Hai-Tao Guan

Antitumor activity of the selective cyclooxygenase-2 inhibitor, celecoxib, on breast cancer in Vitro and in Vivo

BackgroundCyclooxygenase-2(COX-2) promotes carcinogenesis, tumor proliferation, angiogenesis, prevention of apoptosis, and immunosuppression. Meanwhile, COX-2 over-expression has been associated with tumor behavior and prognosis in several cancers. This study investigated the antitumor effects of the selective COX-2 inhibitor, Celecoxib, on breast cancer in vitro and in vivo.MethodsHuman breast cancer MCF-7 and MDA-MB-231 cells were cultured with different concentration (10, 20, 40 μmol/L) of celecoxib after 0-96 hours in vitro. MTT assay was used to determine the growth inhibition of breast cancer cells in vitro. The expression of COX-2 on mRNA was measured by real-time quantitive PCR analysis. Flow cytometry was performed to analyze the cell cycle of MCF-7 cells. Levels of PGE2 were measured by ELISA method. The in vivo therapeutic effects of celecoxib were determined using rat breast cancer chemically induced by 7,12-dimethylben anthracene (DMBA).ResultsThe inhibition of proliferation of both MCF-7 and MDA-MB-231 cells in vitro by celecoxib was observerd in time and dose dependent manner. Celecoxib effectively down-regulated the expression of COX-2. The cell cycle was arrested at G0/G1, and rate of cells in S phase was obviously decreased. Levels of PGE2 were inhibited by Celecoxib. The tumor incidence rate of the celecoxib group was lower than that of the control group. In addition, the tumor latency period of the celecoxib group was longer than that of the control group.ConclusionsCelecoxib inhibited the proliferation of breast cancer cell lines in vitro, and prevented the occurrence of rat breast cancer chemically induced by DMBA. Therefore, celecoxib exhibits an antitumor activity and seems to be effective in anti-tumor therapy.

Up-regulation of hypoxia inducible factor-1α by cobalt chloride correlates with proliferation and apoptosis in PC-2 cells

BackgroundThe exact mechanism of the effects of hypoxia on the proliferation and apoptosis in carcinoma cells is still conflicting. This study investigated the variation of hypoxia-inducible factor-1α(HIF-1α) expression and the apoptosis effect of hypoxia stimulated by cobalt chloride (CoCl2) in pancreatic cancer PC-2 cells.MethodsPC-2 cells were cultured with different concentration (50-200 μmol/L) of CoCl2 after 24-120 hours to simulate hypoxia in vitro. The proliferation of PC-2 cells was examined by MTT assay. The cellular morphology of PC-2 cells were observed by light inverted microscope and transmission electron microscope(EM). The expression of HIF-1α on mRNA and protein level was measured by semi-quantitive RT-PCR and Western blot analysis. Apoptosis of PC-2 cells were demonstrated by flow cytometry with Annexin V-FITC/PI double staining.ResultsMTT assay showed that the proliferation of PC-2 cells were stimulated in the first 72 h, while after treated over 72 h, a dose- dependent inhibition of cell growth could be observed. By using transmission electron microscope, swollen chondrosomes, accumulated chromatin under the nuclear membrane and apoptosis bodies were observed. Flow cytometer(FCM) analysis showed the apoptosis rate was correlated with the dosage of CoCl2. RT-PCR and Western blot analysis indicated that hypoxia could up-regulate the expression of HIF-1α on both mRNA and protein levels.ConclusionHypoxic microenvironment stimulated by CoCl2 could effectively induce apoptosis and influence cell proliferation in PC-2 cells, the mechanism could be related to up-expression of HIF-1α.

Saikosaponin-D reduces cisplatin-induced nephrotoxicity by repressing ROS-mediated activation of MAPK and NF-κB signalling pathways.

