Xia Xu

Combination of curcumin and green tea catechins prevents dimethylhydrazine-induced colon carcinogenesis

The chemopreventive effects of curcumin and green tea catechins individually and in combination on 1,2-dimethylhydrazine (DMH)-induced colon carcinogenesis were studied in male Wister rats following 32 weeks of dietary treatment. The incidence, number and size of colorectal cancer were measured. Colorectal aberrant crypt foci (ACF) were analyzed by methylene blue staining. Proliferation indices and apoptotic indices were determined by PCNA immunostaining and TUNEL assay, respectively. The results showed that dietary curcumin, catechins and combination administration significantly inhibited the total number of ACF per rat. The combination treatment displayed the most potent inhibitory effect, while there was no difference of inhibition between curcumin and catechins-treated groups. The incidence of colorectal cancer in the treated groups was significantly lower than that of positive control group. Compared with the positive control group, the proliferation index was significantly decreased and the apoptotic index was significantly increased in all treatment groups, while the effect of the combination was the greatest among the treated groups. Our findings suggest that the combination of curcumin and catechins may produce a synergistic colon cancer-preventative effect that would be more potent than each of the compounds alone.

Preventive action of curcumin in experimental acute pancreatitis in mouse.

Background & objectives: Curcuma longa (turmeric) has a long history of use in Ayurvedic medicine as a treatment for inflammatory conditions. The purpose of the present study was to investigate the preventive effects of curcumin against acute pancreatitis (AP) induced by caerulein in mouse and to elucidate possible mechanism of curcumin action. Methods: Curcumin (50 mg/kg/day) was intraperitoneally injected to Kun Ming male mice for 6 days, followed by injection of caerulein to induce AP. GW9662 (0.3 mg/kg), a specific peroxisome proliferator-activated receptor gamma (PPARγ) antagonist, was intravenously injected along with curcumin. Murine macrophage RAW264.7 cells were treated with 100 μmol/l curcumin for 2 h, and then stimulated with 0.1 μ g/ml lipopolysaccharide (LPS). Serum amylase and transaminase levels were measured at 10 h after AP. TNF-α level in mouse serum and cell culture medium were detected by ELISA. Expression of PPARγ and NF-κB were analyzed by RT-PCR and Western blot. Results: Curcumin significantly decreased the pancreas injury and reversed the elevation of serum amylase, ALT and AST activities and TNF-α level in mice with AP. Curcumin treatment inhibited the elevation of NF-κB-p65 in the nucleus of mouse pancreas AP group and RAW264.7 cells, but significantly increased the expression of PPARγ. GW9662 could abolish the effects of curcumin on serum levels of amylase, ALT, AST, TNF-α, and NF-κB level. Interpretation & conclusions: Our results suggest that curcumin could attenuate pancreas tissue and other organ injury by inhibiting the release of inflammatory cytokine TNF-α. These effects may involve upregulation of PPARγ and subsequent downregulation of NF-κB.

Oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line

Oridonin, a diterpenoid isolated from Rabdosia rubescencs, has been reported to have antitumor effects. However, low solubility has limited its clinical applications. Preparation of drugs in the form of nanosuspensions is an extensively utilized protocol. In this study, we investigated the anticancer activity of oridonin and oridonin nanosuspension on human pancreatic carcinoma PANC-1 cells. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed to investigate the effect of oridonin on cell growth. Propidium iodide and Hoechst 33342 staining were used to detect morphologic changes. The percentage of apoptosis and cell cycle progression was determined by flow cytometric method staining with propidium iodide. Annexin V-fluorescein isothiocyanate (FITC)/PI staining was used to evaluate cell apoptosis by flow cytometry. Caspase-3 activity was measured by spectrophotometry. The apoptotic and cell cycle protein expression were determined by Western blot analysis. Both oridonin and oridonin nanosuspension induced apoptosis and G2/M phase cell cycle arrest, and the latter had a more significant cytotoxic effect. The ratio of Bcl-2/Bax protein expression was decreased and caspase- 3 activity was stimulated. The expression of cyclin B1 and p-cdc2 (T161) was suppressed. Our results showed that oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line.

miRNA‐532‐5p functions as an oncogenic microRNA in human gastric cancer by directly targeting RUNX3

