IGF-1R down regulates the sensitivity of hepatocellular carcinoma to sorafenib through the PI3K / akt and RAS / raf / ERK signaling pathways.

BACKGROUND: Insulin-like growth factor-1 receptor (IGF-1R) promotes cell proliferation and migration and inhibitsapoptosis, all of which can contribute to the development of cancers. METHOD: This study investigated the effect and mechanism of IGF-1R in mediating the desensitization of hepatocellular carcinoma (HCC) to sorafenib. RESULTS: IGF-1R, highly expressed in the HCC cell lines SK-Hep1 and HepG2, promotes cell proliferation, migration, and anti-apoptosis through PI3K / Akt and RAS / Raf / ERK signaling pathways, resulting in HCC resistance to sorafenib. Knockdown of IGF-1R by RNA interference decreased proliferation and cell migration and upregulation of sorafenib-induced apoptosis of HCC cells. In vivo studies demonstrated that IGF-1R knockdown inhibited the growth of SK-Hep1 xenografts. CONCLUSION: These data are evidence that IGF-1R participates in regulating the survival and cell growth of HCC through the PI3K / Akt and RAS / Raf / ERK signaling pathways. Intervention in the expression of IGF-1R may increase the inhibitory effect of sorafenib on HCC.

IL-6 upregulates the expression of IL-6R through the JAK2/STAT3 signalling pathway to promote progression of hepatocellular carcinoma.

The progression of hepatocellular carcinoma (HCC) involves multifactor, multistep interactions. High expression of interleukin-6 receptor (IL-6R) plays an important role in the occurrence and development of tumours, but the regulatory mechanism of IL-6R expression and its function in HCC have not been fully defined. Western blot was used to evaluate the phosphorylation of key kinases in the JAK2/STAT3 pathway and the protein expression levels of related proliferation molecules, migration molecules and apoptotic molecules. The antiapoptosis, migration and proliferation of cells of each group were analysed with JC-1 to judge the cell apoptosis rate, the EdU method to determine the proliferation vitality of the cells, clone formation experiments and Transwell experiments. High expression of IL-6R in cell lines, lower protein levels of the apoptotic molecules c-Caspase7 and c-Caspase3 and higher protein levels of the proliferative molecules p-P70S6K and migration molecules MMP9 and MMP2 were consistent with stronger antiapoptosis, proliferation and migration. Interestingly, IL-6 upregulated the expression of IL-6R by activating the JAK2/STAT3 signalling pathway. Also, the expression of IL-6R protein was downregulated after lentivirus knockdown of STAT3. In nude mice bearing subcutaneous tumours, upregulation of IL-6R expression after activation of the JAK2/STAT3 signalling pathway by IL-6 significantly increased tumour growth. Moreover, the expression of IL-6R protein was downregulated, and the terminal tumour volume was significantly downregulated in the lentiviral STAT3 knockdown group. IL-6 regulated the transcription of IL-6R through the activation of the JAK2/STAT3 signalling pathway.

Coal dust nanoparticles induced pulmonary fibrosis by promoting inflammation and epithelial-mesenchymal transition via the NF-κB/NLRP3 pathway driven by IGF1/ROS-mediated AKT/GSK3ß signals.

Pneumoconiosis is the most common and serious disease among coal miners. In earlier work on this subject, we documented that coal dust (CD) nanoparticles (CD-NPs) induced pulmonary fibrosis (PF) more profoundly than did CD micron particles (CD-MPs), but the mechanism has not been thoroughly studied. Based on the GEO database, jveen, STRING, and Cytoscape tools were used to screen hub genes regulating PF. Particle size distribution of CD were analyzed with Malvern nanoparticle size potentiometer. Combining 8 computational methods, we found that IGF1, POSTN, MMP7, ASPN, and CXCL14 may act as hub genes regulating PF. Based on the high score of IGF1 and its important regulatory role in various tissue fibrosis, we selected it as the target gene in this study. Activation of the IGF1/IGF1R axis promoted CD-NPs-induced PF, and inhibition of the axis activation had the opposite effect in vitro and in vivo. Furthermore, activation of the IGF1/IGF1R axis induced generation of reactive oxygen species (ROS) to promote epithelial-mesenchymal transition (EMT) in alveolar epithelial cells (AECs) to accelerate PF. High-throughput gene sequencing based on lung tissue suggested that cytokine-cytokine receptor interaction and the NF-kB signaling pathway play a key role in PF. Also, ROS induced inflammation and EMT by the activation of the NF-kB/NLRP3 axis to accelerate PF. ROS can induce the activation of AKT/GSK3ß signaling, and inhibition of it can inhibit ROS-induced inflammation and EMT by the NF-kB/NLRP3 axis, thereby inhibiting PF. CD-NPs induced PF by promoting inflammation and EMT via the NF-κB/NLRP3 pathway driven by IGF1/ROS-mediated AKT/GSK3ß signals. This study provides a valuable experimental basis for the prevention and treatment of coal workers' pneumoconiosis. Illustration of the overall research idea of this study: IGF1 stimulates coal dust nanoparticles induced pulmonary fibrosis by promoting inflammation and EMT via the NF-κB/NLRP3 pathway driven by ROS-mediated AKT/GSK3ß signals.

