Knockdown of TMPRSS3 inhibits cell proliferation,migration/invasion and induces apoptosis of glioma cells

Transmembrane protease serine 3 (TMPRSS3) is a member of type II transmembrane serine proteases (TTSP) family, which play important roles in the development and progression of various cancers. However, the role of TMPRSS3 in glioma remains unclear. In the present study, we evaluated the expression patterns of TMPRSS3 in clinical tumor samples and glioma cell lines. The results showed that TMPRSS3 was highly expressed in both human glioma tissues and cell lines. Knockdown of TMPRSS3 in glioma cells by transfection with small interfering RNA targeting TMPRSS3 (si-TMPRSS3) significantly suppressed cell proliferation and migration/invasion. Moreover, knockdown of TMPRSS3 markedly elevated the apoptotic rate of glioma cells. Si-TMPRSS3 transfection also resulted in a remarkable increase in bax expression and a notable decrease in bcl-2 expression in glioma cells. Furthermore, TMPRSS3 knockdown markedly suppressed the expressions of Notch1 and Hes1. The results indicated that knockdown of TMPRSS3 exhibited antiglioma effect, which is associated with the inactivation of the Notch signaling pathway. These findings suggested that TMPRSS3 might be used as a therapeutic target for glioma treatment.     

Knockdown of TSP50 inhibits cell proliferation and induces apoptosis in P19 cells

Earlier studies identified testes-specific protease 50 (TSP50), which encodes a threonine protease, and showed that it was abnormally reactivated in many breast cancer biopsies. Further, it was shown to be negatively regulated by the p53 gene. However, little is known about the biological function of TSP50. In this study, we applied RNA interference to knockdown TSP50 gene expression in P19 murine embryonal carcinoma stem cells and tested whether this modulated the cell phenotype. The results showed that downregulation of TSP50 expression not only reduced cell proliferation, colony formation, and migration but also induced cell apoptosis. Further investigation revealed that knockdown of TSP50 resulted in greater sensitivity to doxorubicin-induced apoptosis and that activation of caspase-3 was involved in this process.     

Shikonin inhibits the proliferation and induces the apoptosis of human HepG2 cells

This study investigated the potential of shikonin as an anticancer agent against liver cancer and an in vitro human hepatoma cancer model system. The HepG2 cell line was the hepatoma cancer model in the present study. The inhibitory effect of shikonin on the growth of HepG2 cells was measured by MTT assay. To explore the underlying mechanism of cell growth inhibition of shikonin, the cell cycle distribution, DNA fragmentation, mitochondrial membrane potential (ΔΨm) disruption, and expression of Bax and Bcl-2 were measured in HepG2 cells. The activity of shikonin in inducing apoptosis was investigated through the detection of Annexin V signal and CD95 expression by flow cytometry and electron microscopy, respectively. Shikonin inhibited the growth of HepG2 cells in a dose-dependent manner. The IC50 value (inhibiting cell growth by 50%) was 4.30 mg/mL. Shikonin inhibited cell growth in a dose-dependent manner and blocked HepG2 cell cycle progression at the S phase. The changes in mitochondrial morphology, dose-dependently decreased in ΔΨm, were observed in different concentrations of the drug treatment group. Western blot analysis showed that cajanol inhibited Bcl-2 expression and induced Bax expression. Furthermore, we show that shikonin increases Annexin V signal and CD95 (Fas/APO) expression, resulting in apoptotic cell death of HepG2 cells. In addition, lump formation of intranuclear chromatin, pyknosis of cell nucleus, deletion of microvillus, vacuolar degeneration of mitochondria, reduction of rough endoplasmic reticulum, and resolution of free ribosome, etc., associated with apoptosis were discovered by electron microscopy in HepG2 cells after 48 h treatment. Shikonin inhibited HepG2 cells, possibly through the pathway of inducing early apoptosis, and was beneficial for restoring the apoptotic sensitivity of HepG2 cells by CD95, and should therefore be considered as a candidate agent for the prevention or treatment of human hepatoma.     

MicroRNA-1284 inhibits proliferation and induces apoptosis in SGC-7901 human gastric cancer cells

Objectives

To explore the functional effects of miR-1284 on gastric cancer cells.

