scholarly journals Silibinin sensitizes CD133+ hepatocellular carcinoma cells to cisplatin treatment through suppression of OPA1

2020 ◽  
Author(s):  
Jikang Yang ◽  
Zhiyuan Xing
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
Cytochrome C ◽  
Carcinoma Cells ◽  
Flow Cytometry Analysis ◽  
Western Blot Assay ◽  
Huh7 Cells ◽  
Mechanism Research ◽  
Induced Apoptosis ◽  
Hcc Cells

Abstract Background: Drug resistance is still a major obstacle during the cisplatin-based chemotherapy of hepatocellular carcinoma (HCC). Recently, studies have indicated that the population of CD133+ cancer cells is partially responsible for the failure of cancer treatment. However, the potential mechanisms are still unclear.Methods: CD133+ HepG2 and Huh7 cells were sorted via flow cytometry. CCK-8 assay was used to detect the cytotoxicity of cisplatin and silibinin against HCC cells. Western blot assay was performed to detect the protein expression, cleavage of caspases and release of cytochrome c from mitochondria into cytosol. Flow cytometry analysis was used to measure the apoptotic rate of CD133+ HepG2 and Huh7 cells.Results: CD133+ HepG2 and Huh7 cells were observed to exhibit obvious resistance against cisplatin. However, co-treatment with silibinin significantly reduced the cisplatin resistance of CD133+ HepG2 and Huh7 cells. Furthermore, although CD133+ HepG2 and Huh7 cells were resistant to cisplatin-induced apoptosis, co-treatment with silibinin enhanced the cisplatin-induced apoptosis through promoting the release of cytochrome c from mitochondria into cytosol. In the mechanism research, we proved that silibinin inhibited the expression of OPA1 in CD133+ HepG2 and Huh7 cells. Under the stress of cisplatin, silibinin promoted the collapse of mitochondria and increased the release of cytochrome c. As a result, caspases-dependent apoptosis was induced in CD133+ HepG2 and Huh7 cells which were co-treated with cisplatin and silibinin.Conclusion: Silibinin sensitizes CD133+ HCC cells to cisplatin-induced apoptosis through suppression of OPA1.

scholarly journals Intracellular alpha-fetoprotein interferes with all-trans retinoic acid induced ATG7 expression and autophagy in hepatocellular carcinoma cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shanshan Wang ◽  
Rilu Feng ◽  
Ying Shi ◽  
Dexi Chen ◽  
Honglei Weng ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Retinoic Acid ◽  
Carcinoma Cells ◽  
Alpha Fetoprotein ◽  
Cell Counting ◽  
All Trans Retinoic Acid ◽  
Induced Apoptosis ◽  
Anti Tumor Activity ◽  
Retinoid Acid ◽  
Hcc Cells

AbstractRetinoic acid and retinoid acid receptor (RA-RAR) signaling exhibits suppressive functions in the progression of hepatocellular carcinoma (HCC) through multiple mechanisms. However, whether RA-RAR signaling induces autophagy that contributes its anti-tumor activity in HCC remains elusive. In the current study, the effects of RA-RAR pathway on autophagy were investigated in two HCC cell lines: alpha-fetoprotein (AFP) positive PLC/PRF/5 and AFP negative HLE cells. Cell autophagy was analyzed with western blot for detection of LC3 conversion and p62/SQSTM1 degradation while autophagy flux was assayed using the mRFP-GFP-LC3 reporter. Cell apoptosis and viability were analyzed by caspase-3 activity, TdT-mediated dUTP nick end labeling (TUNEL) assay, and Cell Counting Kit (CCK)-8, respectively. Chromatin immunoprecipitation (ChIP) was employed to detect the binding of RAR onto the promoter of autophagy-relevant 7 (ATG7), and co-immunoprecipitation (CoIP) was used to analyze the interaction of AFP and RAR. The results showed that ATRA dosage and time-dependently induced high levels of cell autophagy in both the PLC/PRF/5 and HLE cells, which was accompanied with up-regulation of ATG7. ChIP assay showed that RAR was able to bind to its responsive elements on ATG7 promoter. Impairment of ATG7 induction or blockade of autophagy with chloroquine aggravated ATRA induced apoptosis of HCC cells. Furthermore, intracellular AFP was able to complex with RAR in PLC/PRF/5 cells. Knockdown of AFP in PLC/PRF/5 cells augmented the up-regulation of ATG7 by ATRA while overexpression of AFP in HLE cells attenuated ATRA induced ATG7 expression and autophagy. Thus, ATRA induced ATG7 and autophagy participated in its cytotoxicity on HCC cells and AFP interfere with the induction of ATG7 and autophagy through forming complex with RAR.

