Inhibitory effects of tanshinone II-A on invasion and metastasis of human colon carcinoma cells.

AIM: To investigate the effects and possible mechanisms of tanshinone II-A, an alcohol extract of the root of Salvia miltiorrhiza Bunge, on tumor invasion and metastasis of human colon carcinoma (CRC) cells. METHODS: The effects of tanshinone II-A on invasion and metastasis of CRC cell lines HT29 and SW480 were evaluated by in vitro and in vivo assays. Western blotting was used to investigate possible molecular mechanisms of tanshinone II-A anti-cancer actions. RESULTS: Tanshinone II-A inhibited migration and invasion of CRC cells in a dose-dependent manner. The inhibitory effect also depended on time, with the most significant effects observed at 72 h. Tanshinone II-A also significantly inhibited in vivo metastasis of colon carcinoma SW480 cells. It inhibited in vitro and in vivo invasion and metastasis of CRC cells by reducing levels of urokinase plasminogen activator (uPA) and matrix metalloproteinases (MMP)-2 and MMP-9, and by increasing levels of tissue inhibitor of matrix metalloproteinase protein (TIMP)-1 and TIMP-2. Tanshinone II-A was also shown to suppress the nuclear factor-kappaB (NF-kappaB) signal. CONCLUSION: Tanshinone II-A inhibited in vitro and in vivo invasion and metastasis of CRC cells. The effect resulted from changes in the levels of uPA, MMP-2, MMP-9, TIMP-1 and TIMP-2, and apparent inhibition of the NF-kappaB signal transduction pathway.

[Quantitative detection of DD3 mRNA in prostate cancer tissues by real-time fluorescent quantitative reverse transcription polymerase chain reaction].

OBJECTIVE: To analyze the expression of DD3 mRNA in the prostate tissues. METHODS: DD3 mRNA was detected by realtime fluorescent quantitative reverse transcription polymerase chain reaction (FQ-RT-PCR) based on the Taqman technique in the tissues of 27 patients with non-prostate cancer( NPCa), 21 prostate cancer( PCa), 39 benign prostatic hyperplasia (BPH) and 15 normal prostate (NP). The ROC curve was used to evaluate the diagnostic value of DD3 mRNA. RESULTS: DD3 mRNA expression was not detected in the NPCa tissues. The median expressions of DD3 mRNA in PCa, BPH and NP tissues were 7. 2 x 10(6), 2. 5 x 10(4) and 1.5 x 10(4) copies/mg tissue, respectively. The DD3 mRNA expression levels were significantly different between nonmalignant and malignant tissues (P < 0.01). No significant differences in DD3 mRNA expression were detected between the NP and BPH tissues and no significant correlation was found between the DD3 mRNA expression and clinical pathological parameters. The AUC-ROC was 0.937 (95% CI: 0.879 - 0.995) at cutoff value 1.4 x 10(5) copies/mg tissue. The sensitivity, specificity, accuracy, positive predictive value, negative predictive value, positive likelihood ratio and negative likelihood ratio for DD3 were 90.5%, 85.0%, 86.7%, 76.0%, 94.3%, 6.03 and 0.11 respectively. CONCLUSION: The DD3 mRNA expression is confined to prostate tissues and highly upregulated in PCa tissues. It has a potential application value in the early diagnosis of prostate cancer and the follow-up of the patient.

Effects of nuclear factor-kappaB on rat hepatocyte regeneration and apoptosis after 70% portal branch ligation.

AIM: To detect the DNA binding activity of nuclear factor-kappaB (NF-kappaB) in rat hepatocyte and to investigate the effects of NF-kappaB on rat hepatocyte regeneration and apoptosis after 70% portal branch ligation. METHODS: Sixty Wistar rats were randomly divided into control group and portal branch ligation group. The animals were killed 12 h, 1, 2, 3, 7, and 14 d after surgery to determine the contents of plasma ALT. Hepatocytes were isolated and nuclear protein was extracted. DNA binding activity of NF-kappaB was measured by EMSA. Hepatocyte regeneration and apoptosis were observed under microscope by TUNEL staining. The ultrastructural changes of liver were observed under electron microscope. RESULTS: Seventy percent portal branch ligation produced atrophy of the ligated lobes and the perfused lobes underwent compensatory regeneration, the total liver weight and plasma ALT levels were maintained at the level of sham-operated animals throughout the experiment. After 2 d of portal branch ligation, DNA binding activity of NF-kappaB in hepatocyte increased and reached its peak, the number of apoptotic hepatocyte in the ligated lobes and the number of mitotic hepatocyte in the perfused lobes also reached their peak. Typical apoptotic changes and evident fibrotic changes in the ligated lobes were observed under electron microscope. CONCLUSION: After 70% portal branch ligation, DNA binding activity of NF-kappaB in hepatocyte is significantly increased and NF-kappaB plays an important role in hepatocyte regeneration and apoptosis.