Results

Overexpression of miR-1284 significantly reduced SGC-7901 cell proliferation, but improved apoptosis. However, miR-1284 suppression displayed the inversed impacts. Furthermore, the protein levels of p27, Bax, procaspase-3 and active caspase-3 were up-regulated by miR-1284 overexpression, but were down-regulated by miR-1284 suppression. The level of Bcl-2 was down-regulated by miR-1284 overexpression, while it was up-regulated by miR-1284 suppression. The level of p21 was unaffected.

Conclusion

These results suggest that miR-1284 overexpression might be a suppressor for gastric cancer via controlling of cell proliferation and apoptosis.

     

NK4 inhibits the proliferation and induces apoptosis of human rheumatoid arthritis synovial cells

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by proliferation and insufficient apoptosis of synovial cells. NK4 is a hepatocyte growth factor antagonist and is implicated in cell proliferation, viability, and apoptosis of many tumour cells. This study aimed to investigate the role of NK4 in the regulation of human RA synovial cell proliferation and apoptosis. Fibroblast‐like synoviocytes (FLSs) isolated from RA patients and MH7A synovial cells were subjected to MTT, flow cytometry, and Western blot analysis. We found that NK4 suppressed cell proliferation through cell cycle arrest at the G0/G1 phase and induced apoptosis in RA synovial cells. Furthermore, NK4 altered the expression of cell cycle and apoptosis‐related proteins such as cyclin D1, cyclin B1, PCNA, p21, p53, Bcl‐2, Bax, cleaved caspase‐9, and cleaved caspase‐3. Additionally, NK4 reduced the phosphorylation level of NF‐κB p65 and upregulated the expression of sirt1, but did not change the levels of p38 and p‐p38 in RA‐FLS and MH7A cells. In conclusion, NK4 inhibits the proliferation and induces apoptosis of human RA synovial cells. NK4 is a promising therapeutic target for RA. We demonstrated that NK4 inhibited cell proliferation by inducing apoptosis and arresting cell cycle in RA‐FLS and MH7A cells. The apoptotic effects of NK4 may be mediated in part by decreasing Bcl‐2 protein level, increasing Bax and caspase 3 protein levels, and inhibiting NF‐κB signalling in RA‐FLS and MH7A cells. These findings reveal potential mechanism underlying the role of NK4 in RA synovial cells and suggest that NK4 is a promising agent for RA treatment.     

Knockdown of zinc finger protein, X-linked (ZFX) inhibits cell proliferation and induces apoptosis in human laryngeal squamous cell carcinoma

Apoptosis is a natural form of cell death involved in many physiological changes in the cell. Defects in the process of apoptosis can lead to serious diseases. During some apoptotic pathways, proteins apoptosis-inducing factor (AIF) and endonuclease G (EndoG) are released from the mitochondria and they translocate into the cell nuclei, where they probably participate in chromatin degradation together with other nuclear proteins. Exact mechanism of EndoG activity in cell nucleus is still unknown. Some interacting partners like flap endonuclease 1, DNase I, and exonuclease III were already suggested, but also other interacting partners were proposed. We conducted a living-cell confocal fluorescence microscopy followed by an image analysis of fluorescence resonance energy transfer to analyze the possibility of protein interactions of EndoG with histone H2B and human DNA topoisomerase II alpha (TOPO2a). Our results show that EndoG interacts with both these proteins during apoptotic cell death. Therefore, we can conclude that EndoG and TOPO2a may actively participate in apoptotic chromatin degradation. The possible existence of a degradation complex consisting of EndoG and TOPO2a and possibly other proteins like AIF and cyclophilin A have yet to be investigated.     

Geldanamycin induces mitotic catastrophe and subsequent apoptosis in human glioma cells

