BACKGROUND AND AIM: Heparanase (Hpa) is a kind of endo_D_glucuronidase that degrades heparin sulfate proteoglycans. It played important roles in malignant tumor's invasion and metastasis. Both phosphomannopentaosesulfate (PI_88) and antisense oligodeoxynucleotide (ASODN) are Hpa inhibitors. This study was designed to observe inhibitory effects of PI_88 and Hpa ASODN on growth and angiogenesis of esophageal squamous cell carcinoma(ESCC) xenograft tumor in nude mice. MATERIALS AND METHODS: ESCC cell line TE_13 was injected subcutaneously in nude mice to establish the ESCC subcutaneous invasive model. The nude mice were randomized into 30 mg/kg PI_88 group, 2 mg/kg Hpa ASODN group, 20 mg/kg PI_88 and 1 mg/kg ASODN combined group and sterile saline control group. Each group contained 5 mice. Tumor volume and micro vessel density (MVD) were evaluated. Hpa expression was detected by RT_PCR and immunohistochemistry. RESULTS: On the 6th day after injection, all mice had palpable tumors. On the 21st day after injection, the tumor volume was significantly smaller in ASODN group, PI_88 group and combined group than in control group(P<0.01). The tumor volume in the combined group were less than in PI_88 alone and ASODN alone groups (P<0.01), and there was no difference between the latter two groups(P>0.05). The raitio of suppression in each group was 49.67％ (ASODN), 51.44％ (PI_88) and 83.00％ (combined group). The MVD was significantly less in ASODN, PI_88 and combined group than in control group(P<0.01). The MVD in PI_88 and combined group were less than in ASODN group(P<0.01), and there was no difference between PI_88 and combined group(P>0.05). Hpa gene expression in ASODN group was 0.45±0.12, PI_88 group 1.22±0.01, combined group 0.52±0.05, control group 1.33±0.05. Hpa protein expression in ASODN group was 43.40±7.80, PI_88 group 114.40±14.47, combined group 37.40±7.20, control group 121.20±11.90. Hpa gene and protein in ASODN and combined groups was less than in control group (Hpa gene: P<0.01, Hpa protein: P<0.01). But the expression of PI_88 group showed no difference compared with control group (P>0.05), and similarly between the ASODN and combined groups(P>0.05). CONCLUSION: Administratoring ASODN and PI_88 alone or in combination could all reduce the tumor growth. PI_88 could reduce the growth of tumor and angiogenesis, but had no effects on Hpa expression in vivo. Hpa ASODN could effectively reduce the expression of heparanase to slow down the growth of xenograft and angiogenesis. Combining the two showed synergistic effect, resulting in a much better result.

BACKGROUND AND AIM: To investigate the promoter methylation state of MGMT (O6_methylguanine DNA methyltransferase) gene and p16 gene in esophageal squamous cell carcinoma, and analyze the methylation frequencies of the two genes, thereby understanding the relationship between methylation of the promoters of MGMT gene and p16 gene in esophageal squamous cell carcinoma. MATERIALS AND METHODS: Methylation of the promoters of MGMT gene and p16 gene was evaluated by methylation_specific PCR in 44 esophageal carcinoma tissues, and carry out differential test and association analysis of the methylation frequencies of the two genes using χ2 test. RESULTS: Among the 44 esophageal squamous cell carcinomas, MGMT gene methylation was found in 18 cases and p16 gene methylation in 23 cases. Both MGMT and p16 gene methylation were present in 7 cases. Neither MGMT nor p16 gene methylation was detected in 10 cases. The difference of methylation frequencies of the two genes was insignificant(P>0.05). Differential test and association analysis of the methylation frequency of two genes was also insignificant. CONCLUSION: Methylation of the promoters of MGMT gene and p16 gene may both be closely associated with the initiation and progression of esophageal squamous cell carcinoma, but the methylations of the two genes took place independently.

BACKGROUND AND AIM: To study the roles of Bid in vitamin E succinate (VES)_induced apoptosis of human gastric carcinoma SGC_7901 cells. MATERIALS AND METHODS: The activation of Bid was detected by western blot. Bid was blocked by transient RNA interference (RNAi), and apoptotic rate, mitochondrial transmembrane potential (ΔΨm), the amount of cytosolic cytochrome C and the level of PARP cleavage were determined in Bid_negative cells and Bid_positive cells. RESULTS: Bid was activated by VES. Compared with Bid_positive cell, VES caused lower apoptotic rate, higher mitochondrial transmembrane potential, decreased release of cytochrome c to cytosol and lower level of PARP cleavage in Bid_negative cells. CONCLUSION: This study suggested Bid might play an important role in VES_induced apoptosis of human gastric carcinoma SGC_7901 cells.