scholarly journals Cisplatin Increased the Expression of PD-1 and PD-L1 by YAP1 in Hepatocellular Carcinoma

2021 ◽  
Author(s):  
Liyuan Hao ◽  
Yinglin Guo ◽  
Qing Peng ◽  
Zhiqin Zhang ◽  
Shenghao Li ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
T Cells ◽  
Cancer Cells ◽  
Flow Cytometry Analysis ◽  
Chemotherapy Drugs ◽  
The World ◽  
Hcc Cells

Abstract Hepatocellular carcinoma (HCC) was one of the most malignant cancers in the world. Cisplatin (DDP) was one of the main chemotherapy drugs for HCC, but the mechanism of DDP treatment for HCC remains unclear. In this presentation, we found that DDP inhibited the growth of HCC cells and promoted the expression of PD-1 and its ligand PD-L1 in cancer cells. Meanwhile, flow cytometry analysis revealed that DDP enhanced PD-1-CD8+ T cells expression and decreased PD-1+CD8+ T cells expression. ELISA analysis suggested that DDP decreased TGF-β expression in vivo. Therefore, the study indicated that DDP enhanced PD-1 and PD-L1 expression and inhibited the growth of HCC.

scholarly journals Aurora-a confers radioresistance in human hepatocellular carcinoma by activating NF-κB signaling pathway

BMC Cancer ◽  
2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Ze-Tian Shen ◽  
Ying Chen ◽  
Gui-Chun Huang ◽  
Xi-Xu Zhu ◽  
Rui Wang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Western Blot ◽  
Luciferase Reporter ◽  
Aurora A ◽  
Western Blot Assay ◽  
Induced Apoptosis ◽  
Mtt Assays ◽  
Hcc Cells

Abstract Background Radiotherapy failure is a significant clinical challenge due to the development of resistance in the course of treatment. Therefore, it is necessary to further study the radiation resistance mechanism of HCC. In our early study, we have showed that the expression of Aurora-A mRNA was upregulated in HCC tissue samples or cells, and Aurora-A promoted the malignant phenotype of HCC cells. However, the effect of Aurora-A on the development of HCC radioresistance is not well known. Methods In this study, colony formation assay, MTT assays, flow cytometry assays, RT-PCR assays, Western blot, and tumor xenografts experiments were used to identify Aurora-A promotes the radioresistance of HCC cells by decreasing IR-induced apoptosis in vitro and in vivo. Dual-luciferase reporter assay, MTT assays, flow cytometry assays, and Western blot assay were performed to show the interactions of Aurora-A and NF-κB. Results We established radioresistance HCC cell lines (HepG2-R) and found that Aurora-A was significantly upregulated in those radioresistant HCC cells in comparison with their parental HCC cells. Knockdown of Aurora-A increased radiosensitivity of radioresistant HCC cells both in vivo and in vitro by enhancing irradiation-induced apoptosis, while upregulation of Aurora-A decreased radiosensitivity by reducing irradiation-induced apoptosis of parental cells. In addition, we have showed that Aurora-A could promote the expression of nuclear IkappaB-alpha (IκBα) protein while enhancing the activity of NF-kappaB (κB), thereby promoted expression of NF-κB pathway downstream effectors, including proteins (Mcl-1, Bcl-2, PARP, and caspase-3), all of which are associated with apoptosis. Conclusions Aurora-A reduces radiotherapy-induced apoptosis by activating NF-κB signaling, thereby contributing to HCC radioresistance. Our results provided the first evidence that Aurora-A was essential for radioresistance in HCC and targeting this molecular would be a potential strategy for radiosensitization in HCC.

scholarly journals Notch1 Signaling Sensitizes Tumor Necrosis Factor-related Apoptosis-inducing Ligand-induced Apoptosis in Human Hepatocellular Carcinoma Cells by Inhibiting Akt/Hdm2-mediated p53 Degradation and Up-regulating p53-dependent DR5 Expression