Geldanamycin (GA) binds to heat shock protein 90 (Hsp90) and interferes with its function which is to protect various cellular proteins involved in signaling, growth control, and survival from ubiquitination and subsequent degradation by the proteasome. Recently, we demonstrated that GA inhibited migration of glioma cells in vitro associated with downregulation of hypoxia-inducible factor (HIF-1 alpha) and phosphorylation of focal adhesion kinase (FAK) (Zagzag et al., 2003, J Cell Physiol 196:394-402). Here, we have investigated the mechanisms through which GA treatment of the T98G glioma cell line induces apoptosis. We found that GA treatment induced cell death in a caspase-dependent manner through activation of caspase-3 and PARP cleavage together with release of cytochrome c and apoptosis inducing factor (AIF) from the mitochondria. Use of synchronized T98G cells showed that GA treatment of glioma cells during S-phase enhanced cytotoxicity followed by M-phase arrest, resulting in mitotic catastrophe. In addition, apoptosis was associated with the downregulation of the survival protein, phosphorylated Akt (pAkt), an important signaling protein in the PI3K pathway, that is overexpressed in many cancers including gliomas. Given that many glioma tumors show deregulation of the PI3K signaling pathway, either through loss of the tumor suppressor protein PTEN or overexpression of the growth factor EGFR, the ability to identify different subsets of patients using simple immunohistochemistry for the presence of absence of pAkt could enable selection of the appropriate kinase inhibitor, such as GA, for drug therapy. Based on our data presented here, GA or its analogs may have potential in the treatment of glioma.     

Matrine inhibits proliferation and induces apoptosis of human colon cancer LoVo cells by inactivating Akt pathway

The present study has investigated the anti-tumor activity and the underlying mechanisms of matrine on human colon cancer LoVo cells. Matrine inhibited the proliferation of the cells in dose- and time-dependent manners. The concentration required for 50 % inhibition (IC₅₀) was 1.15, 0.738, and 0.414 mg/ml, when cell were incubated with matrine for 24, 48, and 72 h, respectively. Matrine induced cell cycle arrest at G1 phase by downregulating cyclin D1 and upregulating p27 and p21. Matrine induced cell apoptosis by reducing the ratio of Bcl-2/Bax and increasing the activation of caspase-9 in a dose-dependent manner. Matrine displayed its anti-tumor activity by inactivating Akt, the upstream factor of the above proteins. Matrine significantly reduced the protein levels of pAkt, and increased the protein levels of other downstream factors, pBad and pGSK-3β. Specific inhibition of pAkt induced cell apoptosis, and synergized with matrine to inhibit the proliferation of LoVo cells; whereas activation of Akt neutralized the inhibitory effect of matrine on cell proliferation. The present study has demonstrated that matrine inhibits proliferation and induces apoptosis of human colon cancer LoVo cells by inactivating Akt pathway, indicating matrine may be a potential anti-cancer agent for colon cancer.     

香烟烟雾提取物抑制肺泡上皮细胞的增殖并诱导其凋亡

香烟烟雾提取物（cigarette smoke extract,CSE）中含有丰富的氧化剂和自由基,由它所引起的氧化应激可导致肺泡壁的损伤进而发展为肺气肿.近年来,围绕CSE损伤肺泡壁作用机制的研究较为活跃,但其结果却一直存在着分歧.本实验的目的是观察CSE对肺泡Ⅱ型上皮细胞的损伤作用并探讨与其相关的分子机制.MTT比色法的结果显示,CSE以时间和剂量依赖性的方式降低细胞的增殖活力,流式细胞术的分析结果表明细胞增殖周期被阻滞在G1/S期.Hoechst 33258染色以及透射电镜观察从形态上确认CSE诱导细胞凋亡的发生,DNA梯的出现和Annexin V-FITC/碘化丙啶双染色的结果从分子水平得到进一步的证实.同时,运用流式细胞术检测到CSE诱导的凋亡伴随着Fas受体的高表达和caspase-3的显著活化.另外,使用H2DCFDA染色,经激光共聚焦显微镜术测得细胞内氧自由基在细胞受到CSE刺激以后大量快速积累.结果表明CSE能够抑制肺泡Ⅱ型上皮细胞来源的A549细胞的生长和增殖,并诱导细胞凋亡,由Fas受体所介导的死亡受体途径参与此凋亡过程,而CSE所引起的氧化应激则可能是阻止肺泡上皮细胞生长增殖并诱导其凋亡的始动因素.     

