scholarly journals Decrease in Multidrug Resistance-associated Protein 2 Activities by Knockdown of Phosphatidylinositol 4-phosphate 5-kinase in Hepatocytes and Cancer Cells

2019 ◽  
Vol 22 ◽  
pp. 576-584 ◽  
Author(s):  
Atsushi Kawase ◽  
Yuta Inoue ◽  
Miho Hirosoko ◽  
Yuka Sugihara ◽  
Hiroaki Shimada ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Multidrug Resistance ◽  
Cancer Cells ◽  
Hepg2 Cells ◽  
Carcinoma Cells ◽  
Efflux Transporters ◽  
Membrane Localization ◽  
Intracellular Accumulation ◽  
Primary Hepatocytes ◽  
Phosphatidylinositol 4

Purpose: The plasma membrane localization and transport activity of multidrug resistance-associated protein 2 (MRP2/ABCC2) and P-glycoprotein (P-gp/ABCB1) efflux transporters are governed by transporter-associated proteins. Phosphatidylinositol 4,5-bisphosphate (PIP2) formed by phosphatidylinositol 4-phosphate 5-kinase type 1 (PIP5K1) activates the linker function of radixin for efflux transporters. Radixin is involved in the plasma membrane localization of efflux transporters. We examined whether PIP5K1 could be a target for the modulation of transporter activities in hepatocytes and cancer cells. Methods: The effects of PIP5K1 depletion by siRNA in mouse primary hepatocytes, PANC1 human pancreatic carcinoma cells, and HepG2 human hepatocellular carcinoma cells on the intracellular accumulation of MRP2 and P-gp substrates were examined. Results: PIP5K1A depletion resulted in increased intracellular accumulation of carboxydichlorofluorescein, a MRP2 fluorescent substrate, in mouse primary hepatocytes, PANC1 cells, and HepG2 cells. In PANC1 and HepG2 cells, the transport activities of MRP2 were significantly decreased by PIP5K1C depletion. However, the transport activities of P-gp were unchanged by PIP5K1 depletion. PIP2 levels were unchanged between control and PIP5K1A- or PIP5K1C-depleted HepG2 cells. MRP2 mRNA levels showed few changes in HepG2 cells following PIP5K1A or PIP5K1C depletion. The expression of phosphorylated radixin was decreased by PIP5K1A and PIP5K1C depletion, although total radixin levels were unchanged. Conclusions: These data suggest that PIP5K1A and PIP5K1C could be target proteins for modulating MRP2 function, partly because of the resulting changes of the linker function of radixin.

scholarly journals Sodium citrate inhibits proliferation and induces apoptosis of hepatocellular carcinoma cells

2020 ◽  
Vol 7 (3) ◽  
pp. 3659-3666
Author(s):  
Phuc Hong Vo ◽  
Sinh Truong Nguyen ◽  
Nghia Minh Do ◽  
Kiet Dinh Truong ◽  
Phuc Van Pham
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cancer Cells ◽  
Dna Fragmentation ◽  
Hepg2 Cells ◽  
Annexin V ◽  
Carcinoma Cells ◽  
Human Gastric Cancer ◽  
Apoptosis Pathway ◽  
Hepatocellular Carcinoma Cells ◽  
Activation Assay

Introduction: Cancer cells rely on glycolysis to generate energy and synthesize biomass for cell growth and proliferation (the Warburg effect). Recent studies have shown that citrate has an inhibitory effect on several cancer cells, such as human gastric cancer and ovarian cancer, by inhibiting glycolysis. In this study, we investigated the effects of citrate on the proliferation and apoptosis induction of hepatocellular carcinoma cells. Methods: HepG2 hepatocellular carcinoma cell line was used in this study. The cell proliferation was evaluated by Alamar blue assay. The apoptotic status of the HepG2 cells was recorded by Annexin V/7-AAD assay and caspase 3/7 activation assay. DNA fragmentation was evaluated by nucleus staining assay with Hoechst 33342. Results: The results showed that citrate is able to inhibit the proliferation of HepG2 cells and induce apoptosis in these cells. The initiation time of apoptosis is 4 hours after treatment with 10 mM citrate. Morphology characteristics of DNA fragmentation and broken membranes were also recorded in the apoptotic cells. Conclusion: In conclusion, our study demonstrates that citrate causes HepG2 cell death by the apoptosis pathway.