2009 ◽  
Vol 284 (24) ◽  
pp. 16183-16190 ◽  
Author(s):  
Chunmei Wang ◽  
Runzi Qi ◽  
Nan Li ◽  
Zhengxin Wang ◽  
Huazhang An ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Tumor Necrosis Factor ◽  
Notch Signaling ◽  
Tumor Necrosis ◽  
Carcinoma Cells ◽  
Hepatocellular Carcinoma Cells ◽  
Notch1 Signaling ◽  
Induced Apoptosis ◽  
Necrosis Factor ◽  
Hcc Cells

Notch signaling plays a critical role in regulating cell proliferation, differentiation, and apoptosis. Our previous study showed that overexpression of Notch1 could inhibit human hepatocellular carcinoma (HCC) cell growth by arresting the cell cycle and inducing apoptosis. HCC cells are resistant to apoptotic induction by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), so new therapeutic approaches have been explored to sensitize HCC cells to TRAIL-induced apoptosis. We are wondering whether and how Notch1 signaling can enhance the sensitivity of HCC cells to TRAIL-induced apoptosis. In this study, we found that overexpression of ICN, the constitutive activated form of Notch1, up-regulated p53 protein expression in HCC cells by inhibiting proteasome degradation. p53 up-regulation was further observed in human primary hepatocellular carcinoma cells after activation of Notch signaling. Inhibition of the Akt/Hdm2 pathway by Notch1 signaling was responsible for the suppression of p53 proteasomal degradation, thus contributing to the Notch1 signaling-mediated up-regulation of p53 expression. Accordingly, Notch1 signaling could make HCC cells more sensitive to TRAIL-induced apoptosis, whereas Notch1 signaling lost the synergistic promotion of TRAIL-induced apoptosis in p53-silenced HepG2 HCC cells and p53-defective Hep3B HCC cells. The data suggest that enhancement of TRAIL-induced apoptosis by Notch1 signaling is dependent upon p53 up-regulation. Furthermore, Notch1 signaling could enhance DR5 expression in a p53-dependent manner. Taken together, Notch1 signaling sensitizes TRAIL-induced apoptosis in HCC cells by inhibiting Akt/Hdm2-mediated p53 degradation and up-regulating p53-dependent DR5 expression. Thus, our results suggest that activation of Notch1 signaling may be a promising approach to improve the therapeutic efficacy of TRAIL-resistant HCC.

scholarly journals Luteolin Promotes Cell Apoptosis by Inducing Autophagy in Hepatocellular Carcinoma

2017 ◽  
Vol 43 (5) ◽  
pp. 1803-1812 ◽  
Author(s):  
Zhijia Cao ◽  
Huainian Zhang ◽  
Xiaoyan Cai ◽  
Wei Fang ◽  
Dong Chai ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
Western Blotting ◽  
Cell Apoptosis ◽  
Flow Cytometry Analysis ◽  
Hoechst 33342 ◽  
Qrt Pcr ◽  
Induced Apoptosis ◽  
Luteolin Treatment

Background/Aims: Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and is a leading cause of cancer-related death worldwide. Luteolin, a flavonoid from traditional Chinese medicine, shows anti-cancer activity in many cancer cells, including HCC. However, the mechanism underlying the action of luteolin in HCC, especially its role in regulating cell autophagy, remains unclear. In the present study, we investigated the role of luteolin in regulating cell autophagy and the role of autophagy in luteolin-induced apoptosis. Methods: The 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay (MTT) was used to investigate cell viability. Flow cytometry analysis was used to detect the cell cycle and cell apoptosis. Hoechst 33342 staining was used to detect cell apoptosis. Transmission electron microscopy was used to investigate autophagy. qRT-PCR and western blotting were used to detect apoptosis- and autophagy-related mRNAs and proteins. Results: Luteolin reduced the viability of SMMC-7721 cells in a time and dose-dependent manner, and induced significant G0/G1-phase arrest. In addition, the results of flow cytometry analysis and Hoechst 33342 staining showed that luteolin treatment increased the number of apoptotic cells obviously, and the results of qRT-PCR and western blotting showed that luteolin treatment increased caspase 8 and decreased bcl-2 at the mRNA and protein levels. Furthermore, luteolin increased the number of intracellular autophagosomes, promoted LC3B-I conversion to LC3B-II, and increased Beclin 1 expression. Finally, co-treatment with the autophagy inhibitor chloroquine weakened the effects of luteolin on cell apoptosis. Conclusion: Luteolin induced apoptosis in human liver cancer SMMC-7721 cells, partially via autophagy. Thus, luteolin could be used as a regulator of autophagy in HCC treatment.