BACKGROUND AND AIM: To express NRDRB1 in prokaryotic expression system for detecting enzyme activity and preparing polyclonal antibody. MATERIALS AND METHODS: We investigated the mRNA expression of NRDRB1 in cervical squamous carcinoma using RT_PCR. The total NRDR and NRDRB1 sequences were cloned and the coding regions were constructed to the Gateway_based expression vector (pDEST 17), which was transformed into the Escherichia coli(BL21_AI)for protein expression. The recombinant proteins were purified by affinity chromatography. RESULTS: We identified a novel alternatively spliced variant, NRDRB1, in HeLa cell and human cervical squamous carcinoma cells, characterized by a complete deletion of exon 3. The expression vectors of NRDR and NRDRB1 were constructed. The proteins, harvested at the optimal time point (4 hours after induction), were successfully expressed with a 30－50％ expression leve1. The homogeneous proteins were obtained by a one_step affinity chromatography. CONCLUSION: Recombinant NRDR and NRDRB1 from human cervical squamous carcinoma cells were expressed effectively in BL21_AI by pDEST 17.

BACKGROUND AND AIM:: To explore the effects of peripubertal exposures to bisphenol A on reproduction and development in male rats. MATERIALS AND METHODS: Sprague_Dawley weaning rats received by gavage 0, 5, 50 or 500 mg/kg/day of bisphenol A from post_natal day (PND) 23 to 53. The onset of puberty (progression of preputial separation) was measured on PND 37. Testis and epididymis were weighed and prepared for histological evaluation. Serum testosterone, and gene expression of steroidogenic enzymes in the testis were measured. RESULTS: A decrease in body weight was found in the 500 mg/kg group rats, but there was an increase of testis weight in 5 mg/kg group. There was no difference of serum testosterone between tested groups and control, but the concentration was higher in 50 mg/kg group than in 500 mg/kg group. The gene expressions of steroidogenic enzymes (P450Scc,17β_HSD and P450Arom) were altered in all dosage groups. CONCLUSION: Peripubertal exposure to Bisphenol A could induce changes in gene expression of some steroidogenic enzymes in the testis of rats.

BACKGROUND AND AIM: This study was designed to observe the growth inhibition of the chinese honeylocust fruit extract on esophageal squamous carcinoma cell line EC9706. MATERIALS AND METHODS: The changes of cell morphology were examined by light microscope MTT assay , TUNEL cell apoptosis detection and western blotting were used to get information of EC9706 cells treated with the extract. RESULTS: Different concentrations (100－200 μg /ml)of the chinese honeylocust fruit extract significantly inhibited the growth of EC9706 cells(P<0.01),and displayed a concentration_dependent effect(r=0.768).The extract could induce apoptosis and regulate the expression of bcl_2 protein in EC9706 cells. CONCLUSION: The extract from chinese honeylocust fruit could inhibit the growth and induce the apoptosis of EC9706 cells. These may be associated with the decreased expression of bcl_2 in EC9706.

BACKGROUND AND AIM: To evaluate the effects of selenium_enriched garlic on the gastric juice components and upper gastrointestinal tract stimulation in rats and beagle dogs. MATERIALS AND METHODS: Selenium_ enriched garlic was given to rats and beagle dogs for 10 days, stimulant response of the upper gastrointestinal tract and changes of the mucous membrane, gastric acid and pepsin of rat gastric juice were determined. RESULTS: The free hydrochloric acid, total acidity and activity and volume of pepsin in rats were obviously inhibited. Gastric secretion and total acidity increased at doses of 1 500 mg/kg and 3 750 mg/kg, and total acid output increased at dose of 3 750 mg/kg. There was gentle stimulation on upper gastrointestinal tract mucosa in rats treated with selenium_enriched garlic. There was gentle stimulation on upper gastrointestinal tract mucosa in beagle dogs treated repeatedly with 200 mg/kg selenium_enriched garlic; and medium stimulation in beagle dogs treated with 350 mg/kg and 450 mg/kg once. CONCLUSION: Selenium_enriched garlic could change the stomach intestine environment, suppress gastric acidity and pepsin enzymatic activity, promote gastric secretion and stimulate upper gastrointestinal tract mucosa.

