scholarly journals Inhibition of Siah2 Ubiquitin Ligase By Vitamin K3 Attenuates Hypoxia and Blocks K562-R Cells Resistance

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5214-5214
Author(s):  
Na Xu ◽  
Yajuan Xiao ◽  
Xuan Zhou ◽  
Lin Li ◽  
Fen Huang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Expression ◽  
Western Blot ◽  
Cell Lines ◽  
Oxygen Concentration ◽  
Ubiquitin Ligase ◽  
Vitamin K3 ◽  
Imatinib Resistance ◽  
Hematopoietic Stem ◽  
Expression Levels

Abstract Background and objective: Hypoxia has been shown to favor the self-renewal of murine and human hematopoietic stem cells. Hypoxia as a key feature of the “stem cell niches” in vivo, and studies found that hypoxia modified proliferation and differentiation of chronic myeloid leukemia (CML) stem cells.The E3 ubiquitin ligase Siah2 is an important regulator of the hypoxic response,which has been implicated in the regulation of the hypoxia response, as well as in the control of Ras, JNK/p38/NF-κB, MAPK signaling pathways. In the present study, we identified that SIAH2 induced k562 cells resistance to imatinib by hypoxia-inducible factor (HIF)-1a activated vascular epithelial growth factor (VEGF) pathway in hypoxia micro environment. In this study we show that SIAH2/Hif-1α induced K562 cell remain in G0 stage and resistance to imatinib, and we verified that vitamin K3 (SIAH2 inhibitor) reversed K562-R drug resistance in hypoxia microenvironment. Methods: We detected Siah2 expression levels in K562-wild type (K562-W) and K562-imatinib-resistance type (K562-R) cell lines by western blot analysis. Those two cell lines were further cultured for 24 h and 48 h under the condition of normal and hypoxia concentration of oxygen (1%, 5%), and treated K562-R with 0A5A15A30mM vitamin K3 for 72 hours in hypoxia concentration, explored cell cycle and apoptosis by flow cytometry (FCM) dyed by Annexin-V; analyzed the expression levels of Siah2, HIF-1α respectively by real-time PCR and western blot. Results: The protein of Siah2 and HIF-1α was significantly higher in K562-R compared with K562-w cells (P<0.01). Cell cycle analysis showed a 3% increase in K562-W G0/G1 cells in 1% O2 compared with normal O2, and 7% in K562-R. Under hypoxia condition, the cellular apoptotic ratio of K562-R was 5.46%, much less than 11.08% in K562-W cells. G0 cell proportion increased significantly with the long time Hypoxia (P<0.01); After being cultured in 1% oxygen concentration for 24 hours, we confirmed Siah2,HIF-1α were all up regulated in both cell lines, moreover, it was more obvious in K562-R cells. Siah2 protein expression increased along treated with vitamin K3 concentration (0, 5, 15, 30 mM) (P<0.05), on the contrary ,HIF-1α protein expression decreased with vitamin K3 concentration (P<0.05). the proportion of G0 cell was decreased in K562-R cells treated with Vitamin K3 (15mM)for 48h Under the condition of 1% oxygen concentration compared with control group (P=0.02). Conclusions: Hypoxia up-regulated of Siah2 and Hif-1α expression in K562-R, promoted cell apoptosis and arrested cells in G0 stage, and reduced cell sensitivity to Imatinib. Vitamin K3 could inhibit Siah2 and lowered Hif-1α in K562-R. These findings reveal an effective treatment for the identification of Siah2 inhibitors and would reverse TKI resistance for CML patients. Disclosures No relevant conflicts of interest to declare.

