin inhibitor α1-PDX for 48 h. Wound healing assay and Transwell assay were applied to detect the migration and invasion abilities of the MCF-7 cells. The expression of cell migration-associated proteins, including membrane-type 1 matrix metalloproteinase (MT1-MMP), vascular endothelial growth factor (VEGF)-C and VEGF-D, was determined by Western blotting. The protein levels of MMP2 and MMP9 in the supernatant were measured by ELISA. RESULTS：Compared with control group, 200 nmol/L of furin inhibitor exerted significant inhibitory effects on the cell migration (P<0.05). The expression of cell migration-associated proteins MT1-MMP, VEGF-C and VEGF-D was significantly inhibited after treated with α1-PDX (P<0.05). Significant inhibitory effects of α1-PDX on the expression of MMP9 and MMP2 (P<0.05) in the supernatant were observed. CONCLUSION：Furin inhibitor suppresses the metastasis of MCF-7 cells via down-regulating the expression of MMPs and VEGFs.     

Effect of combined treatment with GnRHa and GH on linear growth in mid- and late pubertal girls at great bone ages with CPP or EFP and relation to CNP signaling pathway

AIM:To investigate the effect of combined treatment with gonadotropin-releasing hormone analogue (GnRHa) and growth hormone (GH) on the linear growth in mid- and late pubertal girls at great bone ages with central precocious puberty (CPP) or early and fast puberty (EFP), and to determine the relation between C-type natriuretic peptide (CNP) signaling pathway and the accelerative effect of GH on long bone growth in these girls. METHODS:Twenty-two girls were diagnosed as CPP or EFP, whose bone ages were older than 11.5 years with impaired predicted adult height (PAH), and divided into GnRHa treatment group (treated with GnRHa alone, slow-release of triptorelin 60~80 μg/kg every 4 weeks, im) and combined treatment group (treated with GnRHa and GH, 1 U/kg GH every week for 6~7 times, sc). The height, weight and pubertal stage were determined every 3 months. At the beginning and after 6 months of the treatment, the bone age was evaluated and the serum concentrations of amino-terminal pro-C-type natriuretic peptide (NTproCNP), insulin-like growth factor 1 (IGF-1) and procollagen type 1 amino-terminal propeptide (P1NP) were measured. Height velocity (HV), height SD score for bone age (HtSDSBA), PAH and the serum indexes mentioned above were compared at the beginning and the end of the treatment. RESULTS:After 6 months of the treatment, HV, ΔHtSDSBA and ΔPAH of the girls treated with GnRHa+GH were statistically higher than those of the girls given GnRHa alone (P<0.01). Serum concentrations of NTproCNP, P1NP and IGF-1 were not significantly different between the beginning and the end of the 6-month combined treatment. The girls treated with GnRHa alone showed a significant decrease in both serum NTproCNP and P1NP levels (P<0.05) and no significant change of serum IGF-1 level after 6 months of the treatment. CONCLUSION: In the CPP or EFP girls who are in mid- and late puberty and at great bone ages, the combined treatment with GnRHa and GH may accelerate linear growth and improve predicted adult height. This effect of GH is not attributed to the change of serum IGF-1 level, and may be related in part to the acceleration of CNP-mediated long bone growth.     

Expression of 2 SYK protein isoforms in breast cancer cells and their effect on cell invasion

AIM: This study was to explore the expression of spleen tyrosine kinase(SYK)(L) and SYK(S) in breast cancer cells and their effects on invasive phenotype of breast cancer cell line MB435. METHODS: The expression of SYK in breast cancer cell lines MB435, ZR75.1, MB468, T47D and MCF 7 were detected by Western blotting. MB435 cells were transfected with pcDNA3.1-SYK(L), pcDNA3.1-SYK(S) or pcDNA3.1 respectively, G418-resistant stable clones were pooled and the SYK expression was measured by Western blotting. Chemoinvasion assays were performed to study the effects of SYK(L) and SYK(S) on breast cancer cell invasion.RESULTS: SYK(L) and SYK(S) were simultaneously expressed in most detected breast cancer cells. Pooled SYK(L) and SYK(S) stable clones were made by Fugene 6 transfection and G418 selection. The expression of SYK(L) suppressed the invasive ability of breast cancer cells, while SYK(S) didnt.CONCLUSION: The simultaneously expression of SYK(L) and SYK(S) in breast cancer cells is a common phenomenon. Among SYK(L) and SYK(S),only SYK(L) protein is capable of suppressing the invasion of breast cancer cells.     

