Bioinspired bimetallic nanozyme with cabozantinib and paclitaxel nanoparticles for effective ROS accumulation for improved ferroptosis in liver cancer cells

Expression Pattern, Regulation, and Functions of Methionine Adenosyltransferase 2β Splicing Variants in Hepatoma Cells

Visfatin is found in adipose tissue and is referred to as nicotinamide phosphoribosyltransferase (Nampt). Visfatin has anti-apoptotic, proliferative, and metastatic properties and may mediate its effects via ERK and PI3K/Akt signaling. Studies have yet to determine whether inhibition of kinase signaling will suppress visfatin-induced liver cancer. The purpose of this study was to determine which signaling pathways visfatin may promote liver cancer progression. HepG2 and SNU-449 liver cancer cells were exposed to visfatin with or without ERK or PI3K/Akt inhibitor, or both inhibitors combined. These processes that were assessed: proliferation, reactive oxygen species (ROS), lipogenesis, invasion, and matrix metalloproteinase (MMP). Inhibition of PI3K/Akt and combination of inhibitors suppressed visfatin-induced viability. ERK inhibition in HepG2 cells decreased visfatin-induced proliferation. ERK inhibitor alone or in combination with PI3K inhibitors effectively suppressed MMP-9 secretion and invasion in liver cancer cells. PI3K and ERK inhibition and PI3K inhibition alone blocked visfatin's ROS production in SNU-449cells. These results corresponded with a decrease in phosphorylated Akt and ERK, β-catenin, and fatty acid synthase. Akt and ERK inhibition differentially regulated physiological changes in liver cancer cells. Inhibition of Akt and ERK signaling pathways suppressed visfatin-induced invasion, viability, MMP-9 activation, and ROS production.

Background Obesity, a major public health concern, increases the risk of developing liver cancer which is the leading cause of cancer-related deaths worldwide. Obesity is associated with increased adiposity and macrophage infiltration both of which promote secretion of adipokines and cytokines in the tumor microenvironment. Specifically, visfatin and resistin have been detected at higher levels in the serum of obese individuals and liver tumors. However, the contribution of these adipocytokines in the progression of liver cancer remains unclear. Materials and methods The objective of this study was to characterize the effects of visfatin and resistin on HepG2, SNU-449 and HuH7 liver cancer cells. Cells exposed to visfatin and resistin were analyzed for fatty acid synthase protein, and phosphorylation of Akt and ERK tumorigenic signaling pathways, cell viability, lipogenesis, reactive oxygen species (ROS), matrix metallopeptidase 9 (MMP-9) enzyme activity and invasion. Results HepG2, SNU-449, and HuH7 liver cancer cells treated with visfatin and resistin increased cell viability, invasion, FASN protein, and Akt and ERK phosphorylation. Visfatin and resistin selectively increased ROS production in HepG2 and SNU-449 cells while there was no statistical difference in HuH7 cells. Visfatin and resistin stimulated lipogenesis in HepG2 cells while visfatin increased lipogenesis in SNU-449 cells, and visfatin nor resistin had an effect on lipogenesis in HuH7 cells. Lastly, visfatin and resistin increased MMP-9 enzyme activity in HepG2 and HuH-7 cells but only visfatin increased MMP-9 activity in SNU-449 cells. Conclusions Future studies are needed to determine if inhibition of ERK and Akt suppresses the visfatin and resistin-induced invasive liver cancer phenotype.

Repressing of NHERF1 inhibits liver cancer progression by promoting the production of ROS

Role of Phagocyte Oxidase in UVA-Induced Oxidative Stress and Apoptosis in Keratinocytes