The E3 Ubiquitin Ligases Siah2 Contributes To Imatinib Resistance In Chronic Myeloid Leukemia In Hypoxia Condition

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2530-2530
Author(s):  
Na Xu ◽  
Xiaozhen Xiao ◽  
Yajuan Xiao ◽  
Xuan Zhou ◽  
Guanlun Gao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Western Blot ◽  
Real Time ◽  
Oxygen Concentration ◽  
Conflicts Of Interest ◽  
Signal Molecule ◽  
Imatinib Resistance ◽  
Hematopoietic Stem ◽  
Significant Difference

Abstract Background and Objective Severe hypoxia has been shown to favor the self-renewal of human hematopoietic stem cells. Recent studies demonstrate that hypoxia via hypoxia-inducible factor (Hif-1a) can modify the proliferation and differentiation of CML stem cells. The ubiquitin E3 ligase SIAH2 is an important regulator of the hypoxic response as it leads to the ubiquitin/proteasomal degradation of prolyl hydroxylases such as PHD3, which in turn increases the stability of Hif-1a. The Hif-1a has been linked to chemosensitivity while the underlying molecular mechanism remains elusive. Therefore, we comprehensively analysed SIAH2 and Hif-1a role in determining chemosensitivity via signal molecule vascular epithelial growth factor (VEGF) pathway. Methods We tested the level of Siah2, Hif-1a and VEGF in Imatinb-sensitive CML Patients (n=15) and insensitive CML patients(n=10) by real-time reverse transcription PCR and western blot analysis. K562-wild type (K562-W) and K562-imatinib-resistance type (K562-R) cell were cultured for 24h and 48h under the condition of normal and hypoxia concentration of oxygen (1%). Knockdown of SAIH2 by RNA interference ,Cell viability and IC50 under 1% concentration oxygen were tested by cck-8;detected cell cycle and apoptosis by flow cytometry (FCM) ; analyzed the expression levels of Siah2, Hif-1a and VEGF respectively by real-time PCR and western blot. Results The level of mRNA and protein of SIAH2 ,Hif-1a and VEGF were significantly higher in IM- resistant CML patients compared to IM sensitive CML patients, respectively (P<0.05). The similar results were observed in K562-R and K562-W cells(P<0.01). Under 21%,1% oxygen concentration cultured for 24h ,the IC50 of K562-W and K562-R cells was no significant difference (P<0.05),but there was significant difference after cultured for 48h Cell cycle analysis showed that more G0/G1 cells in K562-R than K562-W after cultured for 48h under hypoxia condition(P<0.05). After being cultured in 1% oxygen concentration for 48 hours, we confirmed SIAH2, Hif-1a and VEGF were up-regulated in both cell lines, moreover, it was more obvious in K562-R cells. In SIAH2-sh K562-R cells, the apoptosis was higher than K562-R cells obviously under 1% oxygen concentration for 48 hours(P<0.05);the level of Hif-1a Hardly monitored, and the level of VEGF was also lower. Conclusions There were higher level of SIAH2, Hif-1a and VEGF in IM-resistant CML patients. Under hypoxia condition, K562 cells were likely to improve their resistance to Imatinib .After we Knockdown SAIH2, K562-R Cell apoptosis rate increased significantly, along with low level Hif-1a and VEGF. It indicated that in hypoxia micro-environment, Siah-2 might be one of the critical molecules that induce Imatinib-resistance in CML in the way of maintaining leukemic cell survival and stimulating them into quiescence phase. Disclosures: No relevant conflicts of interest to declare.

Protein Expression Patterns of Candidate Genes Involved in Apoptosis and Cell Cycle Control in Genetic Subgroups of Chronic Lymphocytic Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2796-2796
Author(s):  
Christof Schneider ◽  
Dirk Winkler ◽  
Meike Loddenkemper ◽  
Alexander Krober ◽  
Peter Lichter ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Expression ◽  
Cyclin D1 ◽  
Candidate Genes ◽  
Cell Lines ◽  
Normal Karyotype ◽  
Lymphocytic Leukemia ◽  
Expression Levels ◽  
Protein Levels ◽  
Atm Protein

