Evaluation of the Consequence of CTNNB1 & RAB1A Targeting on Hepatocellular Carcinoma Cell Line

QJM ◽  
2021 ◽  
Vol 114 (Supplement_1) ◽  
Author(s):  
Amira Salah Ismail ◽  
Samar Kamal Kassim ◽  
Hanan H Shehata ◽  
Magda I Mohamad ◽  
Marian Maher Salib Roushdy
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Liver Cancer ◽  
Hepg2 Cell ◽  
Cell Lines ◽  
Hepg2 Cells ◽  
Proliferative Activity ◽  
Trypan Blue ◽  
Liver Tumors ◽  
Hepg2 Cell Lines

Abstract Background Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer in adults and the most common cause of death in people with chronic liver diseases. In Egypt, liver cancer forms 11.75% of the malignancies of all digestive organs and 1.68% of the total malignancies. HCC constitutes 70.48% of all liver tumors among Egyptians. In the past few years, early diagnosis and advances in therapeutic measures have greatly improved the outcome of HCC patients. However, the prognosis is still poor with overall survival rates of 3-5%. The alterations in cancer driver genes and associated pathways are the major triggers for HCC. So, the identification and targeting of these genes are beneficial to understand HCC and to develop a new therapy. Aim of the work We aimed to target CTNNB1 and RAB1A oncogenes in HepG2 cell lines by RNAi then evaluate the effect of their targeting on the viability and proliferative activity of HepG2 cells. Materials and methods Using HepG2 cell lines, CTNNB1 & RAB1A oncogenes were targeted using two different siRNAs (small interfering RNA) for each gene. The viability of HepG2 was conducted by Trypan blue test. The cell proliferation was tested by CellTiter 96® AQueous One Solution Cell Proliferation Assay. Results There was significant reduction in the cells’ viability detected by trypan blue test in transfected cells with siRNA targeting either CTNNB1 or RAB1A compared to Mock HepG2 cell lines (p <0.05). In addition, the proliferative activity was significantly lower in both HepG2 cell lines transfected with siRNA targeting the previous genes compared to mock cells (p < 0.05). Furthermore, HepG2 cells transfected with siRNA targeting RAB1A were more proliferative compared to those transfected with siRNA targeting CTNNB1 (p <0.05). Conclusion Targeting CTNNB1or RAB1A in HepG2 cell lines decreased the cell viability and proliferative activity. Moreover, targeting CTNNB1 was effective in decreasing cell proliferative activity compared to targeting RAB1A in HepG2 cell lines. So, targeting CTNNB1 may have a potential therapeutic effect in treatment of HCC.

scholarly journals ID: 1085 Sodium citrate induces apoptosis in HepG2 cell lines

2017 ◽  
Vol 4 (S) ◽  
pp. 174
Author(s):  
Sinh Truong Nguyen ◽  
Phuc Hong Vo ◽  
Oanh Thi-Kieu Nguyen ◽  
Nghia Minh Do ◽  
Phuc Van Pham
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cancer Cells ◽  
Dna Fragmentation ◽  
Hepg2 Cell ◽  
Cell Lines ◽  
Hepg2 Cells ◽  
Caspase 3 ◽  
Sodium Citrate ◽  
Dependent Manner ◽  
Hepg2 Cell Lines

PURPOSES: Cancer cells were observed to increase glucose uptake and fermentation of glucose to lactate to to synthesis rapidly ATP for cell growth, survival and proliferation. Thus, inhibition of glycolysis might be useful in antitumor treatment. This phenomenon occurred even with fully functioning mitochondria, and known as Warberg effect. Sodium citrate, an inhibitor of Warberg effect, was reported to antiproliferate many cancer cells line. However, sodium citrate has not been studied in Hepatocellular Carcinoma cells line yet. Here we aimed to investigate the effect of sodium citrate in HepG2 cells line.   MATERIAL AND METHODS: HepG2 cell lines was treated with sodium citrate at different concentrations. Viable cells were determined by Alamar Blue. The apoptosis induced-cells was detected by Annexin V with FCM technique. Disintegrated nuclei and DNA fragmentation was analyzed. The activity of caspase-3 was also tested.   RESULTS: We observed that the IC50 value of sodium citrate on HepG2 is at 10mM. FCM analysis showed that sodium citrate induced apoptosis in HepG2 cell line in dose-dependent manner. At 10mM sodium citrate, the caspase-3/7 was observed to be activated in time-dependent manner. Sodium citrate also induced nuclei disintergated in HepG2. DNA fragmentation was observed when HepG2 cells were treated with 10mM sodium citrate.   CONCLUSIONS: We have shown that sodium citrate possesses the antiproliferative ability on HepG2 at IC50 10mM. Sodidum citrate induces apoptosis cells in hepatocellular carcinoma HepG2 by capases-3 activation. More investigation of glycolysis inhibition of sodium citrate on HepG2 should be performed in animals