Mansorin-A and its naphthoquinone derivative, mansonone-G are naturally occurring compounds isolated from the trunk of Mansonia gagei, family Sterculariaceae. In a previous work, they showed potential anticancer effects besides their antioxidant properties. Herein, we investigated their potential chemomodulatory effects to 5-fluorouracil (5FU) against two liver cancer cell lines (Huh7 and HepG2). Despite the significant cell killing effect of 5FU to Huh7 and HepG2 cells (IC50’s of 2.6±0.2 µM and 0.82 ± 0.2 µM, respectively), the resistance fractions of both cell lines were considerably high (53.2±1.7% and 39.4±2.7%, respectively). Mansorin-A and mansonone-G induced moderate cytotoxicity against both liver cancer cell lines with IC50’s ranging from 25.8±3.2 µM to 36.3±2.7 µM. In Huh7, mansorin-A and mansonone-G synergized cell killing effect of 5FU and decreased its IC50’s to 0.9±0.1 and 1.0±2.1 µM, respectively. On the other hand, mansorin-A and mansonone-G showed apparent antagonism by combination with 5FU (CI values of 6.2 and 9.4, respectively) against HepG2 cells. However, both compounds significantly abolished the resistant fraction of HepG2 to 5FU (3.3±0.3% and 8.4 ± 2.6%, respectively). Further investigation showed that mansonone-G alone showed antiproliferative effect and arrested cells in G0/G1-phase. Both mansorin-A and mansonone-G enhanced 5FU induced cell cycle arrest at G0/G1-phase with reciprocal decrease in cells entering S-phase and G2/M-phase. In addition, apoptosis due to 5FU treatment alone and in combination with mansorin-A and mansonone-G was assessed using annexin-V/FITC staining coupled with flowcytometry. Yet, both mansorin-A and mansonone-G enhanced 5FU induced apoptotic effect against liver cancer cells. Mansonone-G alone induced significant apoptosis in both HepG2 and Huh7 cells. Besides apoptosis, autophagy induction properties of mansorin-A and mansonone-G were assessed using flowcytometry after acridine orange staining. Mansonone-G aborted autophagy in both HepG2 and Huh7 cell lines and indirectly forced liver cancer cells to undergo apoptotic cell death. On the other hand, both mansorin-A and mansonone-G significantly induced the production of reactive oxygen species within both HepG2 and Huh-7 cells which might directly induce apoptosis in both cell lines. In conclusion, mansonone-G and to a lesser extent mansorin-A, potentiated the anticancer properties of 5FU against liver cancer cells via potentiating its cell cycle arrest and potentiating its apoptotic influence due to ROS production. Citation Format: Hanadi G. Aljohani, Gehan A. Hegazy, Ali M. El-Halawany, Aliaa A. Alamoudi, Ghada M.A. Agabnoor, Ahmed M. Al-Abd. Combination analysis for the potential chemomodulatory effects of mansorin-A and its naphthoquinone derivative (mansonone-G) to 5-fluorouracil against liver cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 305.

The sustained synthesis of gold nanoparticles (GNPs) has gained significant attention in biomedical applications. In this study, we explored the antibacterial and anticancer potential of bakuchiol-mediated gold nanoparticles (Bak-GNPs). Bakuchiol, a natural compound found in Psoralea corylifolia seeds, serves as both a reducing and stabilizing agent for green synthesis of GNPs. Our objectives include network analysis, molecular docking, synthesis of GNPs, characterization, and antipathogenic and anticancer efficacy of Bak-GNPs against lung and liver cancers. Protein-protein interaction networks were analyzed to identify effective protein targets for bakuchiol in lung and liver cancers. A molecular docking study was performed to validate the efficacy of the target protein against lung and liver cancer. Furthermore, Bak-GNPs were synthesized using bakuchiol and characterized by various techniques such as UV-visible spectroscopy, dynamic light scattering (DLS), zeta potential transmission electron microscopy (TEM), and Fourier-transform infrared (FTIR) spectroscopy, and their potential against pathogens and lung and liver cancers. GNAI3 emerged as the most promising target, with a binding energy of -7.5 kcal/mol compared to PTGER3's -6.9 kcal/mol, different characterization techniques revealed the successful synthesis of Bak-GNPs. Bak-GNPs exhibited potent antibacterial activity against both Gram-positive and Gram-negative bacteria, as confirmed by minimum inhibitory concentration (MIC) values. Bak-GNPs demonstrated significant anticancer effects on A549 (lung cancer) and HepG2 (liver cancer) cells, with IC50 values of 11.19 μg/mL and 6.6 μg/mL, respectively. Induction of apoptosis and inhibition of cell proliferation were observed in both the cell lines. The increased production of reactive oxygen species (ROS) contributes to its anticancer effects. This study highlights promising biomedical applications of bakuchiol-mediated GNPs. This green synthesis approach using bakuchiol provides a sustainable method for producing nanoparticles with enhanced biological activities. Further exploration of the pharmacological properties and mechanisms of Bak-GNPs is required to optimize their therapeutic efficacy for clinical use.

