Knockdown of OLR1 weakens glycolytic metabolism to repress colon cancer cell proliferation and chemoresistance by downregulating SULT2B1 via c-MYC

Chemoresistance is one of the major problems of colon cancer treatment. In tumors, glycolytic metabolism has been identified to promote cell proliferation and chemoresistance. However, the molecular mechanisms underlying glycolytic metabolism and chemoresistance in colon cancer remains enigmatic. Hence, this research was designed to explore the mechanism underlying the OLR1/c-MYC/SULT2B1 axis in the regulation of glycolytic metabolism, to affect colon cancer cell proliferation and chemoresistance. Colon cancer tissues and LoVo cells were attained, where OLR1, c-MYC, and SULT2B1 expression was detected by immunohistochemistry, RT-qPCR, and western blot analysis. Next, ectopic expression and knockdown assays were implemented in LoVo cells. Cell proliferation was detected by MTS assay and clone formation. Extracellular acidification, glucose uptake, lactate production, ATP/ADP ratio, and GLUT1 and LDHA expression were measured to evaluate glycolytic metabolism. Then, the transfected cells were treated with chemotherapeutic agents to assess drug resistance by MTS experiments and P-gp and SMAD4 expression by RT-qPCR. A nude mouse model of colon cancer transplantation was constructed for in vivo verification. The levels of OLR1, c-MYC, and SULT2B1 were upregulated in colon cancer tissues and cells. Mechanistically, OLR1 increased c-MYC expression to upregulate SULT2B1 in colon cancer cells. Moreover, knockdown of OLR1, c-MYC, or SULT2B1 weakened glycolytic metabolism, proliferation, and chemoresistance of colon cancer cells. In vivo experiments authenticated that OLR1 knockdown repressed the tumorigenesis and chemoresistance in nude mice by downregulating c-MYC and SULT2B1. Conclusively, knockdown of OLR1 might diminish SULT2B1 expression by downregulating c-MYC, thereby restraining glycolytic metabolism to inhibit colon cancer cell proliferation and chemoresistance.Subject terms: Cancer, Cancer therapy     

Cytosolic sulfotransferase 2B1b promotes hepatocyte proliferation gene expression in vivo and in vitro

Cytosolic sulfotransferase 2B1b (SULT2B1b) catalyzes the sulfation of 3β-hydroxysteroids and functions as a selective cholesterol and oxysterol sulfotransferase. Activation of liver X receptors (LXRs) by oxysterols has been known to be an antiproliferative factor. Overexpression of SULT2B1b impairs LXR's response to oxysterols, by which it regulates lipid metabolism. The aim of this study was to investigate in vivo and in vitro effects of SULT2B1b on liver proliferation and the underlying mechanisms. Primary rat hepatocytes and C57BL/6 mice were infected with adenovirus encoding SULT2B1b. Liver proliferation was determined by measuring the proliferating cell nuclear antigen (PCNA) immunostaining labeling index. The correlation between SULT2B1b and PCNA expression in mouse liver tissues was determined by double immunofluorescence. Gene expressions were evaluated by quantitative real-time PCR and Western blot analysis. SULT2B1b overexpression in mouse liver tissues increased PCNA-positive cells in a dose- and time-dependent manner. The increased expression of PCNA in mouse liver tissues was only observed in the SULT2B1b transgenic cells. Small interference RNA SULT2B1b significantly inhibited cell cycle regulatory gene expressions in primary rat hepatocytes. LXR activation by T0901317 effectively suppressed SULT2B1b-induced gene expression in vivo and in vitro. SULT2B1b may promote hepatocyte proliferation by inactivating oxysterol/LXR signaling.     

Notch1-mediated signaling regulates proliferation of porcine satellite cells (PSCs)

Notch signaling is an evolutionarily conserved cell–cell communication mechanism involved in the regulation of cell proliferation, differentiation and fate decisions of mammalian cells. In the present study, we investigated the possible requirement for Notch signaling in the proliferation and differentiation of porcine satellite cells. We show that Notch1, 2 and 3 are expressed in cultured porcine satellite cells. Knock-down of NOTCH1, but not NOTCH2 and NOTCH3, decreases the proliferation of porcine satellite cells. In contrast, enhancement of NOTCH1 expression via treatment of porcine satellite cells with recombinant NF-κB increases the proliferation of porcine satellite cells. The alteration of porcine satellite cell proliferation is associated with significant changes in the expression of cell cycle related genes (cyclin B1, D1, D2, E1 and p21), myogenic regulatory factors (MyoD and myogenin) and the Notch effector Hes5. In addition, alteration of Notch1 expression in porcine satellite cells causes changes in the expression of GSK3β-3. Taken together, these findings suggest that of the four notch-related genes, Notch1is likely to be required for regulating the proliferation and therefore the maintenance of porcine satellite cells in vivo, and do so through activation of the Notch effector gene Hes5.     