Abstract Chronic lymphocytic leukemia (CLL) is a heterogeneous disease with a highly variable clinical course. Genomic aberrations (such as 13q−, 11q−, +12q, 17p−) can be found in about 80% of CLL cases and define pathogenic as well as clinical subgroups. Similarly, the mutational status of the variable region of the immunoglobulin heavy-chain gene (VH) identifies subgroups with different maturation stage and clinical outcome. In this study protein expression levels of candidate genes involved in cell cycle and apoptosis control (p53, ATM, Akt1, PI3-K, p21, p27, cdk4, Cyclin-D1, D2, D3, Bax, Bcl-2, Apaf-1, Smac, XIAP, cIAP2, survivin) were examined by Western Blotting. A total of 87 CLL cases derived from the subgroups with 11q- (n=22), 17p-/p53 mutation (n=18), +12q (n=24), 13q- (n=8) or a normal karyotype (n=15) were studied and compared to the cell lines EHEB and JVM-2. VH-mutation status was available for 65 cases (unmutated n=48, mutated n=17). Due to limitations in sample availability not all proteins could be examined in all cases. A highly homogenous expression pattern for all the proteins studied was observed in the CLL subgroup with a normal karyotype. This pattern was independent of the VH-status. CLL samples with normal karyotype, +12q and 13q deletion showed equal levels of ATM as compared to EHEB and JVM-2. As compared to cases with a normal karyotype the ATM level within the 11q- subgroup was reduced in 5 cases and absent in 1 case among 11 evaluable 11q- cases. The 17p- subgroup was comprised of 3 cases with concomitant 17p- and 11q- and 15 cases with 17p- but no 11q-. The latter group showed ATM protein levels comparable to the levels of the normal karyotype group. In the group with 17p- and 11q- there was an ATM expression level similar to the groups with 17p- and normal karyotype in two cases while one case had a reduced ATM protein level comparable to the 11q- subgroup. All cases with 17p- exhibited a stronger expression of p53 as compared to the cell lines and all other cases, except for one case with normal karyotype and one with an 11q-. No p53 mutations could be detected in exons 5–9 by sequencing in these two cases. High levels of survivin protein were found in all cases with 17p- and/or 11q-, 13q-, +12q while the subgroup with a normal karyotype showed lower levels. High levels of cdk4 protein were expressed in cases with 17p-, 11q- and 13q- while cdk4 protein levels were low in the subgroup with +12q and normal karyotype. Regarding p21, p27, Bcl2, Bax, Smac, Apaf-1, Cyclin D1–D3, cIAP2, XIAP, Akt1 and PI3K no variation in the expression levels were observed across the genetic CLL subgroups. Comparing the CLL cases to the cell lines the differences in expression levels were found for the cell cycle regulators Cyclin D1, D2, D3, p21 and p27. While the cell lines showed strong protein levels for Cyclin D1, D2, D3 and p21, they were nearly absent in the CLL cases. Expression of p27 was higher in all CLL cases as compared to JVM-2 and EHEB. In conclusion, the 17q- subgroup was the only group with a high level of p53 protein expression indicating that p53 is the affected gene in this subgroup. In contrast, the ATM protein levels are reduced only in a part of the 11q- cases indicating a possible role of additional candidate genes. Cases with +12q and normal karyotype showed weak expression of cdk4 pointing out a possible function in these subgroups.

Treatment with cucurbitacin B alone and in combination with gefitinib induces cell cycle inhibition and apoptosis via EGFR and JAK/STAT pathway in human colorectal cancer cell lines

2015 ◽  
Vol 35 (5) ◽  
pp. 526-543 ◽  
Author(s):  
AS Yar Saglam ◽  
E Alp ◽  
Z Elmazoglu ◽  
S Menevse
Keyword(s):  
Cell Cycle ◽  
Western Blot ◽  
Cell Lines ◽  
Janus Kinase ◽  
Synthesis Rate ◽  
Phosphate Binding ◽  
Active Inhibitor ◽  
Expression Levels ◽  
Cucurbitacin B ◽  
Cell Cycle Inhibition