Survivin-2B induces apoptosis of human breast cancer cells

AIM：To explore the role of survivin-2B in the process of tumor cell apoptosis. METHODS：The survivin-2B gene was cloned into pcDNA3.1 vector and the recombinant plasmid pcDNA3.1-survivin-2B was obtained. Human breast cancer MCF7 cells were transfected with pcDNA3.1 and pcDNA3.1-survivin-2B using Lipofectamine 2000. The cell cycle was determined by propidium iodide staining, and the apoptosis was detected by annexin V/7-AAD staining 48 h after transfection. Meanwhile, tatal RNA was extrated and multiplex polymerase chain reaction based on GenomeLab GeXP Genetic Analysis System was performed to detect the expression of 21 tumor-related genes. RESULTS：Flow cytometry analysis indicated that over-expression of survivin-2B promoted the apoptosis and cell cycle arrest of MCF7 cells. Compared with control group, totally 10 differential expressed genes were related to the over-expressed survivin-2B, among which 2 were up-regulated and 8 were down-regulated. The expression of aldehyde dehydrogenase 4 family member A1 (ALDH4A1) was 48% down-regulated, and the expression of protein regulator of cytokinesis 1 (PRC1) was 1.08 folds up-regulated. CONCLUSION：Survivin-2B induces the expression changes of some tumor-related genes, which results in the apoptosis and G2/M arrest of MCF7 cells.     

Effects of lutein on viability of breast cancer cells

AIM:To investigate the effect of lutein on the viability of breast cancer cells and its possible mechanism. METHODS:The human breast cancer T47D cells were divided into control group and lutein (6.25, 12.5, 25, 50 mg/L) treatment groups. The effect of lutein on the viability of T47D cells was measured by MTT assay. The mRNA expression of nuclear factor erythroid 2-related factor 2 (Nrf2), glutathione peroxidase 1 (GPx1) and superoxide dismutase 2(SOD2) was detected by RT-qPCR. Fluorescent probes DCFH-DA was used to determine the production of reactive oxygen species (ROS). The protein expression of Nrf2 and p65 was determined by Western blot. RESULTS:The MTT results showed that lutein inhibited T47D breast cancer cell viability in a dose-and time-dependent manner. The RT-qPCR results showed that the mRNA levels of Nrf2, GPx1 and SOD2 were higher in lutein treatment groups than those in the control group (P<0.05), and with the increased concentrations and extension of intervention time of lutein, the relative mRNA levels were all increased. The ROS levels were significantly decreased in the lutein-treated groups (P<0.05). The results of Western blot demonstrated that the protein expression of Nrf2 was significantly increased (P<0.05), and p65 protein was decreased (P<0.05) in a dose-dependent manner with lutein treatment for 48 h. CONCLUSION:Lutein significantly inhibits the viability of breast cancer cells, and the inhibition roles may be related to up-regulation of the expression of Nrf2, antioxidant enzymes GPx1 and SOD2 mRNA expression and down-regulation of oxidative stress, thus blocking the NF-κB signaling pathway.     

Relationship between zinc and zinc transporter expression in breast cancer MCF-7 cells

AIM: To study the changes of zinc transporter gene expression in MCF-7 cell line exposed to ZnCl₂ and TPEN. METHODS: Human breast cancer cell line MCF-7 was exposed to different concentrations of ZnCl₂ (0, 50, 100, 150, 200 μmol/L) and TPEN (0, 5, 10, 15 μmol/L), respectively. Twelve hours later, the cell viability was measured by MTT and levels of zinc transporter mRNA by RT-PCR. Zinquin was used to estimate the intracellular zinc concentrations. RESULTS: MCF-7 cells viability rate was significantly decreased when exposed to ZnCl₂ (with 150 μmol/L and 200 μmol/L) and TPEN. The intracellular zinc concentration was significantly increased when exposed to ZnCl₂ and decreased when exposed to TPEN. ZnT-1 mRNA level was increased along with the increasing concentration of ZnCl₂ but decreased when exposed to TPEN. The expressions of ZIP2 and ZIP10 were increased along with the increasing concentration of TPEN. CONCLUSION: ZnT-1 gene expression is induced by zinc supplement and repressed by zinc deficiency. ZIP2 and ZIP10 gene expressions are induced by zinc deficiency.     