scholarly journals Fibrinogen-Like Protein 1 Modulates Sorafenib Resistance in Human Hepatocellular Carcinoma Cells

2021 ◽  
Vol 22 (10) ◽  
pp. 5330
Author(s):  
Yeonghoon Son ◽  
Na-Rae Shin ◽  
Sung-Ho Kim ◽  
Su-Cheol Park ◽  
Hae-June Lee
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
Colony Formation ◽  
Anticancer Therapy ◽  
Carcinoma Cells ◽  
Hep3b Cells ◽  
Human Hepatocellular Carcinoma Cells ◽  
Systemic Drug ◽  
Higher Sensitivity ◽  
Hcc Cells

Despite liver cancer being the second-leading cause of cancer-related death worldwide, few systemic drugs have been approved. Sorafenib, the first FDA-approved systemic drug for unresectable hepatocellular carcinoma (HCC), is limited by resistance. However, the precise mechanisms underlying this phenomenon are unknown. Since fibrinogen-like 1 (FGL1) is involved in HCC progression and upregulated after anticancer therapy, we investigated its role in regulating sorafenib resistance in HCC. FGL1 expression was assessed in six HCC cell lines (HepG2, Huh7, Hep3B, SNU387, SNU449, and SNU475) using western blotting. Correlations between FGL1 expression and sorafenib resistance were examined by cell viability, colony formation, and flow cytometry assays. FGL1 was knocked-down to confirm its effects on sorafenib resistance. FGL1 expression was higher in HepG2, Huh7, and Hep3B cells than in SNU387, SNU449, and SNU475 cells; high FGL1-expressing HCC cells showed a lower IC50 and higher sensitivity to sorafenib. In Huh7 and Hep3B cells, FGL1 knockdown significantly increased colony formation by 61% (p = 0.0013) and 99% (p = 0.0002), respectively, compared to that in controls and abolished sorafenib-induced suppression of colony formation, possibly by modulating ERK and autophagy signals. Our findings demonstrate that sorafenib resistance mediated by FGL1 in HCC cells, suggesting FGL1 as a potential sorafenib-resistance biomarker and target for HCC therapy.

scholarly journals UNDERSTANDING THE MECHANISM OF DRUG-RESISTANT AND TUMOR RECURRENCE IN LIVER CANCER

2018 ◽  
Vol 8 (5) ◽  
pp. 224-229
Author(s):  
Vasanthakumar Sekar ◽  
Muthu Dhandapani ◽  
Babu Balakrishnan ◽  
Kaleeswaran Balasubramanian ◽  
Sivalingam Azhagu Madhavan ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
Liver Cancer ◽  
Tumor Recurrence ◽  
Morphological Analysis ◽  
Multi Drug Resistance ◽  
Flow Cytometry Analysis ◽  
Spheroid Formation ◽  
Entire Study ◽  
Huh7 Cells

Chemo-resistant and tumor recurrence are the major hurdle to overcome the cancer patients. Especially in hepatocellular carcinoma (HCC) is notoriously refractory to chemotherapy because of its tendency to develop multi-drug resistance (MDR), through various mechanisms. Aim: The current research is focussed on understanding the mechanism involved in chemo-resistant and tumor recurrence in liver cancer. Methods: Human hepatocellular carcinoma cell line (Huh7) was used entire study. Huh7 cells were cultured with known chemotherapeutic drugs such as 5-FU, Paclitaxel and Cisplatin-based on their Cmax concentration, and then these drug-treated cells were examined for chemoresistant and tumor recurrence properties through flow cytometry analysis, spheroid formation assay, and morphological analysis. Results: In morphological analysis confirm these all the chemo drugs were shown more cytotoxic effete than control, even though there were few viable cells noticed in cisplatin treatment. In flow cytometry analysis cisplatin pre-treated cells were well expressed LCSC marker such as CD133 and stem cell transduction factors like Oct-4 & Nanog than control. In addition to this, all the CD133 expressed cells also expressed to EpCAM. In spheroid formation assay, cisplatin pre-treated cells shown well-defined spheroid than control. Conclusion: LCSC plays a major role in chemoresistant and tumor recurrence through PI3K/Akt/mTOR, wnt-β catenin signaling, NF-kB signaling. So, targeting LCSC through EpCAM targeted therapy along with chemotherapy might be the better option for enhanced prognosis. Keywords: LCSC, Chemoresistant, Tumor recurrence, Hepatocellular carcinoma.