Downregulation of circDYNC1H1 exhibits inhibitor effect on cell proliferation and migration in hepatocellular carcinoma through miR-140-5p

Circular RNAs have been found to be aberrantly expressed in tumors and their significance in tumorigenesis has been focused on. The role of circDYNC1H1 in hepatocellular carcinoma (HCC) pathogenesis and its relationship with miR-140-5p were explored. The expression of circDYNC1H1, miR-140-5p, and SULT2B1 in HCC tissues and cells was measured, and Pearson's analysis was used to analyze their expression correlation. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and Transwell assays were performed to determine cell proliferation and migration. Binding between circDYNC1H1 and miR-140-5p was evaluated with RNA pull-down assay. A luciferase reporter assay was conducted to assess the interaction between circDYNC1H1 and miR-140-5p and between miR-140-5p and SULT2B1. circDYNC1H1 was highly expressed in HCC tissues (n = 20), and it was negatively associated with the expression of miR-140-5p but positively correlated with SULT2B1 messenger RNA expression. circDYNC1H1 was upregulated in cell lines of HCC. Interference of circDYNC1H1 suppressed cell proliferation and migration of HCC. circDYNC1H1 acted as a sponge of miR-140-5p. miR-140-5p controlled SULT2B1 expression by targeting its 3′-untranslated region. circDYNC1H1 enhanced SULT2B1 expression via sponging miR-140-5p. Downregulation of circDYNC1H1 disturbed cell proliferation and migration of HCC through miR-140-5p/SULT2B1 pathway. Silencing of circDYNC1H1 delayed tumor growth in HCC mouse model. Acting like a sponge of miR-140-5p, silenced circDYNC1H1 downregulated SULT2B1 to restrain HCC cell proliferation and migration, which is adverse to HCC growth and progression.     

Salinomycin Inhibits Proliferation and Induces Apoptosis of Human Hepatocellular Carcinoma Cells In Vitro and In Vivo

The anti-tumor antibiotic salinomycin (Sal) was recently identified as a selective inhibitor of breast cancer stem cells; however, the effect of Sal on hepatocellular carcinoma (HCC) is not clear. This study aimed to determine the anti-tumor efficacy and mechanism of Sal on HCC. HCC cell lines (HepG2, SMMC-7721, and BEL-7402) were treated with Sal. Cell doubling time was determinated by drawing growth curve, cell viability was evaluated using the Cell Counting Kit 8. The fraction of CD133⁺ cell subpopulations was assessed by flow cytometry. We found that Sal inhibits proliferation and decreases PCNA levels as well as the proportion of HCC CD133⁺cell subpopulations in HCC cells. Cell cycle was analyzed using flow cytometry and showed that Sal caused cell cycle arrest of the various HCC cell lines in different phases. Cell apoptosis was evaluated using flow cytometry and Hoechst 33342 staining. Sal induced apoptosis as characterized by an increase in the Bax/Bcl-2 ratio. Several signaling pathways were selected for further mechanistic analyses using real time-PCR and Western blot assays. Compared to control, β-catenin expression is significantly down-regulated upon Sal addition. The Ca²⁺ concentration in HCC cells was examined by flow cytometry and higher Ca²⁺ concentrations were observed in Sal treatment groups. The anti-tumor effect of Sal was further verified in vivo using the hepatoma orthotopic tumor model and the data obtained showed that the size of liver tumors in Sal-treated groups decreased compared to controls. Immunohistochemistry and TUNEL staining also demonstrated that Sal inhibits proliferation and induces apoptosis in vivo. Finally, the role of Sal on in vivo Wnt/β-catenin signaling was evaluated by Western blot and immunohistochemistry. This study demonstrates Sal inhibits proliferation and induces apoptosis of HCC cells in vitro and in vivo and one potential mechanism is inhibition of Wnt/β-catenin signaling via increased intracellular Ca²⁺ levels.     