The epidermal growth factor receptor (EGFR) associated with signaling pathways, such as Janus kinase (JAK)/signal transducer and activator of transcription (STAT), plays an important role in colorectal cancers (CRCs). Gefitinib (Gef) is an orally active inhibitor targeting the adenosine tri phosphate-binding domain of EGFR, and cucurbitacin B (CuB) is a selective inhibitor of JAK/STAT signaling with potent antitumor activity via suppression of STAT3 phosphorylation, but the underlying mechanism is not clear. We aimed to investigate the apoptotic and antiproliferative effects of CuB as a single agent and in combination with Gef on both HT-29 and HCT-116 cell lines. Cell proliferation, cell cycle distribution, and apoptosis were evaluated using viability assay, fluorescent microscopy, cytotoxicity assay, proliferation, DNA fragmentation, and cleaved caspase 3 levels. Real-time polymerase chain reaction and Western blot analyses were performed to determine the expression of relevant genes and proteins including antiapoptotic, proapoptotic, and cell cycle regulation. EGFR, phosphorylated EGFR (pEGFR), STAT3, and pSTAT3 proteins were evalutaed with Western blot analysis. Our results showed that, compared to CuB alone, CuB plus Gef treatment caused a significant growth and cell cycle inhibition and induced apoptosis in both cell lines. Also CuB plus Gef treatment decreased DNA synthesis rate more effectively than CuB alone. Treatment with CuB alone and in combination with Gef decreased the expression levels of B-Cell CLL/Lymphoma 2 (Bcl-2), BCL2-like 1 (BCL2L1), cyclin D1, pSTAT3, and pEGFR and increased the expression levels of Bcl-2-like protein 4, Bcl-2 homologous antagonist/killer, Bcl-2-associated death promoter, Bcl-2-like protein 11, and p27kip1 levels. Our results suggest that treatment with CuB alone and more likely in combination with Gef may be a considerable alternative therapeutic approach for CRC, at least in vitro.

scholarly journals Layers of regulation of cell-cycle gene expression in the budding yeast Saccharomyces cerevisiae

2018 ◽  
Vol 29 (22) ◽  
pp. 2644-2655 ◽  
Author(s):  
Christina M. Kelliher ◽  
Matthew W. Foster ◽  
Francis C. Motta ◽  
Anastasia Deckard ◽  
Erik J. Soderblom ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Protein Expression ◽  
Ubiquitin Ligase ◽  
Budding Yeast ◽  
Cell Cycle Regulators ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Expression Levels ◽  
Mutant Cells

In the budding yeast Saccharomyces cerevisiae, transcription factors (TFs) regulate the periodic expression of many genes during the cell cycle, including gene products required for progression through cell-cycle events. Experimental evidence coupled with quantitative models suggests that a network of interconnected TFs is capable of regulating periodic genes over the cell cycle. Importantly, these dynamical models were built on transcriptomics data and assumed that TF protein levels and activity are directly correlated with mRNA abundance. To ask whether TF transcripts match protein expression levels as cells progress through the cell cycle, we applied a multiplexed targeted mass spectrometry approach (parallel reaction monitoring) to synchronized populations of cells. We found that protein expression of many TFs and cell-cycle regulators closely followed their respective mRNA transcript dynamics in cycling wild-type cells. Discordant mRNA/protein expression dynamics was also observed for a subset of cell-cycle TFs and for proteins targeted for degradation by E3 ubiquitin ligase complexes such as SCF (Skp1/Cul1/F-box) and APC/C (anaphase-promoting complex/cyclosome). We further profiled mutant cells lacking B-type cyclin/CDK activity ( clb1-6) where oscillations in ubiquitin ligase activity, cyclin/CDKs, and cell-cycle progression are halted. We found that a number of proteins were no longer periodically degraded in clb1-6 mutants compared with wild type, highlighting the importance of posttranscriptional regulation. Finally, the TF complexes responsible for activating G1/S transcription (SBF and MBF) were more constitutively expressed at the protein level than at periodic mRNA expression levels in both wild-type and mutant cells. This comprehensive investigation of cell-cycle regulators reveals that multiple layers of regulation (transcription, protein stability, and proteasome targeting) affect protein expression dynamics during the cell cycle.

scholarly journals The Increased lncRNA MIR503HG in Preeclampsia Modulated Trophoblast Cell Proliferation, Invasion, and Migration via Regulating Matrix Metalloproteinases and NF-κB Signaling

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Dan Cheng ◽  
Shan Jiang ◽  
Jiao Chen ◽  
Jie Li ◽  
Liangfei Ao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
Protein Expression ◽  
Western Blot ◽  
Trophoblast Cell ◽  
Migration Flow ◽  
Expression Levels ◽  
Invasion And Migration ◽  
And Migration