The effect of Epitranscriptomic writer (METTL3) silencing on Hepatocellular carcinoma cell line

QJM ◽  
2021 ◽  
Vol 114 (Supplement_1) ◽  
Author(s):  
Afaf R El-sherbeni ◽  
Maivel H Ghattas ◽  
Hanan H Shehata ◽  
Magda I Mohamad
Keyword(s):  
Cell Proliferation ◽  
Cell Count ◽  
Hepg2 Cell ◽  
Cell Lines ◽  
Hepg2 Cells ◽  
Viable Cell Number ◽  
Translation Control ◽  
Trypan Blue Exclusion ◽  
Hepg2 Cell Lines ◽  
Proliferative Cell

Abstract Worldwide, HCC is the sixth most common malignancy and the third most common cause of cancer-related death. RNA epigenetics becomes a hot topic in recent years. Among more than a hundred different RNA modifications, m6A is the most abundant modification. m6A is involved in regulating mRNA stability, splicing, and translation. However, the implications of m6A modification in human carcinogenesis remain poorly understood. METTL3 is a major RNA methyltransferase implicated in mRNA biogenesis, decay, and translation control through m6A modification. We aimed to target METTL3 in HepG2 cell lines by siRNA (small interfering RNA), and then evaluated the effect of this interference on viability and proliferative activity of HepG2 cells. Material and methods Using HepG2 cell lines, METTL3 was targeted using siRNA. The viability of HepG2 was conducted by Trypan blue exclusion test. The cell proliferation was tested by CellTiter 96® AQueous One Solution Cell Proliferation Assay. Results viable cell number and viability percent were significantly reduced in HepG2 cells transfected with siMETTL3 compared to mock cell lines (treated with transfection reagent only) (p<0.05). The active proliferative cell count was lower in cells transfected with siMETTL3 than mock cells (p<0.05). Conclusions Knockdown of METTL3 in HepG2 cell lines successively reduced cell viability and active proliferative cell count. METTL3 may be involved in liver tumorigenesis and its targeting may be of therapeutic benefit.

scholarly journals Kaempferol inhibits proliferation, migration, and invasion of liver cancer HepG2 cells by down-regulation of microRNA-21

2018 ◽  
Vol 32 ◽  
pp. 205873841881434 ◽  
Author(s):  
Genglong Zhu ◽  
Xialei Liu ◽  
Haijing Li ◽  
Yang Yan ◽  
Xiaopeng Hong ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Liver Cancer ◽  
Signaling Pathway ◽  
Hepg2 Cell ◽  
Cell Apoptosis ◽  
Hepg2 Cells ◽  
Mtor Signaling ◽  
Brdu Incorporation ◽  
Migration And Invasion ◽  
Mtor Signaling Pathway

Liver cancer is one of the most common and lethal cancers in human digestive system, which kills more than half a million people every year worldwide. This study aimed to investigate the effects of kaempferol, a flavonoid compound isolated from vegetables and fruits, on hepatic cancer HepG2 cell proliferation, migration, invasion, and apoptosis, as well as microRNA-21 (miR-21) expression. Cell viability was detected using cell counting kit-8 (CCK-8) assay. Cell proliferation was measured using 5-bromo-2′-deoxyuridine (BrdU) incorporation assay. Cell apoptosis was assessed using Guava Nexin assay. Cell migration and invasion were determined using two-chamber migration (invasion) assay. Cell transfection was used to change the expression of miR-21. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to analyze the expressions of miR-21 and phosphatase and tensin homologue (PTEN). Expression of key proteins involved in proliferation, apoptosis, migration, invasion, and phosphatidylinositol 3-kinase/protein kinase 3/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway were evaluated using western blotting. Results showed that kaempferol significantly inhibited HepG2 cell proliferation, migration, and invasion, and induced cell apoptosis. Kaempferol remarkably reduce the expression of miR-21 in HepG2 cells. Overexpression of miR-21 obviously reversed the effects of kaempferol on HepG2 cell proliferation, migration, invasion, and apoptosis. Moreover, miR-21 negatively regulated the expression of PTEN in HepG2 cells. Kaempferol enhanced the expression of PTEN and inactivated PI3K/AKT/mTOR signaling pathway in HepG2 cells. In conclusion, kaempferol inhibited proliferation, migration, and invasion of HepG2 cells by down-regulating miR-21 and up-regulating PTEN, as well as inactivating PI3K/AKT/mTOR signaling pathway.