Brivanib Attenuates Hepatic Fibrosis In Vivo and Stellate Cell Activation In Vitro by Inhibition of FGF,VEGF and PDGF Signaling

MethodsIn vivo, we induced liver fibrosis by bile duct ligation (BDL), chronic carbon tetrachloride (CCl₄), and chronic thioacetamide (TAA) administration. Liver fibrosis was examined by immunohistochemistry and Western immunoblotting. In vitro, we used LX-2 human hepatic stellate cells (HSCs) to assess the effect of brivanib on stellate cell proliferation and activation.ResultsAfter in vivo induction with BDL, CCl₄, and TAA, mice treated with brivanib showed reduced liver fibrosis and decreased expression of collagen Iα1 and α-smooth muscle actin in the liver. In vitro, brivanib decreased proliferation of HSCs induced by platelet-derived growth factor (PDGF), VEGF, and FGF. Brivanib also decreased stellate cell viability and inhibited PDGFBB-induced phosphorylation of its cognate receptor.ConclusionBrivanib reduces liver fibrosis in three different animal models and decreases human hepatic stellate cell activation. Brivanib may represent a novel therapeutic approach to treatment of liver fibrosis and prevention of liver cancer.     

GD3 Synthase Overexpression Sensitizes Hepatocarcinoma Cells to Hypoxia and Reduces Tumor Growth by Suppressing the cSrc/NF-κB Survival Pathway

Background

Hypoxia-mediated HIF-1α stabilization and NF-κB activation play a key role in carcinogenesis by fostering cancer cell survival, angiogenesis and tumor invasion. Gangliosides are integral components of biological membranes with an increasingly recognized role as signaling intermediates. In particular, ganglioside GD3 has been characterized as a proapoptotic lipid effector by promoting cell death signaling and suppression of survival pathways. Thus, our aim was to analyze the role of GD3 in hypoxia susceptibility of hepatocarcinoma cells and in vivo tumor growth.

Methodology/Principal Findings

We generated and characterized a human hepatocarcinoma cell line stably expressing GD3 synthase (Hep3B-GD3), which catalyzes the synthesis of GD3 from GM3. Despite increased GD3 levels (2–3 fold), no significant changes in cell morphology or growth were observed in Hep3B-GD3 cells compared to wild type Hep3B cells under normoxia. However, exposure of Hep3B-GD3 cells to hypoxia (2% O₂) enhanced reactive oxygen species (ROS) generation, resulting in decreased cell survival, with similar findings observed in Hep3B cells exposed to increasing doses of exogenous GD3. In addition, hypoxia-induced c-Src phosphorylation at tyrosine residues, NF-κB activation and subsequent expression of Mn-SOD were observed in Hep3B cells but not in Hep3B-GD3 cells. Moreover, MnTBAP, an antioxidant with predominant SOD mimetic activity, reduced ROS generation, protecting Hep3B-GD3 cells from hypoxia-induced death. Finally, lower tumor growth, higher cell death and reduced Mn-SOD expression were observed in Hep3B-GD3 compared to Hep3B tumor xenografts.

Conclusion

These findings underscore a role for GD3 in hypoxia susceptibility by disabling the c-Src/NF-κB survival pathway resulting in lower Mn-SOD expression, which may be of relevance in hepatocellular carcinoma therapy.     

Effect of bone marrow–derived mesenchymal stromal cells on hepatoma

Background aimsThe aim of the study was to evaluate the effect of mesenchymal stromal cells (MSCs) on tumor cell growth in vitro and in vivo and to elucidate the apoptotic and anti-proliferative mechanisms of MSCs on a hepatocellular carcinoma (HCC) murine model.MethodsThe growth-inhibitory effect of MSCs on the Hepa 1–6 cell line was tested by means of methyl thiazolyl diphenyl-tetrazolium assay. Eighty female mice were randomized into four groups: group 1 consisted of 20 mice that received MSCs only by intrahepatic injection; group 2 consisted of 20 HCC mice induced by inoculation of Hepa 1–6 cells into livers without MSC treatment; group 3 consisted of 20 mice that received MSCs after induction of liver cancer; group 4 consisted of 20 mice that received MSCs after induction of liver cancer on top of induced biliary cirrhosis.ResultsMSCs exhibited a growth-inhibitory effect on Hepa 1–6 murine cell line in vitro. Concerning in vivo study, decreases of serum alanine transaminase, aspartate transaminase and albumin levels after MSC transplantation in groups 2 and 3 were found. Gene expression of α-fetoprotein was significantly downregulated after MSC injection in the HCC groups. We found that gene expression of caspase 3, P21 and P53 was significantly upregulated, whereas gene expression of Bcl-2 and survivin was downregulated in the HCC groups after MSC injection. Liver specimens of the HCC groups confirmed the presence of dysplasia. The histopathological picture was improved after administration of MSCs to groups 2 and 3.ConclusionsMSCs upregulated genes that help apoptosis and downregulated genes that reduce apoptosis. Therefore, MSCs could inhibit cell division of HCC and potentiate their death.     