scholarly journals Effect and Mechanism of Survivin on Hypoxia-Induced Multidrug Resistance of Human Laryngeal Carcinoma Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-6
Author(s):  
Dan Xu ◽  
Da Wei Li ◽  
Jin Xie ◽  
Xin Wei Chen
Keyword(s):  
Multidrug Resistance ◽  
Rna Interference ◽  
Cancer Cells ◽  
Propidium Iodide ◽  
Laryngeal Carcinoma ◽  
Survivin Expression ◽  
Annexin V ◽  
Carcinoma Cells ◽  
Propidium Iodide Staining ◽  
Human Laryngeal Carcinoma

This study aimed at clarifying the mechanism and role of survivin in hypoxia-induced multidrug resistance (MDR) of laryngeal carcinoma cells. Human laryngeal cancer cells were incubated under hypoxia or normoxia. The expression of survivin was silenced by performing RNA interference. Additionally, by Western blot and real-time quantitative RT-PCR, survivin expression was detected. The sensitivity of human laryngeal carcinoma cells to multiple drugs was measured by CCK-8 assay. Meanwhile, the apoptosis of cells induced by cisplatin or paclitaxel was assessed by Annexin-V/propidium iodide staining analysis. Under hypoxic conditions, the upregulation of survivin was abolished by RNA interference. Then, CCK-8 analysis demonstrated that the sensitivity to multiple agents of laryngeal carcinoma cells could be increased by inhibiting survivin expression (P<0.05). Moreover, Annexin-V/propidium iodide staining analysis revealed that decreased expression of survivin could evidently increase the apoptosis rate of laryngeal carcinoma cells that were induced by cisplatin or paclitaxel evidently (P<0.05). Our data suggests that hypoxia-elicited survivin may exert a pivotal role in regulating hypoxia-induced MDR of laryngeal cancer cells by preventing the apoptosis of cells induced by chemotherapeutic drug. Thus, blocking survivin expression in human laryngeal carcinoma cells may provide an avenue for gene therapy.

Tea flavonoid theaflavin induces apoptosis in hepatocellular carcinoma cells through improving ROS level

2020 ◽  
Vol 4 (01) ◽  
pp. 1-5
Author(s):  
V. Chandravadhana ◽  
Vijay Lobo ◽  
R. Vidhyavathi ◽  
E. Mohan Raj ◽  
Arun Kumar Ramu
Keyword(s):  
Hepatocellular Carcinoma ◽  
Liver Cancer ◽  
Cancer Cells ◽  
Mtt Assay ◽  
Hepg2 Cells ◽  
Chemotherapeutic Agent ◽  
Tea Plant ◽  
Carcinoma Cells ◽  
Mitochondrial Morphology ◽  
Anticancer Effect

Flavonoids, a class of normal polyphenolic mixes, restrain cell cycle movement and instigate apoptosis. This examination was performed to explore the anticancer effect of theaflavin, a natural flavonoid found in the leaves of tea plant Camellia sinensis. Although this molecule was found to inhibit several cancer cells, the specific anticancer action in liver cancer remains unexplored, especially in human hepatocellular carcinoma (HepG2) cells. Henceforth, the present study was designed to elucidate the anticancer activity in HepG2 cells, level of reactive oxygen species (ROS) in the cancer cells and tumour cell apoptosis. The action of theaflavin in provocation apoptosis was explored through the improved ROS by MTT assay and 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA) method. As per the results obtained from the MTT assay, theaflavin had cell hindrance effect on HepG2 cells. The IC50 estimation of theaflavin to hindered cell development at 25, 50 and 75 μM concentration and instigating apoptosis through ROS improvement. The progressions in mitochondrial morphology, portion conditionally that diminished cell expansion, were seen in various concentrations of the drug treatment. In this manner, theaflavin might be useful as a chemotherapeutic agent for the treatment of liver cancer.