Combination of Cinobufacini and Doxorubicin Increases Apoptosis of Hepatocellular Carcinoma Cells through the Fas- and Mitochondria-Mediated Pathways

2017 ◽  
Vol 45 (07) ◽  
pp. 1537-1556 ◽  
Author(s):  
Jufeng Xia ◽  
Yoshinori Inagaki ◽  
Jianjun Gao ◽  
Fanghua Qi ◽  
Peipei Song ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Chinese Medicine ◽  
Traditional Chinese Medicine ◽  
Cytochrome C ◽  
Cancer Treatment ◽  
Carcinoma Cells ◽  
Combination Group ◽  
New Strategy ◽  
Apoptotic Pathways ◽  
Hcc Cells

Cinobufacini, a traditional Chinese medicine, has been used widely for cancer treatment, such as hepatocellular carcinoma (HCC), sarcoma, and leukemia. Previous studies done by our lab indicated that cinobufacini could suppress HCC cells through mitochondria-mediated and Fas-mediated apoptotic pathways. Here, we use a combination of cinobufacini and doxorubicin to inhibit the growth of HCC cells. The combination group induced more significant apoptosis by affecting proteins and RNA of apoptosis-related elements, such as Bcl-2, Bax, Bid, and cytochrome c. Furthermore, cinobufacini, as a mixture of a number of components, had stronger apoptosis-inducing activity than particular individual components or a simple mixture of a few components. Overall, these results suggested that the combination of cinobufacini and doxorubicin may provide a new strategy for inhibiting the proliferation of HCC cells.

scholarly journals Cytotoxicity of Human Hepatic Intrasinusoidal CD56bright Natural Killer Cells against Hepatocellular Carcinoma Cells

2019 ◽  
Vol 20 (7) ◽  
pp. 1564 ◽  
Author(s):  
Shin Hwang ◽  
Jaeseok Han ◽  
Ji-Seok Baek ◽  
Eunyoung Tak ◽  
Gi-Won Song ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Nk Cells ◽  
Natural Killer ◽  
Fas Ligand ◽  
Nk Cell ◽  
Carcinoma Cells ◽  
Huh7 Cells ◽  
Ifn Γ ◽  
And Function ◽  
Hcc Cells

Hepatic intrasinusoidal (HI) natural killer (NK) cells from liver perfusate have unique features that are similar to those of liver-resident NK cells. Previously, we have reported that HI CD56bright NK cells effectively degranulate against SNU398 hepatocellular carcinoma (HCC) cells. Thus, the aim of this study was to further investigate the phenotype and function of HI NK cells. We found that HI CD56bright NK cells degranulated much less to Huh7 cells. HI CD56bright NK cells expressed NKG2D, NKp46, TNF-related apoptosis-inducing ligand (TRAIL), and FAS ligand (FASL) at higher levels than CD56dim cells. SNU398 cells expressed more NKG2D ligands and FAS and less PD-L1 than Huh7 cells. Blockade of NKG2D, TRAIL, and FASL significantly reduced the cytotoxicity of HI NK cells against SNU398 cells, but blockade of PD-L1 did not lead to any significant change. However, HI NK cells produced IFN-γ well in response to Huh7 cells. In conclusion, the cytotoxicity of HI CD56bright NK cells was attributed to the expression of NKG2D, TRAIL, and FASL. The results suggest the possible use of HI NK cells for cancer immunotherapy and prescreening of HCC cells to help identify the most effective NK cell therapy recipients.

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