Inhibition of Cancer Cell Growth by Exposure to a Specific Time-Varying Electromagnetic Field Involves T-Type Calcium Channels

Electromagnetic field (EMF) exposures affect many biological systems. The reproducibility of these effects is related to the intensity, duration, frequency, and pattern of the EMF. We have shown that exposure to a specific time-varying EMF can inhibit the growth of malignant cells. Thomas-EMF is a low-intensity, frequency-modulated (25-6 Hz) EMF pattern. Daily, 1 h, exposures to Thomas-EMF inhibited the growth of malignant cell lines including B16-BL6, MDA-MB-231, MCF-7, and HeLa cells but did not affect the growth of non-malignant cells. Thomas-EMF also inhibited B16-BL6 cell proliferation in vivo. B16-BL6 cells implanted in syngeneic C57b mice and exposed daily to Thomas-EMF produced smaller tumours than in sham-treated controls. In vitro studies showed that exposure of malignant cells to Thomas-EMF for > 15 min promoted Ca²⁺ influx which could be blocked by inhibitors of voltage-gated T-type Ca²⁺ channels. Blocking Ca²⁺ uptake also blocked Thomas-EMF-dependent inhibition of cell proliferation. Exposure to Thomas-EMF delayed cell cycle progression and altered cyclin expression consistent with the decrease in cell proliferation. Non-malignant cells did not show any EMF-dependent changes in Ca²⁺ influx or cell growth. These data confirm that exposure to a specific EMF pattern can affect cellular processes and that exposure to Thomas-EMF may provide a potential anti-cancer therapy.     

In Vivo Silencing of A20 via TLR9-Mediated Targeted SiRNA Delivery Potentiates Antitumor Immune Response

A20 is an ubiquitin-editing enzyme that ensures the transient nature of inflammatory signaling pathways induced by cytokines like TNF-α and IL-1 or pathogens via Toll-like receptor (TLR) pathways. It has been identified as a negative regulator of dendritic cell (DC) maturation and attenuator of their immunostimulatory properties. Ex vivo A20-depleted dendritic cells showed enhanced expression of pro-inflammatory cytokines and costimulatory molecules, which resulted in hyperactivation of tumor-infiltrating T lymphocytes and inhibition of regulatory T cells. In the present study, we demonstrate that a synthetic molecule consisting of a CpG oligonucleotide TLR9 agonist linked to A20-specific siRNAs silences its expression in TLR9⁺ mouse dendritic cells in vitro and in vivo. In the B16 mouse melanoma tumor model, silencing of A20 enhances the CpG-triggered induction of NFκB activity followed by elevated expression of IL-6, TNF-α and IL-12. This leads to potentiated antitumor immune responses manifested by increased numbers of tumor-specific cytotoxic T cells, high levels of tumor cell apoptosis and delayed tumor growth. Our findings confirm the central role of A20 in controlling the immunostimulatory potency of DCs and provide a strategy for simultaneous A20 silencing and TLR activation in vivo.     

Multiple Delivery of siRNA against Endoglin into Murine Mammary Adenocarcinoma Prevents Angiogenesis and Delays Tumor Growth

Endoglin is a transforming growth factor-β (TGF- β) co-receptor that participates in the activation of a signaling pathway that mediates endothelial cell proliferation and migration in angiogenic tumor vasculature. Therefore, silencing of endoglin expression is an attractive approach for antiangiogenic therapy of tumors. The aim of our study was to evaluate the therapeutic potential of small interfering RNA (siRNA) molecules against endoglin in vitro and in vivo. Therapeutic potential in vitro was assessed in human and murine endothelial cells (HMEC-1, 2H11) by determining endoglin expression level, cell proliferation and tube formation. In vivo, the therapeutic potential of siRNA molecules was evaluated in TS/A mammary adenocarcinoma growing in BALB/c mice. Results of our study showed that siRNA molecules against endoglin have a good antiangiogenic therapeutic potential in vitro, as expression of endoglin mRNA and protein levels in mouse and human microvascular endothelial cells after lipofection were efficiently reduced, which resulted in the inhibition of endothelial cell proliferation and tube formation. In vivo, silencing of endoglin with triple electrotransfer of siRNA molecules into TS/A mammary adenocarcinoma also significantly reduced the mRNA levels, number of tumor blood vessels and the growth of tumors. The obtained results demonstrate that silencing of endoglin is a promising antiangiogenic therapy of tumors that could not be used as single treatment, but as an adjunct to the established cytotoxic treatment approaches.     