Green Synthesis of Silver Nanocomposites of Nigella sativa Seeds Extract for Hepatocellular Carcinoma

2019 ◽  
Vol 4 (3) ◽  
pp. 191-200 ◽  
Author(s):  
Afreen Usmani ◽  
Anuradha Mishra ◽  
Asif Jafri ◽  
Md Arshad ◽  
Mohd Aftab Siddiqui
Keyword(s):  
Hepatocellular Carcinoma ◽  
Green Synthesis ◽  
Hepg2 Cell ◽  
Cell Lines ◽  
Nigella Sativa ◽  
Uv Spectroscopy ◽  
Silver Nanocomposites ◽  
Natural Drugs ◽  
Hepg2 Cell Lines

Background: Silver nanoparticles play a significant role in bioavailability and refining the compatibility of natural drugs in the treatment of various chronic diseases including different types of cancer. Objective: Green synthesis of silver nanocomposites of Nigella sativa seeds extract to evaluate the anticancer effects against hepatocellular carcinoma using HepG2 cell lines. Methods: The AgNCs were developed by treating aqueous extract of N. sativa seeds treated with silver nitrate (1mM) solution and were used to test its efficacy against hepatocellular carcinoma using HepG2 cell lines. Results and Discussion: The Surface Plasmon Resonance (SPR) of prepared AgNCs showed a peak at 432 nm via UV spectroscopy. The selected N. sativa AgNCs were characterized for polydispersity, surface charge and size and the results showed 0.215±0.093 polydispersity index (PDI), zeta potential 18.8±0.372 mV and size range 10-20 nm, respectively. The Fourier transform infrared spectroscopy (FTIR) also showed various peak of functional groups that are possibly involved in the reduction of silver ion and synthesized the N. sativa silver nanocomposites, respectively. N. sativa AgNCs showed 89.954% drug release while in the case of extract release, it was only 33.821% in 24 hrs. Further, in vitro studies of N. sativa AgNCs against hepatocellular carcinoma showed good cytotoxic effect p<0.05 with 7.16 µg/ml IC50 value. Conclusion: Thus, the present results revealed that green synthesis of N. sativa AgNCs can be an alternative tool for clinical application in cancer therapy; however, there is a need to find the mechanism and role of AgNCs inside the individual.

scholarly journals Deletion of low-density lipoprotein-related receptor 5 inhibits liver Cancer cell proliferation via destabilizing Nucleoporin 37

2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Jinxiao Chen ◽  
Da Wo ◽  
En Ma ◽  
Hongwei Yan ◽  
Jun Peng ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Liver Cancer ◽  
Hepg2 Cell ◽  
Cancer Cell ◽  
Hepg2 Cells ◽  
Nuclear Pore ◽  
Cancer Cell Proliferation ◽  
Liver Cancer Cell ◽  
Independent Manner ◽  
Independent Functions