Objective Liver cancer (LC), one of the familiar malignancies, has a very high morbidity all over the world. The onset of the disease is hidden, and the patients usually do not express any special symptoms. Most of them will have been developed to the middle and later stage when they are diagnosed. This is one of the main reasons why the prognosis of LC is extremely pessimistic all the year round. Recently, researchers have focused mainly on molecular studies, among which LncRNA is a hot spot. This research aims to explore the biological behaviors of LncRNA NKILA and miR-485-5p in LC cells and verify the relationship between them, thereby providing a new theoretical basis for future prevention and treatment. Methods Ninety-four early LC patients admitted to our hospital from January 2015 to January 2017 were regarded as the research objects. In addition, human LC cells SMMC-7721, HepG2, and normal liver cells HL-7702 were purchased. The LncRNA NKILA and miR-485-5p level in cancer and adjacent tissues, LC, and normal liver cells of patients was tested by PCR. Patients were followed up for 3 years. Then, LncRNA NKILA and miR-485-5p's effects on prognosis and cell biological behavior were analyzed. At last, the relationship between LncRNA NKILA and miR-485-5p was assessed by a dual-luciferase reporter assay. Results The LncRNA NKILA expression was high in LC tissues and cells (P < 0.050), while miR-485-5p was low compared with the normal adjacent tissues (P < 0.050). Prognostic follow-up manifested that high LncRNA NKILA or low miR-485-5p could predict the poor prognosis and high mortality risk of the patients (P < 0.050). LC cells with downregulated LncRNA NKILA documented inhibited proliferation, invasion, and EMT, while the apoptosis level of the cells increased (P < 0.050). The proliferation, invasion, and EMT were inhibited by miR-485-5p increase, while the apoptosis of the cells decreased after upregulating miR-485-5p (P < 0.050). Online websites predicted that LncRNA NKILA had a binding site with miR-485-5p, and dual-luciferase reporter assay confirmed that LncRNA NKILA could directly target with miR-485-5p (P < 0.050). The miR-485-5p in LC cells increased after LncRNA NKILA was silenced (P < 0.050). The rescue experiment documented that LncRNA NKILA inhibition on LC cells was reversed by inhibiting miR-485-5p (P < 0.050). Conclusion The LncRNA NKILA with high expression advances LC cell proliferation, invasion, and EMT by targeting miR-485-5p.

Citrate synthase (CS) is a key rate-limiting enzyme in the tricarboxylic acid (TCA) cycle and plays a crucial role in cancer progression. However, the mechanism by which CS promotes liver cancer growth remains unclear. The aim of this study is to elucidate the role of CS and its post-translational modifications (PTMs) in the initiation and progression of hepatocellular carcinoma (HCC). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to detect protein lysine succinylation in human liver cancer and adjacent non-cancerous tissues. A HCC model was established in male C57BL/6 mice through intraperitoneal injection of DEN. The expression of SIRT5 and CS in HCC mice was assessed by RT-qPCR, immunohistochemistry, and Western blotting. HepG2 cells were cultured, and co-immunoprecipitation (Co-IP) was performed to evaluate the interaction between SIRT5 and CS. Western blotting was used to measure the succinylation levels of CS. In addition, Mito-Tracker Red CMXRos staining, reactive oxygen species (ROS) measurement, ATP level assay, EdU cell proliferation assay, colony formation assay, TUNEL staining, and flow cytometry were used to investigate the effects of CS succinylation and desuccinylation on mitochondrial function and cell proliferation in hepatocellular carcinoma cells. A total of 358 differentially modified proteins were identified in human liver cancer tissues. These differentially modified proteins were primarily enriched in the mitochondria, and CS exhibited high levels of succinylation in HCC tissues. In mouse liver cancer tissues, SIRT5 expression was reduced while CS expression was increased. Furthermore, SIRT5 was found to interact with CS, mediating the de-succinylation of CS at the lysine 375 site. Additionally, succinylation at the K375 site of CS was shown to enhance mitochondrial activity and ATP content in HepG2 cells, while reducing intracellular ROS levels and promoting cell proliferation. In contrast, de-succinylation of CS at the K375 site significantly impaired mitochondrial function and ATP levels, increased ROS levels, and induced apoptosis in HepG2 cells. Succinylation of CS is crucial for maintaining mitochondrial function and promoting cell proliferation in liver cancer cells. Targeting SIRT5-mediated de-succinylation of CS may represent a promising therapeutic strategy for the treatment of hepatocellular carcinoma.