scholarly journals Correlations of mRNA Levels among Efflux Transporters, Transcriptional Regulators, and Scaffold Proteins in Non-Small-Cell Lung Cancer

2021 ◽  
Vol 2021 ◽  
pp. 1-6
Author(s):  
Xieyi Zhang ◽  
Wangyang Liu ◽  
Kazue Edaki ◽  
Yuta Nakazawa ◽  
Hiroki Kamioka ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Multidrug Resistance ◽  
Mrna Expression ◽  
Cancer Cells ◽  
Transcriptional Regulators ◽  
Scaffold Proteins ◽  
Lung Cancer Cells ◽  
Efflux Transporters ◽  
Expression Levels ◽  
Mrna Expression Levels

Multidrug resistance (MDR) due to enhanced drug efflux activity of tumor cells can severely impact the efficacy of antitumor therapies. We recently showed that increased activity of the efflux transporter P-glycoprotein (P-gp) associated with activation of Snail transcriptional regulators may be mediated mainly by moesin in lung cancer cells. Here, we aimed to systematically evaluate the relationships among mRNA expression levels of efflux transporters (P-gp, breast cancer resistance protein (BCRP), and multidrug resistance-associated protein 2 (MRP2)), scaffold proteins (ezrin (Ezr), radixin (Rdx), and moesin (Msn); ERM proteins), and SNAI family members (Snail, Slug, and Smac) in clinical lung cancer and noncancer samples. We found high correlations between relative (cancer/noncancer) mRNA expression levels of Snail and Msn, Msn and P-gp, Slug and MRP2, and Smuc and BCRP. These findings support our previous conclusion that Snail regulates P-gp activity via Msn and further suggest that Slug and Smuc may contribute to the functional regulation of MRP2 and BCRP, respectively, in lung cancer cells. This trial is registered with UMIN000023923.

scholarly journals Subcellular distribution of ezrin/radixin/moesin and their roles in the cell surface localization and transport function of P-glycoprotein in human colon adenocarcinoma LS180 cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0250889
Author(s):  
Takuro Kobori ◽  
Mayuka Tameishi ◽  
Chihiro Tanaka ◽  
Yoko Urashima ◽  
Tokio Obata
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Cancer Cells ◽  
Colon Adenocarcinoma ◽  
Human Colon ◽  
Membrane Localization ◽  
Transport Function ◽  
Erm Proteins ◽  
Plasma Membrane Localization ◽  
Surface Localization

The ezrin/radixin/moesin (ERM) family proteins act as linkers between the actin cytoskeleton and P-glycoprotein (P-gp) and regulate the plasma membrane localization and functionality of the latter in various cancer cells. Notably, P-gp overexpression in the plasma membrane of cancer cells is a principal factor responsible for multidrug resistance and drug-induced mutagenesis. However, it remains unknown whether the ERM proteins contribute to the plasma membrane localization and transport function of P-gp in human colorectal cancer cells in which the subcellular localization of ERM has yet to be determined. This study aimed to determine the gene expression patterns and subcellular localization of ERM and P-gp and investigate the role of ERM proteins in the plasma membrane localization and transport function of P-gp using the human colon adenocarcinoma cell line LS180. Using real-time reverse transcription polymerase chain reaction and immunofluorescence analyses, we showed higher levels of ezrin and moesin mRNAs than those of radixin mRNA in these cells and preferential distribution of all three ERM proteins on the plasma membrane. The ERM proteins were highly colocalized with P-gp. Additionally, we show that the knockdown of ezrin, but not of radixin and moesin, by RNA interference significantly decreased the cell surface expression of P-gp in LS180 cells without affecting the mRNA expression of P-gp. Furthermore, gene silencing of ezrin substantially increased the intracellular accumulation of rhodamine123, a typical P-gp substrate, with no alterations in the plasma membrane permeability of Evans blue, a passive transport marker. In conclusion, ezrin may primarily regulate the cell surface localization and transport function of P-gp as a scaffold protein without influencing the transcriptional activity of P-gp in LS180 cells. These findings should be relevant for treating colorectal cancer, which is the second leading cause of cancer-related deaths in males and females combined.