Differences in the characteristics and pulmonary toxicity of nano- and micron-sized respirable coal dust.

BACKGROUND: The characteristics of coal dust (CD) particles affect the inhalation of CD, which causes coal worker's pneumoconiosis (CWP). CD nanoparticles (CD-NPs, < 500 nm) and micron particles (CD-MPs, < 5 µm) are components of the respirable CD. However, the differences in physicochemical properties and pulmonary toxicity between CD-NPs and CD-MPs remain unclear. METHODS: CD was analyzed by scanning electron microscopy, Malvern nanoparticle size potentiometer, energy dispersive spectroscopy, infrared spectroscopy, and electron paramagnetic resonance spectroscopy. CCK-8 assay, ELISA, transmission electron microscope, JC-1 staining, reactive oxygen species activity probe, calcium ion fluorescent probe, AO/EB staining, flow cytometry, and western blot were used to determine the differences between CD-NPs and CD-MPs on acute pulmonary toxicity. CCK-8, scratch healing and Transwell assay, hematoxylin-eosin and Masson staining, immunohistochemistry, immunofluorescence, and western blot were applied to examine the effects of CD-NPs and CD-MPs on pneumoconiosis. RESULTS: Analysis of the size distribution of CD revealed that the samples had been size segregated. The carbon content of CD-NPs was greater than that of CD-MPs, and the oxygen, aluminum, and silicon contents were less. In in vitro experiments with A549 and BEAS-2B cells, CD-NPs, compared with CD-MPs, had more inflammatory vacuoles, release of pro-inflammatory cytokines (IL-6, IL-1ß, TNFα) and profibrotic cytokines (CXCL2, TGFß1), mitochondrial damage (reactive oxygen species and Ca2+ levels and decreased mitochondrial membrane potential), and cell death (apoptosis, pyroptosis, and necrosis). CD-NPs-induced fibrosis model cells had stronger proliferation, migration, and invasion than did CD-MPs. In in vivo experiments, lung coefficient, alveolar inflammation score, and lung tissue fibrosis score (mean: 1.1%, 1.33, 1.33) of CD-NPs were higher than those of CD-MPs (mean: 1.3%, 2.67, 2.67). CD-NPs accelerated the progression of pulmonary fibrosis by upregulating the expression of pro-fibrotic proteins and promoting epithelial-mesenchymal transition. The regulatory molecules involved were E-cadherin, N-cadherin, COL-1, COL-3, ZO-1, ZEB1, Slug, α-SMA, TGFß1, and Vimentin. CONCLUSIONS: Stimulation with CD-NPs resulted in more pronounced acute and chronic lung toxicity than did stimulation with CD-MPs. These effects included acute inflammatory response, mitochondrial damage, pyroptosis, and necrosis, and more pulmonary fibrosis induced by epithelial-mesenchymal transition.

ASIC1α up-regulates MMP-2/9 expression to enhance mobility and proliferation of liver cancer cells via the PI3K/AKT/mTOR pathway.

A major challenge in the treatment of liver cancer is that a large proportion of patients fail to achieve long-term disease control, with death from liver cancer cell migration and invasion. Acid-sensitive ion channel 1α (ASIC1α) is involved in the migration, invasion, and proliferation of liver cancer cells. Therefore, we explored the mechanism of ASIC1α-mediated liver cancer cell migration and invasion. We determined the levels of ASIC1α by western blotting and immunofluorescence in HepG2 and SK-Hep1 cells cultured in various acidic conditions. In addition, wound healing assay, transwell invasion assay, and MTT assay were conducted to assess the migration, invasion, and proliferation abilities of liver cancer cells. Western blotting was conducted to determine the levels of MMP2, MMP9, ASIC1α, p-PI3Kp85, t-PI3Kp85, p-AKT(Ser473), t-AKT, p-mTOR (Ser2448), t-mTOR. We first found that the levels of ASIC1α in the HepG2 and SK-Hep1 cells in acidic conditions (pH 6.5) were significantly increased. Inhibition and knockdown of ASIC1α down-regulated MMP-2/9 expression and inhibited the migration, invasion, and proliferation of HepG2 and SK-Hep1 cells; overexpression of ASIC1α had the opposite effect. We further demonstrated that ASIC1α up-regulates MMP-2/9 via activation of the PI3K/AKT/mTOR pathway, thereby promoting migration, invasion, and proliferation of liver cancer cells. Overexpression of MMP-2/9 and activation of AKT reversed these effects on liver cancer cells caused by inhibition of ASIC1α. We conclude that ASIC1α can regulate migration, invasion, and proliferation of liver cancer cells through the MMP-2/9/PI3K/AKT/mTOR pathway. These observations may provide a new reference for liver cancer chemotherapy.