Accumulating data reveal that microRNAs are involved in gastric carcinogenesis. To date, no information was reported about the function and regulatory mechanism of miR‐532‐5p in human gastric cancer (GC). Thus, our study aims to determine the role and regulation of miR‐532‐5p in GC. Here, we found that transient and stable overexpression of miR‐532‐5p dramatically increased the potential of colony formation and migration of GC cells, decreased the percentage of cells in G1 phase and cell apoptosis in vitro, and increased the weight of mice lungs and number of lung xenografts in vivo. Gain‐of‐function, loss‐of‐function and luciferase activity assays demonstrated that miR‐532‐5p negatively regulated the expression of RUNX3 and its targets directly. We also found that miR‐532‐5p level was negatively correlated with RUNX3 gene expression in various GC cell lines. Our results indicate that miR‐532‐5p functions as an oncogenic miRNA by promoting cell growth, migration and invasion in human GC cells.

RUNX3 inhibits survivin expression and induces cell apoptosis in gastric cancer

Transcription factor RUNX3 is associated with gastric tumorigenesis and progression through regulating the expression of its target genes. Survivin is a member of the inhibitor of apoptosis (IAP) family and has been shown to inhibit cell apoptosis and promote cell proliferation. Increased survivin expression has been found in various cancer types, including gastric cancer. In this study, we found that restoration of RUNX3 promotes cell apoptosis through inhibiting the survivin expression, while RUNX3 inhibition increases the expression of survivin in gastric cancer cell lines. Moreover, RUNX3 over-expression inhibits,whereas its inhibition increases, the promoter activity of survivin gene, respectively. RUNX3-R122C, a mutation located in the conserved Runt domain, has no effect on the promoter activity of survivin gene. We further identified a RUNX3-binding site in the promoter of survivin gene and the binding of RUNX3 on survivin promoter leads to significantly inhibition of survivin expression. Finally, we confirmed the negative correlation of RUNX3 and survivin expression in clinical specimens of gastric cancer. These findings reveal a novel mechanism of RUNX3 for the induction of cell apoptosis in human gastric cancer.

Metformin prevents DMH-induced colorectal cancer in diabetic rats by reversing the warburg effect.

Epidemiologic studies have shown that the treatment of diabetics with metformin reduced the risk of cancer‐related mortality. Here, we investigated the chemopreventive effects of metformin on dimethylhydrazine (DMH)‐induced colorectal carcinogenesis in diabetic SD rats following metformin treatment and the effect on Warburg effect involved in this process. Diabetic rat models were induced with high‐fat feeding in combination with a low dose of Streptozotocin (STZ) and then induce colorectal cancer with a low dose of DMH. The formation of colorectal Aberrant crypt foci (ACF) and the incidence, number and size of the tumor were measured. The proliferation indices of colonic tissues were determined through Proliferating cell nuclear antigen (PCNA) immunostaining. Then detect the expression of PK and IDH in colonic tissues using immunohistochemistry and Western blot. The enzyme activities of HK and PDH in colonic tissues were measured. The growth and expression of PK and IDH and activity of HK and PDH in cell lines LoVo and HT‐29 were measured after metformin treatment. The results showed that metformin treatment significantly inhibited the formation of ACF and tumors. The proliferation index of colonic tissues was significantly decreased following metformin treatment. In addition, metformin inhibited cell growth and decreased the imbalance in the expression of the enzymes involved in glycolysis and the TCA cycle. These findings suggested that metformin might produce a synergistic colon cancer‐preventative effect in diabetic patients through the regulation of the enzymes expression involved in glucose metabolism.

RUNX3-mediated up-regulation of miR-29b suppresses the proliferation and migration of gastric cancer cells by targeting KDM2A.

RUNX3 is a transcriptional factor that has been shown to regulate protein-coding gene expression at the transcriptional level. However, the regulation of RUNX3 on miRNAs is not fully understood. In this study, we used miRNA microarray to identify the miRNAs that are regulated by RUNX3 and found that miR-29b showed the most up-regulation in RUNX3 over-expressed cells compared with the control cells. We used qRT-PCR to confirm the miRNA microarray results in several gastric cancer cells and found that RUNX3 could bind to the miR-29b promoter directly and cooperate with Smad3 to increase the promoter activity of miR-29b. In the clinical setting, both RUNX3 and miR-29b are down-regulated significantly in human gastric cancer tissues. A positive correlation between miR-29b and RUNX3 was found in the gastric cancer tissues. Additionally, we found that miR-29b suppressed the proliferation and metastasis of gastric cancer cells by directly targeting KDM2A. The miR-29b/KDM2A axis was involved in the RUNX3-mediated inhibition of gastric cancer cell proliferation and metastasis. Taken together, our results suggested that RUNX3-mediated up-regulation of miR-29b inhibited the proliferation and migration of gastric cancer cells by targeting KDM2A, representing a novel molecular mechanism for the tumor suppression action of RUNX3.