13-Methyltetradecanoic Acid Exhibits Anti-Tumor Activity on T-Cell Lymphomas In Vitro and In Vivo by Down-Regulating p-AKT and Activating Caspase-3

13-Methyltetradecanoic acid (13-MTD), a saturated branched-chain fatty acid purified from soy fermentation products, induces apoptosis in human cancer cells. We investigated the inhibitory effects and mechanism of action of 13-MTD on T-cell non-Hodgkin’s lymphoma (T-NHL) cell lines both in vitro and in vivo. Growth inhibition in response to 13-MTD was evaluated by the cell counting kit-8 (CCK-8) assay in three T-NHL cell lines (Jurkat, Hut78, EL4 cells). Flow cytometry analyses were used to monitor the cell cycle and apoptosis. Proteins involved in 13-MTD-induced apoptosis were examined in Jurkat cells by western blotting. We found that 13-MTD inhibited proliferation and induced the apoptosis of T-NHL cell lines. 13-MTD treatment also induced a concentration-dependent arrest of Jurkat cells in the G₁-phase. During 13-MTD-induced apoptosis in Jurkat cells, the cleavage of caspase-3 and poly ADP-ribose polymerase (PARP, a caspase enzymolysis product) were detected after incubation for 2 h, and increased after extending the incubation time. However, there was no change in the expression of Bcl-2 or c-myc proteins. The appearance of apoptotic Jurkat cells was accompanied by the inhibition of AKT and nuclear factor-kappa B (NF-κB) phosphorylation. In addition, 13-MTD could also effectively inhibit the growth of T-NHL tumors in vivo in a xenograft model. The tumor inhibition rate in the experimental group was 40%. These data indicate that 13-MTD inhibits proliferation and induces apoptosis through the down-regulation of AKT phosphorylation followed by caspase activation, which may provide a new approach for treating T-cell lymphomas.     

In Vitro and In Vivo Effect of 5-FC Combined Gene Therapy with TNF-α and CD Suicide Gene on Human Laryngeal Carcinoma Cell Line Hep-2

This study was aimed to investigate the effect of combined cancer gene therapy with exogenous tumor necrosis factor-alpha (TNF-α) and cytosine deaminase (CD) suicide gene on laryngeal carcinoma cell line Hep-2 in vitro and in vivo. Transfection of the recombinant eukaryotic vectors of pcDNA3.1 (+) containing TNF-α and/or CD into Hep-2 cells resulted in expression of TNF-α and/or CD gene in vitro. The significant increase in apoptotic Hep-2 cells and decrease of Hep-2 cell proliferation were observed using 5-FC treatment combined with TNF-a expression by CD/5-FC suicide system. Moreover, bystander effect was also observed in the TNF-α and CD gene co-expression group. Laryngeal squamous cell carcinoma (LSCC) mice model was established by using BALB/c mice which different transfected Hep-2 cells with pcDNA3.1 (+) containing TNF-α and/or CD were applied subcutaneously. So these mice are divided into four groups, namely, Hep-2/TIC group; Hep-2/CD group; Hep-2/TNF-α group; Hep-2/0 group. At day 29 after cell inoculation, volume of grafted tumor had significant difference between each two of them (P<0.05). These results showed that the products of combined CD and TNF-α genes inhibited the growth of transplanted LSCC in mice model. So by our observed parameters and many others results, we hypothesized that 5-FC combined gene therapy with TNF-αand CD suicide gene should be an effective treatment on Laryngeal carcinoma.     

The Intramembrane Proteases Signal Peptide Peptidase-Like 2a and 2b Have Distinct Functions In Vivo