The nephrotoxicity induced by cisplatin (DDP) severely limits the clinical efficacy of this widely used anticancer agent. The observed nephrotoxicity may be the result of DDP-induced inflammation and apoptosis. Saikosaponin-D (SSD), a triterpenoid saponin, has numerous pharmacological properties. The goal of the present study was to investigate whether and how SSD protected against DDP-induced nephrotoxicity. Non-cytotoxic levels of SSD significantly increased the viability rate, improved the nuclear morphology, and attenuated the caspase-3 activation and programmed apoptosis of DDP-treated HK-2 cells. In addition, SSD treatment markedly inhibited the release of tumour necrosis factor (TNF)-α, interleukin-1β (IL-1β), and interleukin-6 (IL-6), as well as the production of nitric oxide and the expression of inducible nitric oxide synthase (iNOS) by these cells. More importantly, SSD effectively blocked the DDP-induced activation of NF-κB, P38, JNK, and MAPKs. Furthermore, we found that U0126 (a specific inhibitor of MAPKs) strongly inhibited the IKK/IκB/NF-κB-dependent release of pro-inflammatory cytokines and iNOS gene expression. Finally, we demonstrated that SSD decreased the level of reactive oxygen species (ROS) accumulation and that the specific ROS scavenger N-acetylcysteine (NAC) markedly inhibited the DDP-induced activation of MAPK and phosphorylation of the downstream signal NF-κB, which in turn reduced the levels of pro-inflammatory cytokine release and iNOS gene expression. Our results suggest that the SSD-mediated alleviation of DDP-induced nephrotoxicity was due to uncoupling of the ROS, P38, and JNK/NF-κB signalling pathways.

In Vitro and In Vivo Antitumor Activity of Scutellaria barbate Extract on Murine Liver Cancer

In the present study, we investigated the in vitro and in vivo antitumor effects of crude extract of Scutellaria barbate (CE-SB) on mouse hepatoma H22 cells. The MTT assay was used to determine the growth inhibition of H22 cells in vitro. The in vivo therapeutic effects of CE-SB were determined using H22 tumor bearing mice. Besides, the body weight, tumor weight, thymus index and spleen index of H22 bearing mice were also measured. The tumor inhibitory rate (IR) was calculated according to the mean weight of tumor (MWT). The phagocytotic function of macrophages was examined by observing peritoneal macrophages phagocytize chicken RBC. The results showed that CE-SB could inhibit the growth of hepatoma H22 Cells in vitro and in vivo. Furthermore, CE-SB could improve immune function of H22 tumor bearing mice. Together these results indicate that CE-SB has antitumor activity and seems to be safe and effective for the use of anti-tumor therapy.

Saikosaponin-D Enhances Radiosensitivity of Hepatoma Cells under Hypoxic Conditions by Inhibiting Hypoxia-Inducible Factor-1α

Background: Our previous study revealed that the combination of Saikosaponin-d (SSd) and radiation is more effective in the treatment of liver cancer than the application of either of these monotherapeutic methods. However, the molecular mechanisms of the radiosensitizing effect of SSd on liver cancer remained ill defined. Methods: Cells were treated with different interventions; afterward, cell viability, apoptosis, and cell survival of SMMC-7721 and HepG2 hepatoma cells were examined. Xenograft tumor models were established by subcutaneously injecting SMMC-7721 cells. The molecular mechanism was assessed by western blot. Results: SSd dose-dependently increased radiosensitivity of hepatoma cells under hypoxic condition. The growth inhibitory effect of the combined treatment was correlated with cell apoptosis. Further mechanistic analysis indicated that SSd induced the upregulation of p53 and Bax as well as the downregulation of Bcl-2 by attenuating HIF-1α expression under hypoxic condition. These effects were enhanced when the HIF-1α inhibitor PX-478 was introduced. In vivo data also presented a more significant suppression of tumor xenograft growth from the combined therapy than from either of the monotherapeutic methods. Conclusions: Our study provides evidence for a radiosensitizing effect of SSd on hepatoma cells under hypoxic conditions by inhibiting HIF-1α expression. Thus, SSd can be used as a potential sensitizer in hepatoma radiotherapy.

Knockdown of SPOCK1 Inhibits the Proliferation and Invasion in Colorectal Cancer Cells by Suppressing the PI3K/Akt Pathway

Sparc/osteonectin, cwcv, and kazal-like domains proteoglycan (testican) 1 (SPOCK1), known as testican-1, were found to be involved in the development and progression of tumors. However, in colorectal cancer (CRC), the expression pattern of SPOCK1 and its functional role remain poorly investigated. In the present study, we explored the role of SPOCK1 in CRC. Our results demonstrated that SPOCK1 is overexpressed in CRC cell lines. SPOCK1 silencing significantly inhibited the proliferation in vitro and the tumor growth in vivo. Furthermore, SPOCK1 silencing significantly attenuated the migration/invasion by reversing the EMT process in CRC cells. Finally, knockdown of SPOCK1 obviously decreased the protein expression levels of p-PI3K and p-Akt in HCT116 cells. In total, our study demonstrated for the first time that knockdown of SPOCK1 inhibits the proliferation and invasion in CRC cells, possibly through the PI3K/Akt signaling pathway. Therefore, SPOCK1 may be a potential therapeutic target for the treatment of CRC.