The size of nanomaterials influences physicochemical parameters, and variations in the size of nanomaterials can have a significant effect on their biological activities in cells. Due to the potential applicability of nanoparticles (NPs), the current work was designed to carry out a size-dependent study of gold nanoparticles (GNPs) in different dimensions, synthesized via a colloidal solution process. Three dissimilar-sized GNPs, GNPs-1 (10–15 nm), GNPs-2 (20–30 nm), and GNPs-3 (45 nm), were prepared and characterized via transmission electron microscopy (TEM), high-resolution TEM (HR-TEM), hydrodynamic size, zeta potential, and UV-visible spectroscopy, and applied against liver cancer (HepG2) cells. Various concentrations of GNPs (1, 2, 5, 10, 50, and 100 µg/mL) were applied against the HepG2 cancer cells to assess the percentage of cell viability via MTT and NRU assays; reactive oxygen species (ROS) generation was also used. ROS generation was increased by 194%, 164%, and 153% for GNPs-1, GNPs-2, and GNPs-3, respectively, in the HepG2 cells. The quantitative polymerase chain reaction (qPCR) data for the HepG2 cells showed up-regulation in gene expression of apoptotic genes (Bax, p53, and caspase-3) when exposed to the different-sized GNPs, and defined their respective roles. Based on the results, it was concluded that GNPs of different sizes have the potential to induce cancer cell death.

Circulating levels of soluble lectin-like oxidized low-density lipoprotein receptor-1 (sLOX-1) play an important role in the development and progression of atherosclerosis. We hypothesized that the inflammatory marker C-reactive protein (CRP) might stimulate sLOX-1 release by activating tumor necrosis factor-α converting enzyme (TACE). Macrophages differentiated from THP-1 cells were stimulated with TNF-α and further treated with CRP in the absence or presence of specific inhibitors or small interfering RNA (siRNA). Our results showed that CRP increased sLOX-1 release from activated macrophages in a dose-dependent manner and that these effects were regulated by Fc γ receptor II (FcγRII)-mediated p47(phox) phosphorylation, reactive oxygen species (ROS) production, and TACE activation. CRP also enhanced sLOX-1 release from macrophages derived from peripheral blood mononuclear cells (PBMC) of patients with acute coronary syndrome (ACS). Pretreatment with antibody against FcγRII or with CD32 siRNA, p47(phox) siRNA, apocynin, N-acetylcysteine, tumor necrosis factor-α protease inhibitor 1 (TAPI-1) or TACE siRNA attenuated sLOX-1 release induced by CRP. CRP also elevated serum sLOX-1 levels in a rabbit model of atherosclerosis. Thus, CRP might stimulate sLOX-1 release, and the underlying mechanisms possibly involved FcγRII-mediated p47(phox) phosphorylation, ROS production, and TACE activation.

The calcium-ROS-pH triangle and mitochondrial permeability transition: challenges to mimic cardiac ischemia-reperfusion

Vascular permeability is a complex process involving the coordinated regulation of multiple signaling pathways in the endothelial cell. It has long been documented that vascular endothelial growth factor (VEGF) greatly enhances microvascular permeability; however, the molecular mechanisms controlling VEGF-induced permeability remain unknown. Treatment of microvascular endothelial cells with VEGF led to an increase in reactive oxygen species (ROS) production. ROS are required for VEGF-induced permeability as treatment with the free radical scavenger, N-acetylcysteine, inhibited this effect. Additionally, treatment with VEGF caused ROS-dependent tyrosine phosphorylation of both vascular-endothelial (VE)-cadherin and beta-catenin. Rac1 was required for the VEGF-induced increase in permeability and adherens junction protein phosphorylation. Knockdown of Rac1 inhibited VEGF-induced ROS production consistent with Rac lying upstream of ROS in this pathway. Collectively, these data suggest that VEGF leads to a Rac-mediated generation of ROS, which, in turn, elevates the tyrosine phosphorylation of VE-cadherin and beta-catenin, ultimately regulating adherens junction integrity.