Effects of p21-activated protein kinase 2 down-regulation on proliferation and apoptosis of human breast cancer cells

AIM：To study the effect of p21-activated protein kinase 2 (PAK2) knockdown by RNA interference on the proliferation and apoptosis of human breast cancer cells. METHODS：The short hairpin RNA (shRNA) targeting PAK2 gene was designed and used for packing lentivirus in 293T cells.Human breast cancer MCF-7 cells were infected by the virus particles and PAK2 knockdown stable cell line was established by puromycin selection. The knockdown efficiency was assessed by Western blotting. The proliferation ability of MCF-7 cells was evaluated by CellTiter 96 AQueous and anchorage-independent growth assays. The cell apoptosis induced by staurosporine was detected by flow cytometry. RESULTS：The protein level of PAK2 was significantly suppressed after silencing of PAK2 gene in MCF-7 cells (P<0.01). Furthermore, knockdown of PAK2 caused remarkable inhibition of the cell proliferation and colony formation (P<0.01). Staurosporine induced more apoptosis in the PAK2 knockdown cells compared with the control cells (P<0.01). CONCLUSION：Knockdown of PAK2 inhibits the proliferation of MCF-7 cells and increases the sensitivity of chemotherapeutic drug-induced cell apoptosis, suggesting that PAK2 might be a new therapeutic target in breast cancer treatment.     

Effects of curcumin analogue B50 on proliferation and apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance

AIM: To compare the effects of B50, a mono-carbonyl analogue of curcumin, on the proliferation and apoptosis between homologous nasopharyngeal carcinoma cells CNE-2R and CNE-2 with different radioresistance.METHODS: The effects of B50 on cell viability and cell growth were detected by MTT assay and colony-forming experiment, respectively. The changes of cell cycle, apoptosis and mitochondrial membrane potential (MMP) were determined by flow cytometry.RESULTS: B50 inhibited the cell viability of CNE-2R cells in a time-and dose-dependent manner with the IC₅₀ of (8.06±0.14) μmol/L (24 h), (2.49±0.02)μmol/L (48 h) and (1.42±0.02) μmol/L (72 h), which was more effective than that in CNE-2 cells . The inhibitory effect of B50 on CNE-2R cell growth was more effective than that on CNE-2 cells . After treated with B50 for 48 h, the proportion of CNE-2R cells in G₂/M stage was increased from 7.1% to 34.9%, which was better than that of CNE-2 cells (from 12.4% to 35.7%). After treated with B50 for 24 h, the early apoptotic rate in CNE-2R cells was increased from 3.7% to 19.5%, which was better than that in CNE-2 cells (from 4.4% to 14.8%), and the MMP in CNE-2R cells was decreased by (43.17±3.11)%, which was better than that in CNE-2 cells .CONCLUSION: B50 is more effective on inhibiting the cell viability and cell growth, blocking the cell cycle at G₂/M stage, inducing apoptosis and decreasing MMP in CNE-2R cells than those in CNE-2 cells, indicating that B50 may enhance the radio-sensitivity of CNE-2R cells by blocking the cell cycle and inducing apoptosis through mitochondrial pathway.     

Effect of norcantharidin on proliferation and invasion of human breast cancer cell line SKBR3 in vitro

AIM: To investigate the effect of norcantharidin（NCTD）on proliferation and invasion of human breast cancer cell line SKBR3 in vitro and its anticancer mechanisms.METHODS: MTT assay was used to determine SKBR3 cell proliferation. Light and FACScan were used to detect apoptosis and cell cycle. The invasiveness of SKBR3 was evaluated by the adhesion test，Matrigel experiment and the crossing-river test.RESULTS: NCTD had inhibitive effects on growth of SKBR3 cells in a dose and time-dependent manner, with the IC₅₀ value of 12.5 mg/L at 24 h．The cells treated with 10 mg/L NCTD for 24 h and 48 h showed typical apoptotic morphology and hypodiploid peak before G₁ phase. The cell cycle was arrested at G₂/M phase. The apoptosis percentage was up to 3.44% and 6.17%, and the G₂/M percentage was up to 35.82% and 38.70%. NCTD also could inhibit obviously the adhesion, movement and invasive capability simulating human basement membrane of SKBR3. Its effect was also in a dose-dependent manner. In the NCTD-treated group, crossing-river time was prolonged significantly and passing-membrane cells markedly decreased. CONCLUSION: NCTD in vitro inhibits not only the proliferation and growth of human breast cancer cells but also invasion and metastasis of the cells at relatively low concentration. NCTD shows prominent anti-tumor effects.     