MicroRNA-155 promotes the proliferation and invasion abilities of colon cancer cells by targeting quaking.

The increasing expression of microRNA­155 (miR­155) and decreasing expression of RNA­binding protein quaking (QKI) in colon cells have been observed previously. In this study, we attempted to establish the correlation between miR­155 and QKI. In addition, we assessed whether the expression of miR­155 and QKI is linked to the proliferation and invasion capabilities of colon cells. Firstly, nineteen tumor samples, divided into two groups according to the presence or absence of lymphatic metastasis, were obtained from colon cancer patients at the First Affiliated Hospital of Wenzhou Medical University, China. The expression level of miR­155 and QKI was measured by quantitative polymerase chain reaction (qPCR). Secondly, the GES­1, SW480 and COLO205 cell lines were cultured and the expression level of QKI and miR­155 was also assessed by qPCR. Thirdly, a luciferase reporter gene assay was performed to detect the association between miR­155 and QKI, and qPCR and western blot analysis were performed to confirm the effects of miR­155 on the expression of QKI at the mRNA and protein level. Subsequently, the SW480 cells were used in the following experiments. Following treatment with miR­155 inhibitor and QKI overexpression vector, western blot analysis, propidium iodide (PI) staining and a cell scratch assay were carried out to assess the effects of miR­155 on the proliferation and invasion potential of colon cancer cells. qPCR findings revealed higher miR­155 expression and lower QKI expression in colon cancer tissues as well as the colon cancer cell lines SW480 and COLO205. The relative luciferase activity of the 3' untranslated region (3'UTR) was decreased by approximately 45% when SW480 cells stimulated by mimic­miR­155 were combined with the wild­type 3'UTR constructs. In addition, when the cells were treated with mimic­miR­155, QKI expression was significantly decreased at the mRNA and protein level. These outcomes revealed that miR­155 decreased the production of QKI by acting on the 3'UTR of the QKI gene. Furthermore, PI staining and the cell scratch assay revealed that miR­155 influenced the cell cycle and invasion abilities of colon cancer cells by directly targeting QKI and decreased the production of QKI by acting on the 3'UTR of the QKI gene. This study has demonstrated the correlation between miR­155 and QKI, in which miR­155 regulates the cell cycle and invasion ability of colon cancer cells via the modulation of QKI expression. Our study provides novel therapeutic strategies for colon cancer therapy.

Decreased expression of LKB1 correlates with poor prognosis in hepatocellular carcinoma patients undergoing hepatectomy.

AIM: To study any correlation of LKB1 expression with prognosis in hepatocellular carcinoma (HCC) cases. METHODS: A total of 70 HCC patients and 20 primary intrahepatic stone patients in the first affiliated hospital of Wenzhou Medical College were enrolled in this study. LKB1 expression was detected by immunohistochemistry. Patients were followed-up and prognostic factors were evaluated. RESULT: LKB1 expression was decreased in the HCC samples. Loss of LKB1 expression in HCC was significantly related to histologic grade (P=0.010), vascular invasion (P=0.025) and TMN stage (P=0.011). Patients showing negative LKB1 expression had a significantly shorter disease-free and overall survival than those with positive expression (P = 0.001, P=0.000, respectively). Multivariate Cox regression analysis indicated that LKB1 expression level was an independent factor of survival (P = 0.033). CONCLUSION: HCC patients with decreased expression LKB1 have a poor prognosis. The loss of LKB1 expression is correlated with a lower survival rate.

Angiogenesis and clinicopathologic characteristics in different hepatocellular carcinoma subtypes defined by EpCAM and α-fetoprotein expression status.

Recently, two hepatic lineage markers epithelial cell adhesion molecule (EpCAM) and α-fetoprotein (AFP) were used to classify hepatocellular carcinoma (HCC) into four subtypes with prognostic implication. In the present study, we further evaluated the clinicopathologic and angiogenic characteristics among these HCC subtypes. EpCAM expression was investigated by immunohistochemistry in 115 HCC primary tumors. Based on EpCAM immunostaining and serum AFP levels, 115 HCC cases were classified into four subtypes: EpCAM+AFP+ (26.1%), EpCAM-AFP+ (20.0%), EpCAM+AFP- (20.8%), and EpCAM-AFP- (33.1%). EpCAM+AFP+ and EpCAM-AFP+ HCC were associated with late TNM stages and high frequencies of venous invasion, whereas EpCAM+AFP- and EpCAM-AFP- subtypes were associated with early TNM stages and low frequencies of venous invasion. Furthermore, EpCAM+AFP+ HCC had a significantly higher microvessel density (MVD) and higher level of VEGF (Vascular epithelial growth factor) expression than the other three subtypes. In conclusion, our study indicated that subtype classification of HCC based on EpCAM and AFP status had clinicopathologic and biologic implications in aggressive phenotype and angiogenesis. We also suggest that the EpCAM+AFP+ HCC patients might be potential therapeutic candidates for anti-angiogenesis therapy.