Abstract Background LRP5/6 are co-receptors in Wnt/β-catenin pathway. Recently, we discovered multiple β-catenin independent functions of LRP5/6 in tumor cells and in the diseased heart. Nucleoporin 37 (NUP37) is an important component of the nuclear pore complex (NPC), whose elevated expression is associated with worsened prognosis in liver cancer. Previous studies have shown that NUP37 interacted with YAP and activated YAP/TEAD signaling in liver cancer. Our preliminary findings showed a nuclear location of LRP5. We thus tested the hypothesis that LRP5 may act as a genuine regulator of YAP/TEAD signaling via modulating NUP37 in a β-catenin-independent way. Methods We performed siRNA knockdown of LRP5, LRP6, or β-catenin in liver cancer HepG2 cells to determine the effect on tumor cell proliferation. Protein expressions and interaction between LRP5 and NUP37 were determined using immunoprecipitation and western blot analyses. Results HepG2 cell proliferation was markedly inhibited by knockdown of LRP5 but not LRP6 or β-catenin, suggesting that LRP5 has a specific, β-catenin-independent role in inhibiting HepG2 cell proliferation. Knockdown of NUP37 by siRNA inhibited the proliferation of HepG2 cells, whereas overexpression of NUP37 reversed the decrease in cell proliferation induced by LRP5 knockdown. Immunoprecipitation assays confirmed that LRP5 bound to NUP37. Furthermore, LRP5 overexpression restored NUP37 knockdown-induced downregulation of YAP/TEAD pathway. Conclusions LRP5 deletion attenuates cell proliferation via destabilization of NUP37, in a β-catenin-independent manner. LRP5 therefore acts as a genuine regulator of YAP/TEAD signaling via maintaining the integrity of the NPC, and implicates a therapeutic strategy in targeting LRP5 for inhibiting liver cancer cell proliferation.

scholarly journals MiR-196b-5p regulates the proliferation of drug-resistant hepatocellular carcinoma cell lines by activating NFκB/ABCB1 signaling pathway

2020 ◽  
Vol 19 (1) ◽  
pp. 39-44
Author(s):  
Bangming Pu ◽  
Yong Cao ◽  
Yan Li ◽  
Li Tang ◽  
Jiyi Xia ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Cell Proliferation ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Hepg2 Cells ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Drug Resistant ◽  
Reporter Assay

Purpose: To explore the molecular function of miR-196b-5p in hepatocellular carcinoma (HCC).Methods: MiR-196b-5p expression levels in HCC tissue samples were assessed by qRT-PCR. MiR-196b-5p was knocked-down or over-expressed in HepG2 cells by transfecting the cells with plasmids expressing either a miR-196b-5p inhibitor or mimic, respectively, while cell proliferation was  assessed by MTT assay. The interaction of miR-196b-5p with target molecules was confirmed using luciferase reporter assay. Cell cycle was investigated by flow cytometry, while NFκBIA expression was assessed by western blotting.Results: MiR-196b-5p was over-expressed in HCC, and miR-196b-5p expression levels in patients with HCC were related to tumor grade. MiR-196b-5p over-expression promoted cell proliferation and colony formation and suppressed cell cycle arrest and apoptosis. The results of luciferase reporter assay showed that miR-196b-5p reduced NFκBIA expression in HepG2 cells by binding to a response element in the 3′ UTR of NFκBIA. Further investigation showed that NFκBIA interacts with NFκB1 and reduces the concentration of NFκB1 in HepG2 cells. The promoter of ATP-binding cassette sub-family B member 1 (ABCB1) was also targeted and bound by NFκB1, which altered the expression of ABCB1 in HepG2 cells.Conclusion: MiR-196b-5p regulates cell proliferation in drug-resistant HCC cell lines via activation of the NFκB/ABCB1 signaling pathway. Keywords: Hepatocellular carcinoma, miR-196b-5p, NFκBIA, NFκB1, ABCB1

scholarly journals Effects of miR-489 targeting on SOX4 gene on proliferation and apoptosis of human hepatocellular carcinoma cells

2020 ◽  
Vol 20 (3) ◽  
pp. 1292-1298
Author(s):  
Bing Wang ◽  
Wang-Xun Jin ◽  
Yun-Li Zhang ◽  
Ling Huang ◽  
Hai-Bin Ni ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Cycle ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
Liver Cancer ◽  
Hepg2 Cells ◽  
Carcinoma Cells ◽  
Human Hepatocellular Carcinoma ◽  
Hepatocellular Carcinoma Cells ◽  
Human Hepatocellular Carcinoma Cells