scholarly journals Notch1 Signaling Modulates Hypoxia-induced Multidrug Resistance of Human Laryngeal Cancer Cells

2021 ◽  
Author(s):  
Dawei Li ◽  
Dan Xu ◽  
Penghui Chen ◽  
Jin Xie
Keyword(s):  
Multidrug Resistance ◽  
Cancer Cells ◽  
Laryngeal Cancer ◽  
Laryngeal Carcinoma ◽  
Drug Transport ◽  
Malignant Tumors ◽  
Carcinoma Cells ◽  
Cell Counting ◽  
Apoptosis Resistance ◽  
Notch1 Signaling

Abstract Background: Laryngeal carcinoma is one of the common malignant tumors of the head and neck. Multidrug resistance (MDR) remains a critical problem in the chemotherapy for patients with laryngeal cancer. This study aims to clarify the role and mechanisms of Notch1 signaling on MDR induced by hypoxia in laryngeal cancer cells.Methods and Results: Laryngeal carcinoma cells were cultured under normoxia or hypoxia. Notch1 expression was inhibited by small interfering RNA (siRNA). The expression of Notch1, Hes1, Hey1, MDR1 and survivin mRNA was determined by Real-time PCR. The expression of Notch1, Notch1 intracellular domain (N1ICD), MDR1/P-gp and survivin protein was detected by Western blot. Current research showed that hypoxia could upregulate Notch1 expression and the activity of Notch1 signaling. Furthermore, suppression of Notch1 expression could effectively down-regulate the activity of Notch1 signaling and the expression of MDR and survivin genes in laryngeal cancer cells under hypoxia (P<0.05). Cell Counting Kit-8 (CCK-8) assay confirmed that the sensitivity of hypoxic laryngeal cancer cells to a variety of drugs could be up-regulated by suppressing Notch1 expression (P<0.05). Additionally, flow cytometry (FCM) showed that suppression of Notch1 expression significantly increased cisplatin-induced apoptosis and intracellular Rh123 (Rh123) accumulation in hypoxic laryngeal carcinoma cells (P<0.05). Conclusions: Notch1 signalling could be regarded as a pivotal regulator for mediating hypoxia-induced MDR in laryngeal cancer cells by regulating survivin-mediated apoptosis resistance and MDR1/P-gp-mediated drug transport.

scholarly journals Drug Efflux Transporters Are Overexpressed in Short-Term Tamoxifen-Induced MCF7 Breast Cancer Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Desak Gede Budi Krisnamurti ◽  
Melva Louisa ◽  
Erlia Anggraeni ◽  
Septelia Inawati Wanandi
Keyword(s):  
Breast Cancer ◽  
Multidrug Resistance ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Efflux Transporters ◽  
Drug Efflux ◽  
Cancer Resistance ◽  
P Glycoprotein ◽  
Short Period ◽  
Drug Efflux Transporters

Tamoxifen is the first line drug used in the treatment of estrogen receptor-positive (ER+) breast cancer. The development of multidrug resistance (MDR) to tamoxifen remains a major challenge in the treatment of cancer. One of the mechanisms related to MDR is decrease of drug influx via overexpression of drug efflux transporters such as P-glycoprotein (P-gp/MDR1), multidrug resistance associated protein (MRP), or BCRP (breast cancer resistance protein). We aimed to investigate whether the sensitivity of tamoxifen to the cells is maintained through the short period and whether the expressions of several drug efflux transporters have been upregulated. We exposed MCF7 breast cancer cells with tamoxifen 1 μM for 10 passages (MCF7 (T)). The result showed that MCF7 began to lose their sensitivity to tamoxifen from the second passage. MCF7 (T) also showed a significant increase in all transporters examined compared with MCF7 parent cells. The result also showed a significant increase of CC50 in MCF7 (T) compared to that in MCF7 (97.54 μM and 3.04 μM, resp.). In conclusion, we suggest that the expression of several drug efflux transporters such as P-glycoprotein, MRP2, and BCRP might be used and further studied as a marker in the development of tamoxifen resistance.

Sign in / Sign up

Export Citation Format

Share Document