ASIC1a stimulates the resistance of human hepatocellular carcinoma by promoting EMT via the AKT/GSK3ß/Snail pathway driven by TGFß/Smad signals.

Multidrug resistance is the main obstacle to curing hepatocellular carcinoma (HCC). Acid-sensing ion channel 1a (ASIC1a) has critical roles in all stages of cancer progression, especially invasion and metastasis, and in resistance to therapy. Epithelial to mesenchymal transition (EMT) transforms epithelial cells into mesenchymal cells after being stimulated by extracellular factors and is closely related to tumour infiltration and resistance. We used Western blotting, immunofluorescence, qRT-PCR, immunohistochemical staining, MTT, colony formation and scratch healing assay to determine ASIC1a levels and its relationship to cell proliferation, migration and invasion. ASIC1a is overexpressed in HCC tissues, and the amount increased in resistant HCC cells. EMT occurred more frequently in drug-resistant cells than in parental cells. Inactivation of ASIC1a inhibited cell migration and invasion and increased the chemosensitivity of cells through EMT. Overexpression of ASIC1a upregulated EMT and increased the cells' proliferation, migration and invasion and induced drug resistance; knocking down ASIC1a with shRNA had the opposite effects. ASIC1a increased cell migration and invasion through EMT by regulating α and ß-catenin, vimentin and fibronectin expression via the AKT/GSK-3ß/Snail pathway driven by TGFß/Smad signals. ASIC1a mediates drug resistance of HCC through EMT via the AKT/GSK-3ß/Snail pathway.

Autophagy up-regulated by MEK/ERK promotes the repair of DNA damage caused by aflatoxin B1.

Aflatoxin B1 (AFB1), a kind of mycotoxin, exerts its cytotoxicity by increasing the oxidative damage of target organs, especially the liver. In vivo and in vitro experiments were carried out to elucidate the toxic mechanism of AFB1. The results of MTT, cloning-formation, flow cytometry, immunocytochemistry, Reverse transcription PCR (RT-PCR) and western blot showed that AFB1 activated NOX2 gp91 phox, inhibited proliferation and migration, and blocked cell cycle at G0/G1 period of HHL-5 cells. Autophagy promoted the repair of NOX2-dependent DNA damage. NOX2/gp91 phox mainly activates MEK/ERK pathway and then up-regulates autophagy. In vivo experiments have shown that AFB1 (0.75 mg/kg daily orally, 4 weeks) had no significant changes in the size and shape of the liver in mice. However, these treatments lead to structural abnormalities of hepatocytes and DNA damage. In summary, AFB1 caused intracellular oxidative stress and DNA damage, NOX2/gp91-phox activates the MEK/ERK pathway, and upregulated autophagy to promote the repair of DNA damage. We concluded that by increasing the level of autophagy, the ability of anti-AFB1 toxicity of liver can be increased.

Curcumin reverses hepatic epithelial mesenchymal transition induced by trichloroethylene by inhibiting IL-6R/STAT3.

OBJECTIVE: Epithelial mesenchymal transition (EMT) and inflammation have been identified as carcinogenic agents. This study aims to investigate whether inhibition of trichloroethylene (TCE) associated hepatocellular carcinoma (HCC) by curcumin is associated with inflammation and EMT. METHODS: In the current study, TCE sub-chronic cell model was induced in vitro, and the effects of TCE on cell proliferation, migration, invasion, and expression of functional proteins were verified by Western blot, MTT, clone formation, wound healing, Transwell. The detoxification effect of curcumin on TCE was explored by a mouse tumor-bearing experiment. RESULTS: TCE induces hepatocyte migration, colony formation, and EMT in vitro. In vivo studies have shown that curcumin significantly reduces the mortality of mice and control the occurrence and size of liver tumors by inhibiting the IL-6/STAT3 signaling pathway. In vitro, curcumin inhibits the proliferation of HepG2 cells as determined by MTT assay. In addition, curcumin significantly inhibited the protein expression of IL-6R, STAT3, snail, survivin, and cyclin D1 in THLE-2 and HepG2 cells induced by IL-6. CONCLUSION: Curcumin has anti-inflammatory and anti-proliferative effects, and inhibits the development of HCC induced by TCE by reversing IL-6/STAT3 mediated EMT.