Expression of Survivin, p53 and its relationship with apoptosis, proliferation in hepatocellular carcinoma（HCC）

Abstract Objective To investigate the expression of Survivin p53 and its relationship with apoptosis proliferation in hepatocellular carcinoma (HCC). Methods The expression of Survivin, p53 and the proliferation of tumor cells marked by proliferation cell nuclear antigen (PCNA) in 42 cases of HCC were assessed by immunohistochemical method. TUNEL was used to detect apoptosis. Results Survivin protein was expressed in 30 of 42 cases of HCC (71.4%) and in 4 of 34 cases of adjacent cirrhosis tissues (11.8%). Expression of Survivin protein was negative in 10 cases of normal tissues. Survivin protein positive expression rate in HCC was significantly higher than adjacent cirrhosis tissues and normal tissues (P 0.05). Conclusion There is a marked increased expression of Survivin in HCC, which may play an important role in breaking the balance of proliferation and apoptosis of HCC cells. The correlation between Survivin and p53 expression in HCC indicates that cooperation between Survivin and p53 plays a certain role in occurrence and /or development of HCC.

Knockdown of Tripartite Motif-Containing Protein 37 (TRIM37) Inhibits the Proliferation and Tumorigenesis in Colorectal Cancer Cells

Tripartite motif-containing protein 37 (TRIM37), a new member of the RING-B-box-coiled-coil (RBCC) subfamily of zinc finger proteins, was found to be involved in the development and progression of several cancers. However, the expression pattern and biological functions of TRIM37 in colorectal cancer (CRC) remain unknown. Therefore, in the present study, we examined the expression pattern of TRIM37 in CRC and investigated the function of TRIM37 in the progression of CRC. Our results showed that TRIM37 expression was upregulated in CRC cell lines. Knockdown of TRIM37 inhibited CRC cell proliferation and tumor growth in vivo. Furthermore, knockdown of TRIM37 inhibited the migration and invasion in CRC cells. Last, knockdown of TRIM37 inhibited the protein level expression of β-catenin, cyclin D1, and c-Myc in CRC cells. In conclusion, these results demonstrate that TRIM37 may play an important role in the proliferation, invasion, and tumorigenesis of CRC cells. Thus, TRIM37 may be a potential therapeutic target for the treatment of CRC.

MiR-429 suppresses neurotrophin-3 to alleviate perineural invasion of pancreatic cancer

Perineural invasion (PNI) potentially increases the risk of relapse and abdominal pain in patients with pancreatic ductal adenocarcinoma (PDAC). However, the underlying mechanisms of PNI of PDAC is incompletely revealed. Our study aimed to investigate roles of miR-429 in modulating PNI in PDAC. We found that miR-429 was downregulated in PDAC cancer tissues and was profoundly decreased in tissues with PNI. It was reduced in nine of the ten examined pancreatic cancer cell lines. MiR-429 mimics restored its cellular expressions in MIA PaCa-2 and BxCP3 cells and significantly suppressed cell viability and invasion of the cancer cells. The online bioinformatic software predicted that neurotrophin-3 (NT-3) was a potential target gene of miR-429. It was showed that NT-3 mRNA elevated in PC cancer tissues, especially in patients presenting PNI. MiR-429 upregulation substantially suppressed the NT-3 mRNA and secretion in cancer cells. Also, the dual luciferase reporter assays confirmed the interaction between miR-429 and NT-3. When co-culturing the two PDAC cells with PC-12 cells, the invaded cell counts significantly increased comparing with the sole culture of cancer cells. However, miR-429 mimic transfection or NT-3 blocking retarded the cancer invasion in the co-culture system. Besides, we found that cancer cells conditioned medium (CM) treatment significantly increased the neurite outgrowth percentage in PC-12 cells, which was suppressed by culturing with CM from miR-429 mimics-transfected cells. In the CM cultured PC-12 cells, NT-3 receptor TrkC as well as pain-related proteins TRPV1 and TRPV2 significantly elevated. Collectively, miR-429 potentially suppressed neurotrophin-3 to alleviate PNI of PDAC.