BACKGROUND AND AIM: To evaluate the expression of 3 different transcripts and their clinical significance in human gastric adenocarcinomas. MATERIALS AND METHODS: The mRNA expressions of RASSF1A、RASSF1B and RASSF1C were determined by RT_PCR in gastric carcinomas BGC_823 cell lines、50 specimens of gastric carcinoma and matched normal tissues and 30 gastric biopsy specimens of high_risk group and those with possible gastric carcinoma. H.pylori in the 50 of gastric carcinomas was tested by PCR technique. RESULTS: Expression of RASSF1A mRNA was abnormally low in BGC_823 cell line, loss of expression rates of RASSF1A and RASSF1B in 50 gastric carcinomas were 52％(26/50) and 22％(11/50), respectively. RASSF1C mRNA was fully expressed. RASSF1A mRNA expression loss was positively related with histological differentiation degree (P<0.05),lymph node metastasis (P<0.05) and TNM stages(P<0.05). There was no relationship between RASSF1A expression and H.pylori infection. The rate of expression loss of RASSF1A was 20％(6/30) in gastric biopsy specimens of high_risk group and patients with possible carcinoma. However, RASSF1B and RASSF1C mRNA were fully expressed in both patient groups. The expression of RASSF1B and RASSF1C mRNA showed no statistical relation with gastric carcinoma clinical pathology indexes. CONCLUSION: RASSF1A gene is related to tumor suppression in gastric carcinoma, and correlated with the clinical pathological stage and the histological differentiation. The expression of RASSF1A was lost in gastroc biopsy specimens of high_risk group and patients with possible carcinoma. So,it could be a possible marker of early gastric carcinoma.There was no relationship between RASSF1A expression and H.pylori infection. Both are thus probably independent etiological factors.

BACKGROUND AND AIM: To study the antitumor effect of colon26 cells transfected with 4_1BBL gene in vivo. MATERIALS AND METHODS: The eukaryotic expression vector pMKIT/4_1BBL was transfected into colon26 cells with lipofectamine, The transfected cells were selected by G418 and termed colon26/4_1BBL. Then the tumor cell vaccine(TCV) was obtained by treatment with MMC. Three mice models (colon26, colon26/pMKITneo , and colon26/4_1BBL ) were established to study the antitumor effects of TCV_4_1BBL. The cytolytic activity of CTL and NK were detected by variable MTT method. The content of IL_2 and IFN_γ in serum were measured by ELISA . The early cancer model was made by inoculating colon26, then the mice were treated with TCV_4_1BBL on the second and seven days. RESULTS: Compared with colon26 and colon26/pMKITneo, colon26/4_1BBL cells grew much slower in mice. Mostly syngeneic mice were protected by inoculation with TCV_4_1BBL, survived free from tumor for a long period (over 100 days). The cytolytic effect of CTL and NK in colon26/4_1BBL mice were improved significantly, and the contents of IL_2 and IFN_γ were also increased. Cured by TCV_4_1BBL,some mice could survive free from tumor,and other bearing_tumor mice whose tumor grew slowly.The survival periods of all these mice were significantly prolonged. CONCLUSION: The antitumor effect against syngeneic marine colon carcinoma and the immune response in vivo were obviously enhanced by treating them with TCV_4_1BBL.

BACKGROUND AND AIM: A new approach to kill tumor cells in the blood through investigating apoptosis of K562 cells induced by high intensity pulsed electric fields. MATERIALS AND METHODS: Apoptosis of K562 cells was evaluated by transmission electron microscope and Annexin V_FITC and PI double labeled flow cytometry. RESULTS: There were many vacuoles and cracks in the cells, with heterochromatin margination, nuclear pycnosis and nuclear fragmentation. The high intensity pulsed electric fields caused damage in K562 cells, and the apoptotic percentage went up with the increase in frequency and intensity of PEF. CONCLUSION: K562 cell apoptosis was induced by high intensity pulsed electric fields,providing a new approach to kill tumors in the blood.

BACKGROUND AND AIM: To investigate polymorphisms of CYP3A7 gene of population in Guangxi with heavy aflatoxin B1 (AFB1)contamination, and the relationship between CYP3A7 genotypes and susceptibility of hepatocellular carcinoma (HCC). MATERIALS AND METHODS: Single nucleotide polymorphisms of CYP3A71A and 1C were analyzed in 210 cases and control samples (97 HCC, 38 HBV_positive but with no other disease and 75 HBV_ negative no disease) using the polymerase chain reaction_restriction fragment length polymorphism assays. RESULTS: All of the samples were CYP3A71A/1A genotype ,CYP3A71C allele was not detected in this cohort of Guangxi population. CONCLUSION: CYP3A71C allele was not detected in this cohort,suggesting that CYP3A7 might have a small influence on the development of AFB1_related HCC.