Cloning,transfer and anti-human breast cancer cell growth of the extracellular domain of cadherin 5

AIM: In order to study the effect of the extracellular domain of cadherin 5 on the growth of a human breast cancer cell line MDA-MB435.METHODS: Cadherin extracellular domain repeats 1 to 4(CED1-4) was cloned by using RT-PCR technique,and inserted into the plasmid vector pMSCV.pMSCV-CED1-4 was propagated in XL-blue strain of Escherichia coli,extracted and purified.CED1-4 was cut by restriction endonuclease,examined by using agar gel electrophoresis,and finally sequenced.CED1-4 gene was transferred into MDA-MB435 cell line.The expression of CED1-4 gene in MDA-MB435 cell was analyzed by methods of RT-PCR and Western blotting.The effect of CED1-4 on the growth of MDA-MB435 cell was observed by the methods of proliferation experiments in vitro and the experiments in nude mice in vivo.RESULTS: The recombinant vector pMSCV-CED1-4 was successfully constructed.CED1-4 band appeared between the 1 636 bp and 1 018 bp in agar gel electrophoresis.The sequence result showed that CED1-4 had 1 452 bp and codes 484 amino acids.PCR and Western blotting identified that CED1-4 mRNA and protein were expressed in the transfected MDA-MB435 cells.Cell proliferation experiments showed that the proliferation rate of MDA-MB435 cells was lower in the experimental group than that in the experimental control group and the blank control group.The mean volume and weight of tumors in nude mice were lower in the experimental group than those in the experimental control group and the blank control group.CONCLUSION: The growth of a human breast cancer cell line MDA-MB435 is inhibited in vitro and in vivo by cadherin 5 extracellular domain CED1-4.     

Anti-angiogenic effect of thymoquinone on breast cancer

AIM: To investigate the anti-angiogenic effect of thymoquinone on human breast cancer and the possible mechanism. METHODS: The activities of human umbilical vein endothelial cells (HUVECs) and human breast cancer MCF-7 cells were assessed by CCK-8 assay. The apoptotic rate was measured by flow cytometry. Furthermore, the assay of capillary tube formation was used to observe the effect of thymoquinone on the tube formation of HUVECs. The protein levels of cleaved caspase-3, p-ERK and p-AKT were detected by Western blot. MCF-7 cells were subcutaneously injected into nude mice to establish breast xenograft tumors. After 3 weeks of implantation, the mice were randomized into control group and thymoquinone group. After the mice were sacrificed, immunohistochemistry was used to detect the expression of CD31 in the tumors, and the TUNEL test kit was used to explore cell apoptosis in the tumor tissues. RESULTS: Thymoquinone at concentrations of 20~80 nmol/L exerted no growth inhibitiory effect on MCF-7 cells. However, the cell viability of HUVECs were (66.1±8.3)％, (53.7±3.4)％ and (41.6±4.9)％ when the concentrations of thymoquinone were 20, 40 and 80 nmol/L, respectively. The apoptotic ratio of MCF-7 cells were (2.6±0.3)％, (2.4±0.3)％ and (4.6±0.4)％ and the apoptotic ratio of HUVECs were (21.5±3.7)％, (23.8±2.9)％ and (27.8±3.1)％ when the concentrations of thymoquinone were 20, 40 and 80 nmol/L, respectively. HUVECs were more sensitive to thymoquinone-induced apoptosis and inhibition in the cell activity than MCF-7 cells. Incubation of HUVECs with diffe-rent concentrations of thymoquinone (20, 40 and 80 nmol/L) for 1 h decreased their tube formation capacity (P<0.05). The protein level of cleaved caspase-3 was up-regulated, but the phosphorylation of AKT and ERK were inhibited in a concentration-dependent manner. The immunohistochemical analysis of CD31 showed significant difference of the integral absorbance between control group and thymoquinone group, and the TUNEL-positive cells in thymoquinone group was significantly more than that in control group. CONCLUSION: Thymoquinone has the anti-angiogenic effect on breast cancer both in vitro and in vivo, which may be related to the decreases in p-ERK and p-AKT.     

Role of miR-183 in breast cancer progression

AIM:To investigate the expression of miR-183 in breast cell lines and tissue specimens and its effects on the biological behaviors of breast cancer cells. METHODS:Human breast cell lines and clinical tissue specimens of breast diseases were used in the study. The expression of miR-183 was determined by real-time PCR. The characteristics of cell proliferation, invasion and migration were examined after miR-183 was transfected by lipofection. RESULTS:Altered expression of miR-183 was found in the highly invasive breast cancer cells as compared with weakly invasive breast cells. The expression of miR-183 was also significantly decreased in the breast cancer tissues as compared with that in the benign breast disease tissues. The capacities of breast cancer cell invasion and migration were significantly increased after transfection of miR-183 inhibitor and were decreased after transfection of miR-183 mimic. No significant change of cell proliferation was observed after miR-183 transfection. CONCLUSION:miR-183 may play an important role in breast cancer progression, especially in the cell invasion and migration.     