Background. Preeclampsia (PE) is a pregnancy-related syndrome characterized by hypertension and proteinuria after the 20th week of gestation. The long noncoding RNAs (lncRNAs) have been recently discovered for their roles in the pathogenesis of PE. This study is aimed at determining the expression of lncRNA MIR503 host gene (MIR503HG) in PE placental tissues and exploring the molecular mechanism underlying MIR503HG-mediated trophoblast cell proliferation, invasion, and migration. Methods. The expression level of MIR503HG in placental tissues, HTR-8/SVneo, and JEG3 cells was determined by quantitative real-time PCR; western blot detected the relevant protein expression levels in HTR-8/SVneo and JEG3 cells; flow cytometry determined cell apoptosis and cell cycle of HTR-8/SVneo and JEG3 cells; trophoblast cell proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells were measured by CCK-8, transwell invasion, and wound healing assays, respectively. Results. The highly expressed MIR503HG was detected in PE placental tissues compared to normal placental tissues. MIR503HG overexpression suppressed cell proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells, while knockdown of MIR503HG increased trophoblast cell proliferation, invasion, and migration. Flow cytometry results showed that MIR503HG overexpression induced apoptosis and caused cell cycle arrest at the G0/G1 phase, while MIR503HG knockdown had the opposite actions in HTR-8/SVneo and JEG3 cells. Western blot assay results showed that MIR503HG overexpression suppressed the matrix metalloproteinase-2/-9 and the snail protein expression and increased the E-cadherin expression in trophoblast cells. In addition, MIR503HG overexpression suppressed the NF-κB signaling pathway by inhibiting the phosphorylation of IκBα and the nuclear translocation of NF-κB signaling subunit p65. On the other hand, MIR503HG knockdown played an opposite role in these protein expression levels. Conclusion. Our results showed that MIR503HG inhibited the proliferation, invasion, and migration of HTR-8/SVneo and JEG3 cells, which may be related to the pathogenesis of PE.

Proteasome Inhibition Leads to Dephosphorylation and Downregulation of Protein Expression of Members of the Akt/mTOR Pathway In MCL

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4449-4449
Author(s):  
Grit Hutter ◽  
Yvonne Zimmermann ◽  
Malte Rieken ◽  
Elena Hartmann ◽  
Wolfgang Hiddemann ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Expression ◽  
Western Blot ◽  
Cell Lines ◽  
Translation Initiation ◽  
Proteasome Inhibition ◽  
Mtor Pathway ◽  
Blue Staining ◽  
Erk Pathway ◽  
Bortezomib Treatment

Abstract Abstract 4449 Background: Mantle cell Lymphoma (MCL) is a distinct B-cell subtype characterized by the chromosomal translocation t(11;14)(q13;q32), an especially poor clinical outcome and low response to chemotherapy. The proteasome inhibitor bortezomib is approved for treatment of relapsed and refractory MCL and achieves a response rate of 30–40%. However, little is known which molecules represent the critical targets of proteasome inhibition and how different regulators of cell cycle and apoptosis are affected. Bortezomib has been shown to inactivate the NFkB pathway in MCL, but recent findings indicate Bortezomib is also active in a proteasome –independent manner suggesting Bortezomib targets multiple pathways. Method: Four MCL cell lines (HBL2, Granta 519, Jeko-1, NCEB-1), two CLL cell lines (Mec1, Mec2) and two hematological control cell lines (Jurkat, Karpas 422) were exposed to Bortezomib at a previously defined cytotoxic concentration (25nmol). Western blot and mRNA analysis were performed for various members of the PI3K/Akt/mTOR and the MEK/ERK pathway after 24h Bortezomib exposure. Results were compared to cell proliferation (WST1, trypan blue staining), induction of apoptosis (Annexin V PE/7-AAD staining) and cell cycle data (FACS). Result: Western blot analysis revealed reduced phosphorylation of Akt at Ser473 in all cell lines while autophosphorylation of mTOR at Ser2481 was completely downregulated only in the susceptible cell lines. In addition further members of the mTOR pathway were affected by bortezomib treatment. Dephosphorylation of the 4EBP1, downregulation of p70S6 protein expression was detected in all cell lines whereas EIF4E dephosphorylation was only observed in the sensitive MCL cell lines. In contrast Bortezomib did not affect members of the MEK/ERK pathway (MEK1/2, p42/44MAPK). Interestingly the mRNA and protein expression profile of CCND1 were differently altered suggesting an involvement of bortezomib in the regulation of translation initiation. This data were also confirmed by microarray analysis. Conclusion: In this study Bortezomib treatment was shown to target the Akt/mTOR pathway especially by dephosphorylation of the translation initiation factor EIF4E and other molecules of this signal pathway. This knowledge will play a crucial role in the development of future combinations of biologicals to target the molecular pathogenesis of MCL. Disclosures: Dreyling: Johnson & Johnson: support of investigators initial trials.