BACKGROUND AND AIM: To evaluate the protective effects of Huangqi on the damage of high glucose on vascular endothelial cell(VEC_304)in vitro． MATERIALS AND METHODS: The model of human umbilical vein endothelial cells (VEC_304) damage was induced by 50％ glucose for 4 h.The cells were pretreated with three concentrations of Huangqi solution for 24 h,before they were exposed to glucose. MTT and protein levels in cells were determined after 4 h. The expression of VEGF was investigated by RT_PCR and immunohistochemistry methods. RESULTS: MTT and protein levels in cells of the normal control group amd Huangqi_treated groups were higher than in glucose_damaged group without huangqi(P<0.05). The expression of VEGF was inhibited by glucose. CONCLUSION: Huangqi could protect the damages of VEC_304 endothelial cells induced by high glucose.

BACKGROUND AND AIM: The study was undertaken to assess the oxidative stress and DNA damage in human amnion cells induced by TBT. MATERIALS AND METHODS: Cultured FL cells were exposed to different concentrations of TBT (0, 2, 4, 6, 8, 10 μmol/L) for different durations (2 h and 4 h). Cell proliferation rate was evaluated by MTT assay. After exposure to different concentrations of TBT (0,1,2,3,4 μmol/L) for 2 h,the ROS level and DNA damage of FL cells were measured by DCFH_DA method and the single cell gel electrophoresis method, respectively. RESULTS: After exposure to different concentrations of TBT for 4 h, the FL cell proliferation rates in the 2, 8, 10 μmol/L groups were significantly decreased as compared to control(P<0.05, P<0.01, P<0.001) in a dose_dependent manner. The ROS level was increased in the 3 and 4 μmol/L groups in the dose_dependent manner and the difference was significant at 4 μmol/L group compared to the control(P<0.05). Meanwhile, TBT exposure led to the interrelated increase of nucleus tail length and tail moment in a dose－dependent manner and the differences were significant at 2, 3 and 4 μmol/L group compared to the control(P<0.05). CONCLUSION: TBT exposure could cause oxidative stress and DNA damage to FL cells.

BACKGROUND AND AIM: We evaluated the expression of CXCR4 and HER2, the inhibition of Herceptin on CXCR4 expression and the in vitro metastatic action in breast cancer cells. MATERIALS AND METHODS: Biomarker expression levels in paraffin_embedded tissue sections of breast cancer were evaluated using immunohistochemical staining. The protein expression of CXCR4 was studied by Western blot and the mRNA expression was by RT_PCR after treatment with Herceptin . Adhesion and chemotaxis assays were used to evalvate the effect of Herceptin on breast cancer cells with different HER2 expressions. RESULTS: Cytoplasmic CXCR4 was positively correlated with lymph node－positive tumors(P=0.032)and different stage of breast cancer(P=0.000)and the expression of HER2(P=0.015). The protein and mRNA expressions of CXCR4 were decreased after treatment with Herceptin in breast cancer cells with HER2 overexpression(P<0.05)and activity of cell adherence to fibronectin(FN) and migration to SDF_1α were inhibited. CONCLUSION: HER2_mediated homing to metastatic organs and upregulation of CXCR4 may be key factors for HER2_mediated breast cancer metastasis.

BACKGROUND AND AIM: To detect the expression of receptor tyrosine kinase EphB2 as well as its relationship with angiogenesis in colon carcinoma，and explore the possible mechanism. MATERIALS AND METHODS: The expressions of EphB2 in 10 normal tissues and 43 colon carcinoma were detected by immunohistochemisty, and stromal microvessel density (MVD) was counted according to the results of CD34 detectedby immunohistochemisty. RESULTS:The majority of colon carcinomas expressed EphB2 in the cytoplasm. There was positive correlation between EphB2 expression in the cytoplasm and tumor differentiation，but negative correlation between EphB2 and tumor infiltration depth. Expression of EphB2 was positive correlation with MVD in immunoreation. CONCLUSION: EphB2 may play suppressive role in carcinogenesis and progression of colon carcinoma.EphB2 may achieve its tumor suppressor through regulation of tumor angiogenesis.

BACKGROUND AND AIM: To study the relationship between the expression of Survivin and PTEN in skin scar and scar carcinoma . MATERIALS AND METHODS:The expressions of Survivin and PTEN in 10 skin scar carcinoma, 10 skin scar and 10 normal skin tissues were evaluated by the immunohistochemical method of S_P.Pictures were collected and analyzed, statistical analysis was performed by SPSS system. RESULTS:The expressions of Survivin in skin scar carcinoma and scar tissues were significantly higher than normal skin(P<0.01).However,no statistical difference was found between skin scar carcinoma and scar tissues(P>0.05).The expressions of PTEN in skin scar carcinoma and scar tissues were significantly lower than normal skin(P<0.01),and with no statistical difference between scar carcinoma and scar tissues(P>0.05). CONCLUSION: The abnormal expressions of Survivin and PTEN in skin scar and scar carcinoma tissues were correlated with malignant transformation of skin scar.