Effects of propofol on expression of CXCR4/CXCR7 and migration ability of human breast cancer MCF-7 cells in vitro

AIM:To investigate the effects of propofol on the expression of CXC-chemokine receptor (CXCR)4/CXCR7 and migration ability of human breast cancer MCF-7 cells in vitro. METHODS:MCF-7 cells were randomly divided into 4 groups,control group, lipid emulsion group, 3 mg/L and 8 mg/L propofol group. The cell viability was measured by MTT assay. The migration ability was detected by wound-healing assay and Transwell assay. The mRNA level of CXCR4/CXCR7 was detected by RT-qPCR. The protein expression leve of CXCR4/CXCR7 was determined by Western blot. RESULTS:Compared with control group, the scratching healing rates in 3 and 8 mg/L propofol group were decreased (P<0.05), and the chemotactic index was also decreased (P<0.05). The protein expression level of CXCR4/CXCR7 was decreased in 3 and 8 mg/L propofol group(P<0.05). However, both the mRNA level of CXCR4/CXCR7 and the viability of the MCF-7 cells kept no change. CONCLUSION:Propofol down-regulates the protein expression of CXCR4/CXCR7 and inhibits the migration ability of breast cancer MCF-7 cells in vitro.     

Relationship between expression of somatostatin and DNA ploidy in breast cancer

AIM: To investigate the relationship between somatostatin and the pathologic type, estrogen receptor,DNA ploidy of nuclei in tumor cells of breast cancer.METHODS: 67 cases of primary breast cancer and 25 cases of benign breast tumor were examined by immunohistochemical stretomyces avidin peroxidase method. 26 cases of breast cancer selected at random were analyzed by flow cytometry.RESULTS: Somatostatin expressed significantly higher in low malignant breast cancer than that in high malignant breast cancer (P<0.05). Most of cancers with positive staining of somatostatin were diploidy,most of cancers with negative staining were aneuploidy,there had significant difference between two groups (P<0.05).CONCLUSION: Somatostatin may delay the progress of breast cancer,and somatostatin levels in cancer tissues may become a useful indicator for assessing prognosis of patients with breast cancer.     

Effect of heat shock protein 90 on proliferation of human breast cancer cells and its molecular mechanisms

AIM: To investigate the effect of heat shock protein 90 (HSP90) on proliferation of human breast cancer cells by a specific inhibitor for HSP90,geldanamycin (GA),and to analyze its molecular mechanisms.METHODS: Human breast cancer cell line MDA-MB-435s was used as a model.Proliferation of MDA-MB-435s cell was measured with MTT assay.Cell cycle was analyzed by flow cytometry.Alteration of phosphorylated Stat3 protein and mutant p53 protein in cells was detected by Western blotting.RESULTS: Geldanamycin inhibited the proliferation of MDA-MB-435s cells in time-and dose-depending manners.After treatment with GA,MDA-MB-435s cell cycle was arrested at G2/M phase.The levels of phosphorylated protein Stat3 and mutant p53 protein in MDA-MB-435s cells were reduced obviously by 55% and 48% respectively after 48 hour treatment with GA at the concentration of 400 nmol/L,compared to control cells.CONCLUSION: Inhibition of HSP90 function can prevent breast cancer cell proliferation significantly,which may be related to down-regulatin of the phosphorylated Stat3 protein and mutant p53 protein in the cells.     

Relationship between c-erbB-2, BCSG1 expression and recurrence,metastasis in breast cancer

AIM: To assess the significance of c-erbB-2, BCSG1 (breast cancer specific gene-1) expression and other parameters in recurrence or metastasis of breast cancer. METHODS: The expression of c-erbB-2, BCSG1, and ER, PR, MVD, VEGF, VEGF-C, FLT-4, LVD were determined with the SP immunohistochemical method in 58 cases of invasive breast cancer patients occurred over 5 years. The cases were used to analyze the effect of c-erbB-2, BCSG1, VEGF-C and ER, PR, MVD, VEGF, FLT-4, LVD expression on clinical-pathological manifestations and prognosis in breast cancer. RESULTS: The expression rates of c-erbB-2, BCSG1, VEGF-C, LVD were respectively 25.9%, 62.1%, 36.2%, 32.8% in association with the lymph node metastasis and recurrence of breast cancer (P<0.05), the expression rate of MVD was also increased significantly (P＜0.05). CONCLUSION: The c-erbB-2, BCSG1, VEGF-C, LVD are highly expressed and strongly correlated with the lymph node metastasis and recurrence of breast cancer, of which BCSG1 may be used as a predictor of prognosis.