Guggulsterone induces apoptosis and inhibits lysosomal-dependent migration in human bladder cancer cells

BackgroundThe survival rate and therapeutic options for patients with bladder cancer have improved little in recent decades. Guggulsterone (GS), a phytoestrogen, has been investigated as an anticancer drug in various malignancies.PurposeThe present study aimed to evaluate the anticancer effects of E-isomer and Z-isomer GS in the human bladder cancer cell lines TSGH8301 (low-grade) and T24 (high-grade) and their underlying mechanisms.MethodsThe cell survival effect of GS was investigated by the MTT and colony formation assays in bladder cancer cell lines. Flow cytometry was used to analyze the cell cycle and cell death. Migration ability was measured by wound healing and transwell assays. Protein expression was determined by Western blot after GS treatment. The potency of GS on subcutaneous TSGH8301 bladder tumors was evaluated using an in vivo imaging system.ResultsE-isomer GS reduced the survival rate of both low- and high-grade human bladder cancer cells. GS caused cell cycle arrest, accompanied by the decrease and increase in cyclin A and p21 levels, respectively. Additionally, caspase-dependent apoptosis was observed following GS treatment. Furthermore, GS treatment downregulated mTOR-Akt signaling and induced autophagy with p62 and LC3β-II expression. Moreover, the farnesoid X receptor was involved in GS-inhibited cell growth. In addition, GS reduced the migration ability with a decrease in integrin-focal adhesion kinase and myosin light chain. Interestingly, the suppression of GS-mediated migration was prevented by the lysosomal inhibitor ammonium chloride (NH₄Cl). GS also reduced TSGH8301 bladder cancer cell progression by increasing the level of p21, cleaved caspase 3, cleaved poly (ADP-ribose) polymerase (PARP), and LC3β-II in vivo.ConclusionsThe current findings suggest that GS treatment may serve as a potential anticancer therapy for different grades of urothelial carcinoma.     

Matrine inhibits proliferation and induces apoptosis via BID-mediated mitochondrial pathway in esophageal cancer cells

Matrine, as a member of Sophora family, is an alkaloid found in plants, and produces plethora pharmacological effects, including anti-cancer effects. However, the mechanism involved remains largely unknown. This study is conducted to investigate the anti-cancer mechanisms of matrine in human esophageal cancer in vitro and in vivo. In human esophageal cancer cell Eca-109, matrine significantly decreased the cell viability in a dose-dependent manner, and induced apoptosis as well as cell cycle arrest in G0/G1 phase by up-regulation of P53 and P21. The expression of several apoptosis-related proteins in cells and tumor tissues were evaluated by Western blot analysis. We found that matrine induced cell apoptosis by down-regulation of the ratio of BCL-2/BID and increasing activation of caspase-9. Further studies indicated that matrine induced apoptosis of Eca-109 was through the mitochondria-mediated internal pathway, but not by death receptor-mediated extrinsic apoptotic pathway, which was confirmed by the fact that Bid translocated from the nucleus to mitochondria during the process of the apoptosis induced by matrine. In vivo study found that matrine effectively inhibited the tumor formation of Eca-109 cells in nude mice. Our study suggests that matrine could serve as a potential novel agent from natural products to treat esophageal cancer.     

7,8-Dihydroxyflavone suppresses proliferation and induces apoptosis of human osteosarcoma cells

Recent studies suggest that 7,8-dihydroxyflavone (7,8-DHF) inhibits the development of several tumors.However,its role in osteosarcoma (OS) remains unknown.This...     

Analgesic-antitumor peptide inhibits proliferation and migration of SHG-44 human malignant glioma cells

Malignant gliomas, the most common subtype of primary brain tumors, are characterized by high proliferation, great invasion, and neurological destruction and considered to be the deadliest of human cancers. Analgesic-antitumor peptide (AGAP), one of scorpion toxic polypeptides, has been shown to have antitumor activity. Here, we show that recombinant AGAP (rAGAP) not only inhibits the proliferation of gliomas cell SHG-44 and rat glioma cell C6, but also suppresses the migration of SHG-44 cells during wound healing. To explain these phenomena, we find that rAGAP leads to cell cycle of SHG-44 arrested in G1 phase accompanied by suppressing G1 cell cycle regulatory proteins CDK2, CDK6, and p-RB by means of the down-regulated protein expression of p-AKT. Meanwhile, rAGAP significantly decreases the production of NF-κB, BCL-2, p-p38, p-c-Jun, and p-Erk1/2 and further suppresses the activation of VEGF and MMP-9 in SHG-44 cells. These findings suggest rAGAP inhibit proliferation and migration of SHG-44 cells by arresting cell cycle and interfering p-AKT, NF-κB, BCL-2, and MAPK signaling pathways.