Regulation of SOD2 and β-arrestin1 by interleukin-6 contributes to the increase of IGF-1R expression in docetaxel resistant prostate cancer cells

Although several mechanisms behind resistance to docetaxel in castration-refractory prostate cancer (CRPC) have been investigated, molecular determinants of evolved resistance are still not entirely understood. Proteomics-based analysis in this study revealed that SOD2, associated with downregulation of reactive oxygen species (ROS), was significantly up-regulated in docetaxel-resistant (PC3/Doc) cells if compared to sensitive cells, and the expression of redox-regulated genes such as IGF-1R, CXCR4, and BCL2 was increased as well. Forced expression of SOD2 in sensitive cells led to the increase of IGF-1R and association with drug resistance, whereas silencing of SOD2 resulted in the decrease of IGF-1R at the protein level in resistant cells. Further study revealed that SOD2 acted as a negative regulator of β-arrestin1 that is an important adaptor responsible for degradation of IGF-1R via the changes in ROS, as evidenced by observations that an antioxidant agent substantially attenuated β-arrestin1 expression in vitro and in vivo. Finally, we found that blocking of IL6 that was up-regulated in resistant cells resulted in attenuation of SOD2 and STAT3, and simultaneously in increased expression of β-arrestin1. The modulation consequently led to the decreased IGF-1R at both protein and transcription levels. Together, our data provide a novel explanation that high level of IL6 stimulated SOD2 expression that, at least partially, contributed to the low level of ROS that would likely result in a sustained increase in the expression of IGF-1R through abolishment of β-arrestin1 in docetaxel resistant cells.

Associations of saposin C, Src, and androgen receptor upregulate the expression and function of androgen receptor in human prostate cancer cells

We previously demonstrated that ectopic expression of neurotrophic peptide (NP) derived from saposin C promotes androgen receptor (AR) expression and transactivation in human prostate cancer cells. This prompted us to investigate how NP or saposin C can function in cells. We constructed plasmids expressing saposin C or a chimeric peptide of a viral TAT transduction domain and saposin C (TAT‐saposin C) with His‐tag. Intracellular localization of saposin C and NP was predominantly shown in transfected cells, while TAT‐saposin C was detected around membrane and in cytosol by immunofluorescence staining. Furthermore, induction of the AR expression and activation of the AR transcriptional function were observed in cells transfected with saposin C or TAT‐saposin C, compared to control cells transfected with an empty plasmid. The effects of saposin C and TAT‐saposin C on AR activity were examined in the presence of inhibitors of GPCR, MAPK1/2, and PI3K/Akt. Interestingly, we found that these inhibitors only affect AR activities in cells with TAT‐saposin C expression but not with saposin C expression. Immunostaining images showed that co‐localization of saposin C, Src, and the AR occurred in transfected cells. Physical interactions of saposin C/NP, Src, and the AR were then demonstrated by co‐immunoprecipitation assays. Blockage of Src activity by specific inhibitor led to a decrease in the saposin C‐mediated enhancement of AR transactivity, suggesting that intracellular expression of saposin C caused stimulation of AR expression and activity by associations with Src in LNCaP cells. This effect may not be mediated by GPCR. J. Cell. Biochem. 112: 818–828, 2011.

Oxidative stress induced autophagy in cancer associated fibroblast enhances proliferation and metabolism of colorectal cancer cells

ABSTRACT Tumors are comprised of malignant cancer cells and stromal cells which constitute the tumor microenvironment (TME). Previous studies have shown that cancer associated fibroblast (CAF) in TME is an important promoter of tumor initiation and progression. However, the underlying molecular mechanisms by which CAFs influence the growth of colorectal cancer cells (CRCs) have not been clearly elucidated. In this study, by using a non-contact co-culture system between human colorectal fibroblasts (CCD-18-co) and CRCs (LoVo, SW480, and SW620), we found that fibroblasts existing in tumor microenvironment positively influenced the metabolism of colorectal cancer cells, through its autophagy and oxidative stress pathway which were initially induced by neighboring tumor cells. Therefore, our data provided a novel possibility to develop fibroblasts as a potential target to treat CRC.