Background: Hepatocellular carcinoma is one of the most common malignant tumors found all over the globe. Despite advances in surgery and chemotherapy, the five-year survival rate of patients with hepatocellular carcinoma is still low. It is known that the proliferation of hepatocellular carcinoma cells is closely related to the occurrence, development and prog- nosis of hepatocellular carcinoma. The present work investigates the expression of microRNA-489 (miR-489) in human hepatocellular carcinoma cells and its effect on the biological behavior of human hepatocellular carcinoma cells. Methods: The expression of miR-489 by fluorescence quantitative PCR detection in 30 patients with hepatoblastoma of liver cancer tissues and adjacent tissues was studied. Also, the determination of hepatoblastoma in four cell lines with differ- ent metastatic potential (HR8348, HCT116, HT29 and HEPG2) and the expression of miR-489 during miR-489 simulation process was studied. MTT assay, flow cytometry and Western blot analysis were performed to know the cell proliferation to detect the changes in cell cycle, apoptosis of cells, and SOX4 gene expression respectively. Results: RT-PCR results showed that the cells compared with pre-cancerous tissue, the expression level of miR-489 in hepatocellular carcinoma tissues than in adjacent tissue significantly decreased (P<0.05), and with liver cancer cell metastasis increased (P<0.05); analogue transfection constructed miR-489 overexpressing HEPG2 cell line by microRNA. MTT results showed that miR-489 can inhibit the proliferation of HEPG2 cells, the differences were statistically significant (P<0.05); flow cytometry results showed that miR-489 mimics was transfected into HEPG2 cells at 48 hours had no significant effect on cell cycle distribution (P > 0.05); but miR-489 expression could induce apoptosis, compared with the control group, the apoptosis of miR-489 mimics was significantly increased and the difference was statistically significant (P < 0.05). Conclusion: In conclusion, miR-489 can significantly inhibit the occurrence and development of hepatocellular carcinoma cells. The mechanism may be down regulated by the expression of SOX4 and inhibit cell proliferation. Further this study showed that the tumor cells SOX4 gene as a regulatory factor target the genes of miR-489 in hepatocellular carcinoma. Keywords: Hepatocellular carcinoma; mircroRNA-489; SOX4; apoptosis.

Effect of the BBC3 Gene on the Proliferation and Apoptosis of Hepatocellular Carcinoma Cells Through p53-Regulated Signaling

2021 ◽  
Vol 11 (1) ◽  
pp. 135-141
Author(s):  
Sheng Zheng ◽  
Hua Yang ◽  
Yefei Chang ◽  
Dan Zhao ◽  
Juan Yang
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Proliferation ◽  
Hepg2 Cell ◽  
Hepg2 Cells ◽  
Caspase 3 ◽  
Carcinoma Cells ◽  
Caspase 9 ◽  
Hepatocellular Carcinoma Cells ◽  
P53 Signaling ◽  
Normal Human Liver

We examined the effect of the BBC3 gene on hyperplasia and apoptosis in HepG2 cells and its underlying mechanism. Quantitative RT-PCR was used to determine the level of BBC3 expression in HL-7702 normal human liver cells and four different hepatocellular carcinoma cell lines (HepG2, HuH-7, HCCLM3 and MHCC97H). Transfection was performed with Lipofectamine 2000 reagent and the transfectants were divided into three groups: pcDNA-BBC3 group (transfected BBC3 over-expressing plasmid), pcDNA-NC group (transfected empty plasmid), and a Ctrl group (not transfected). Quantitative RT-PCR and western blot analysis were used to measure BBC3 expression. The CCK-8 assay was used to determine the effect of BBC3 on HepG2 cell proliferation. Flow cytometry was used for testing the effect of overexpressing BBC3 on apoptosis in HepG2 cells. The levels of cleaved-Caspase-3 (C-Caspase-3), cleaved-Caspase-9 (C-Caspase-9), and proteins associated with the p53 signaling pathway were assessed by western blot analysis. The level of BBC3 mRNA in HL-7702 normal human liver cells was significantly higher compared with that in human hepatocellular carcinoma cells including HepG2, HuH-7, HCCLM3 and MHCC97H (P < 0.05). The lowest level of BBC3 mRNA was observed in HepG2 cells. The level of BBC3 mRNA and protein in HepG2 cells were significantly higher compared with that of the pcDNA-NC group following transfection with a BBC3 overexpressing plasmid. HepG2 cell proliferation in the pcDNA-NC group was higher compared with that of the pcDNA-BBC3-transfected group (P < 0.05). The apoptotic rate and levels of cleaved-Caspase-3, cleaved-Caspase-9, p53, phospho-p53, and p21 protein in cells were higher compared with that of the pcDNA-NC group. No change was observed in the pcDNA-NC and Ctrl groups. The BBC3 gene was down-regulated in hepatocellular carcinoma cells. HepG2 cell proliferation can be inhibited and HepG2 cell apoptosis can be induced by the overexpression of BBC3 through activation of the p53 signaling pathway.

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