BACKGROUND AND AIM: Surgery is the main choice in treating early cervix cancer, and preoperative radiotherapy or chemotherapy can increase surgical resection rate.We analyzed the postoperative complications of radical hysterectomy, and further explored its prevention and control measures. MATERIALS AND METHODS: Data of 247 patients with cervical cancer confirmed by pathology were collected from the Affiliated Tumor Hospital, Shantou University Medical College in China, during Jan. 2000 to Dec. 2006. All patients were treated by radical hysterectomy. 130 patients were treated by preoperative brachytherapy, among them 52 patients received a course of preoperative chemotherapy simultaneously. RESULTS: The major complications of radical hysterectomy were urinary retention, lymphocele, and urinary tract infection with rates of 24.7％, 7.69％ and 8.50％, respectively. The incidences of three complications in the 130 patients treated with preoperative radiotherapy or chemotherapy compared to the 117 patients treated only by radical hysterectomy were 23.1％/26.5％，6.93％/8.55％ and 10.77％/5.99％, respectively, without significant differences(P> 0.05). The amount of average intra_operative hemorrhage in former group was higher than that of latter group [(490.74±47.67) ml VS (438±61.38) ml], but the difference between two groups was not statistically significant (P<0.05). CONCLUSION: The main postoperative complications were urinary retention, lymphocele and urinary tract infection. The preoperative radiotherapy or chemotherapy did not increase the complications and the amount of intra_operative bleeding.

BACKGROUND AND AIM: To obtain highly purified cultured Sertoli cells from rat testis. MATERIALS AND METHODS: Testes from 18－22 day old SD rats were removed and decapsulated，then chopped and sequentially digested with two enzymes at 37 ℃：first with 0.25％ trypsin for 20－40 minutes，then with 0.05％ collagenase V for 40－60 minutes．The speed of centrifugation was 800 r/min. The isolated cells were incubated at 35 ℃ in a humidified atmosphere of 5％ CO2．To increase the purity of Sertoli cells，cultured cells were subjected to hypotonic shock(treatment)with 20 mmol Tris_HCl after 48 h of incubation．After 0.25％ trypsin digestion and adherent culture, the cultured cells were identified by HE staining，oil red O staining and transmission electron microscope. The growth curve of Sertoli cells was evaluated as well. RESULTS: Over 95％ of the cultured cells were Sertoli cells．The exponential phase of growth was 4－9 days. CONCLUSION: Highly purified Sertoli cells could be obtained by two step enzymes digestion and hypotonic shock, and identification of Sertoli cells by oil red O was a convenient method.

BACKGROUND AND AIM: To assess the protective effects of β_carotene(βC) on DNA damage induced by mitomycin_C (MMC) in bone marrow cells of mice. MATERIALS AND METHODS: Two batches of mice were primed orally with βC in concentrations of 3 and 30 mg/kg for 8 days and then followed by MMC intraperitoneal injection at dose of 2 mg/kg on day 9. DNA and chromosome damage were examined by single cell gel electrophoresis(SCGE) and chromosome aberrations analysis. RESULTS: The comet frequency, tail length and the chromosomal aberration rate of two doses of βC were significantly lower than those of the MMC group. CONCLUSION: To some extent,βC could protect the bone marrow cells of mice from damage induced by MMC in vivo.

BACKGROUND AND AIM:To study the teratogenic toxicities of carthamus in SD rats during the period of organ formation． MATERIALS AND METHODS: Eighty_two pregnant SD rats were randomly divided into four groups with about 20 rats in each, three carthamus dosage groups (7.5,15.0,30.0 mg/kg),and one negative control group. carthamus or normal saline was given via vena caudalis injection for 10 days during the period of organ formation (the 6th to 15th day of gestation). The rats were sacrificed to examine the fetuses on the 20th day. RESULTS: Carthamus didn't produce significant difference on living fetuses ratio, absorbed fetuses ratio, dead fetus ratio, body length, tail length and weight of fetuses .There were statistical differences on average weight of placenta ,number of caudal vertebra and distant digital bone between treated groups and the negative control group. It didn' t induce any teratogenic effect on the appearance, bones and bowels of the fetuses when used under 30.0 mg/kg. CONCLUSION: Carthamus had no maternal toxicity, teratogenic toxicity, embryotoxicity and fetotoxicity when used under 30.0 mg/kg.