scholarly journals Cannabidiol Induces Cell Cycle Arrest and Cell Apoptosis in Human Gastric Cancer SGC-7901 Cells

Biomolecules ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 302 ◽  
Author(s):  
Xin Zhang ◽  
Yao Qin ◽  
Zhaohai Pan ◽  
Minjing Li ◽  
Xiaona Liu ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Cycle ◽  
Protein Expression ◽  
Cell Cycle Arrest ◽  
Phase Cell ◽  
G1 Phase ◽  
Therapeutic Effects ◽  
Human Gastric Cancer ◽  
Expression Levels ◽  
Cycle Arrest

The main chemical component of cannabis, cannabidiol (CBD), has been shown to have antitumor properties. The present study examined the in vitro effects of CBD on human gastric cancer SGC-7901 cells. We found that CBD significantly inhibited the proliferation and colony formation of SGC-7901 cells. Further investigation showed that CBD significantly upregulated ataxia telangiectasia-mutated gene (ATM) and p53 protein expression and downregulated p21 protein expression in SGC-7901 cells, which subsequently inhibited the levels of CDK2 and cyclin E, thereby resulting in cell cycle arrest at the G0–G1 phase. In addition, CBD significantly increased Bax expression levels, decreased Bcl-2 expression levels and mitochondrial membrane potential, and then upregulated the levels of cleaved caspase-3 and cleaved caspase-9, thereby inducing apoptosis in SGC-7901 cells. Finally, we found that intracellular reactive oxygen species (ROS) increased after CBD treatment. These results indicated that CBD could induce G0–G1 phase cell cycle arrest and apoptosis by increasing ROS production, leading to the inhibition of SGC-7901 cell proliferation, thereby suggesting that CBD may have therapeutic effects on gastric cancer.

scholarly journals ENHANCED p53-DEPENDENT GROWTH INHIBITION OF HUMAN GLIOBLASTOMA CELLS BY COMBINATORIAL TREATMENT OF TEMOZOLOMIDE AND NOVEL PURIFIED NATURAL CARBOHYDRATE OF PLEUROTUS FLORIDA

2017 ◽  
Vol 9 (6) ◽  
pp. 189
Author(s):  
Priyankar Maji ◽  
Ranodeep Chatterjee ◽  
Biswa P. Choudhury ◽  
Urmi Chatterji ◽  
Jhuma Ganguly
Keyword(s):  
Cell Cycle ◽  
Protein Expression ◽  
Cell Lines ◽  
P53 Protein ◽  
Glioblastoma Cells ◽  
Human Glioblastoma ◽  
Pleurotus Florida ◽  
P53 Protein Expression ◽  
Human Glioblastoma Cells ◽  
Combinatorial Treatment

Objective: This study was designed to analyze the combinatorial chemotherapeutic effect of temozolomide (TMZ), the most common drug in glioblastoma treatment and a purified carbohydrate (Fr-II) from the edible mushroom Pleurotus florida, on human glioblastoma cell lines.Methods: Fr-II was purified by size-exclusion chromatography and characterised by different mass spectroscopy analysis. Human glioblastoma cells were treated with TMZ, Fr-II, and combination of TMZ and Fr-II. Cell cytotoxicity was measured by MTT assay, cell cycle phase distribution was determined by cell cycle analysis and followed by the relative p53 protein expression was analyzed by western blot analysis.Results: Chemical analysis of Fr-II confirmed the glycosidically linked two units of glucose with terminally attached mannitol with mass of 506 Da. Fr-II treatment exhibited cytotoxicity in both the cell lines in a dose-dependent manner with most effective dose at 200µg/ml. When Fr-II (200µg/ml) was combined with a dose range of TMZ it showed a more cellular cytotoxicity compared to the cytotoxicity of TMZ alone with most oppressive combinatorial dose at 400µM (TMZ)+200µg/ml (Fr-II). In compliance, with the above results, both cell lines showed a 10% increase in no. of cells (p<0.05) in G2/M phase indicating an arrest of cell cycle and increased p53 protein expression (p<0.05) at the combinatorial dose than TMZ alone at 400µM, but Fr-II alone didn’t show any cell cycle arrest nor did it show increased p53 expression.Conclusion: Therefore it confirms that Fr-II synergizes with TMZ to significantly intensify its anti-proliferative properties, thereby emerging as an effective element for combinatorial treatment of glioblastoma.