We reported recently that the presenilin homologue signal peptide peptidase-like 2a (SPPL2a) is essential for B cell development by cleaving the N-terminal fragment (NTF) of the invariant chain (li, CD74). Based on this, we suggested that pharmacological modulation of SPPL2a may represent a novel approach to deplete B cells in autoimmune disorders. With regard to reported overlapping substrate spectra of SPPL2a and its close homologue, SPPL2b, we investigated the role of SPPL2b in CD74 NTF proteolysis and its impact on B and dendritic cell homeostasis. In heterologous expression experiments, SPPL2b was found to cleave CD74 NTF with an efficiency simliar to that of SPPL2a. For in vivo analysis, SPPL2b single-deficient and SPPL2a/SPPL2b double-deficient mice were generated and examined for CD74 NTF turnover/accumulation, B cell maturation and functionality, and dendritic cell homeostasis. We demonstrate that in vivo SPPL2b does not exhibit a physiologically relevant contribution to CD74 proteolysis in B and dendritic cells. Furthermore, we reveal that both proteases exhibit divergent subcellular localizations in B cells and different expression profiles in murine tissues. These findings suggest distinct functions of SPPL2a and SPPL2b and, based on a high abundance of SPPL2b in brain, a physiological role of this protease in the central nervous system.     

The RNA Binding Protein Igf2bp1 Is Required for Zebrafish RGC Axon Outgrowth In Vivo

Attractive growth cone turning requires Igf2bp1-dependent local translation of β-actin mRNA in response to external cues in vitro. While in vivo studies have shown that Igf2bp1 is required for cell migration and axon terminal branching, a requirement for Igf2bp1 function during axon outgrowth has not been demonstrated. Using a timelapse assay in the zebrafish retinotectal system, we demonstrate that the β-actin 3’UTR is sufficient to target local translation of the photoconvertible fluorescent protein Kaede in growth cones of pathfinding retinal ganglion cells (RGCs) in vivo. Igf2bp1 knockdown reduced RGC axonal outgrowth and tectal coverage and retinal cell survival. RGC-specific expression of a phosphomimetic Igf2bp1 reduced the density of axonal projections in the optic tract while sparing RGCs, demonstrating for the first time that Igf2bp1 is required during axon outgrowth in vivo. Therefore, regulation of local translation mediated by Igf2bp proteins may be required at all stages of axon development.     

Divergent expression and roles for caveolin-1 in mouse hepatocarcinoma cell lines with varying invasive ability

Caveolin-1 is the major component protein of caveolae and associated with a lot of cellular events such as endocytosis, cholesterol homeostasis, signal transduction, and tumorigenesis. The majority of results suggest that caveolin-1 might not only act as a tumor suppressor gene but also a promoting metastasis gene. In this study, the divergent expression and roles of caveolin-1 were investigated in mouse hepatocarcinoma cell lines Hca-F, Hca-P, and Hepa1-6, which have high, low, and no metastatic potential in the lymph nodes, as compared with normal mouse liver cell line IAR-20. The results showed that expression of caveolin-1 mRNA and protein along with the amount of caveolae number in Hca-F cells was higher than that in Hca-P cells, but was not detectable in Hepa1-6 cells. When caveolin-1 expression in Hca-F cells was down-regulated by RNAi approach, Hca-F cells proliferation rate in vitro declined and the expression of lymphangiogenic factor VEGFA in Hca-F decreased as well. Furthermore, in vivo implantation assay indicated that reduction of caveolin-1 expression in Hca-F prevented the lymphatic metastasis tumor burden of Hca-F cells in 615 mice. These results suggest that caveolin-1 facilities the lymphatic metastasis ability of mouse hepatocarcinoma cells via regulation tumor cell growth and VEGFA expression.     

Up-Regulation of RACK1 by TGF-β1 Promotes Hepatic Fibrosis in Mice

Liver fibrosis represents the consequences of a sustained wound healing response to chronic liver injury, and activation of quiescent hepatic stellate cells (HSCs) into a myofibroblast-like phenotype is considered as the central event of liver fibrosis. RACK1, the receptor for activated C-kinase 1, is a classical scaffold protein implicated in numerous signaling pathways and cellular processes; however, the role of RACK1 in liver fibrosis is little defined. Herein, we report that RACK1 is up-regulated in activated HSCs in transforming growth factor beta 1 (TGF-β1)-dependent manner both in vitro and in vivo, and TGF-β1 stimulates the expression of RACK1 through NF-κB signaling. Moreover, RACK1 promotes TGF-β1 and platelet-derived growth factor (PDGF)-mediated activation of pro-fibrogenic pathways as well as the differentiation, proliferation and migration of HSCs. Depletion of RACK1 suppresses the progression of TAA-induced liver fibrosis in vivo. In addition, the expression of RACK1 in fibrogenic cells also positively correlates well with the stage of liver fibrosis in clinical cases. Our results suggest RACK1 as a downstream target gene of TGF-β1 involved in the modulation of liver fibrosis progression in vitro and in vivo, and propose a strategy to target RACK1 for liver fibrosis treatment.