Cancer is a cataclysmic disease that affects not only the target organ, but also the whole body. Metal-based nanoparticles (NPs) have recently emerged as a better option for the treatment of this deadly disease. Accordingly, the present work describes a means to control the growth of cancer cells by using colloidal silver nanoparticles (AgNPs) processed via homemade solutions and the characterization of these materials. The AgNPs may become an instantaneous solution for the treatment of these deadly diseases and to minimize or remove these problems. The AgNPs exhibit excellent control of the growth rate of human liver (HepG2) and breast (MCF-7) cancer cells, even at a very low concentrations. The cytotoxic effects of AgNPs on HepG2 and MCF-7 cancer cells were dose dependent (2–200 μg/mL), as evaluated using MTT and NRU assays. The production of reactive oxygen species (ROS) was increased by 136% and 142% in HepG2 and MCF-7 cells treated with AgNPs, respectively. The quantitative polymerase chain reaction (qPCR) data for both cell types (HepG2 and MCF-7) after exposure to AgNPs showed up- and downregulation of the expression of apoptotic (p53, Bax, caspase-3) and anti-apoptotic (BCl2) genes; moreover, their roles were described. This work shows that NPs were successfully prepared and controlled the growth of both types of cancer cells.

The aim of the present study was to investigate the biological role of glypican 3 (GPC3), and to identify its mechanism and clinical significance in the carcinogenesis of liver cancer. A total of 114 patients with liver cancer were involved. Their clinical data, hematoxylin and eosin-stained and Antigen Ki-67 protein (Ki-67) and GPC3 immunohistochemically-stained liver cancer tissue sections were analyzed to evaluate the correlation between the liver cancer proliferation, differentiation and GPC3 expression. Fluorescence microscopy, western blotting, MTT and reverse transcription quantitative polymerase chain reaction (RT-qPCR) assays were performed in HepG2 and HLE cell lines to investigate the potential mechanisms of action. Among the 114 patients with liver cancer enrolled in the present study, 12 exhibited well-differentiated liver cancer, of which 6 (50%) were positive for GPC3. A total of 30 cases exhibited poorly differentiated liver cancer; 26 (87%) of these expressed GPC3 and 11 cases (37%) demonstrated strong positive expression levels. The other 72 liver cancer cases were moderately differentiated; 75% (54/72) of these expressed GPC3 and 12.5% (9/72) exhibited strong positive expression levels. There was a significant association between the levels of GPC3 expression and liver cancer differentiation (χ2=16.306, P=0.008). Ki-67 staining as the criteria of the liver cancer cell proliferation index also indicated a cross correlation between liver cancer proliferation and GPC3 levels. Among the 39 liver cancer samples with a cell proliferation index <5%, only 2.6% (1/39) exhibited strong positive GPC3 staining, but of the 16 cases with a high cell proliferation index >50%, 6 exhibited strong GPC3 staining (37.5%). The difference of cell proliferation indexes between cancer cells were well, moderate and poorly differentiated, and was markedly significant (χ2=26.334, P=0.002), and suggested that liver cancer cell proliferation was positively correlated with GPC3 expression (r=0.316, P=0.001). Consistently, in vitro analysis indicated that GPC3 promoted HepG2 and HLE cell growth, which was more apparent in HepG2 cells. The RT-qPCR results indicated that GPC3 promoted proliferation through the Hedgehog (Hh) pathway in HepG2 cells, but not in HLE cells. In the present study, it was demonstrated that patients with liver cancer with higher GPC3 levels exhibited poorer differentiation and higher proliferation levels. In vitro GPC3 may promote liver cancer cell lines proliferation through the Hh pathway.