scholarly journals Combination of axitinib and dasatinib for anti-cancer activities in two prostate cancer cell lines

2015 ◽  
Vol 11 (1) ◽  
pp. 130
Author(s):  
Nai-Xiong Peng ◽  
Chun-Xiao Liu ◽  
Xi-Sheng Wang ◽  
Ze-Jian Zhang ◽  
Su-Cai Liao
Keyword(s):  
Prostate Cancer ◽  
Cell Cycle ◽  
Survival Rate ◽  
Cell Lines ◽  
Cancer Cell ◽  
Prostate Cancer Cell ◽  
Cancer Cell Lines ◽  
Prostate Cancer Cell Lines ◽  
Expression Levels ◽  
Different Response

<p class="Abstract">Prostate cancer is major cause of cancer related deaths worldwide in men. There are new treatment methods and drugs are being developed with promising results in two of the prostate cancer cell lines (PPC-1 and TSU-Pr1). These two cells were treated with 20 uM of axitinib combined with dasatinib for 6-72 hours. The cell viability assessed by the cytotoxicity assay. Various regulatory genes such as c-KIT, cell cycle and apoptosis and angiogenic factors were also studied. The enzyme activity of apoptosis efector caspase-3 was colorimetrically determined. Axitinib and dasatinib combination lowered the survival rate of PPC-1 cells but enhanced the survival rate of TSU-Pr1 cells. The protein expression levels in apoptosis and angiogenesis factors were also found to be in contrast between the two cell lines. PPC-1 and TSU-Pr1 cells displayed a different response to axitinib with dasatinib, which explains different expression levels of regulators of cell-cycle, apoptosis and angiogenesis.</p><p> </p>

scholarly journals Silencing of the TROP2 gene suppresses proliferation and invasion of hepatocellular carcinoma HepG2 cells

2019 ◽  
Vol 47 (3) ◽  
pp. 1319-1329 ◽  
Author(s):  
Jian Zhang ◽  
Hai Ma ◽  
Liu Yang ◽  
Hongchun Yang ◽  
Zhenxing He
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Protein Expression ◽  
Cell Lines ◽  
Epithelial Mesenchymal Transition ◽  
Migration And Invasion ◽  
Human Trophoblast ◽  
Mesenchymal Transition ◽  
Expression Levels ◽  
Proliferation And Invasion

Objectives Overexpression of human trophoblast cell surface antigen 2 (Trop2) has been observed in many cancers; however, its roles in proliferation, apoptosis, migration, and invasion of hepatocellular carcinoma (HCC) remain unclear. Thus, this study aimed to characterize the function of Trop2 in HCC. Methods Trop2 protein expression was detected by immunohistochemistry in HCC tissues. Cell proliferation, apoptosis, and invasion were respectively measured by CCK-8, flow cytometry, Transwell, and wound healing assays. Expression levels of epithelial–mesenchymal transition-related proteins and Trop2 protein in HCC cell lines were detected by western blotting after silencing of the TROP2 gene. Results Trop2 protein was highly expressed in HCC tissues and HCC cell lines. Trop2 mRNA and protein expression levels decreased in HepG2 and HCCLM3 cells after transfection with Trop2 siRNA. Silencing of the TROP2 gene in HepG2 and HCCLM3 cells strongly inhibited cell proliferation and migration, while enhancing cell apoptosis. Investigation of the molecular mechanism revealed that silencing of the TROP2 gene suppressed epithelial–mesenchymal transition of HepG2 and HCCLM3 cells. Conclusions The results of the present study may improve understanding of the role of Trop2 in regulation of cell proliferation and invasion, and may aid in development of novel therapy for HCC.

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