Exogenous S1P via S1P receptor 2 induces CTGF expression through Src-RhoA-ROCK-YAP pathway in hepatic stellate cells.

BACKGROUND: Hepatic fibrosis, a prevalent chronic liver condition, involves excessive extracellular matrix production associated with aberrant wound healing. Hepatic stellate cells (HSCs) play a pivotal role in liver fibrosis, activated by inflammatory factors such as sphingosine 1-phosphate (S1P). Despite S1P's involvement in fibrosis, its specific role and downstream pathway in HSCs remain controversial. METHODS: In this study, we investigated the regulatory role of S1P/S1P receptor (S1PR) in Hippo-YAP activation in both LX-2 cell lines and primary HSCs. Real-time PCR, western blot, pharmacological inhibitors, siRNAs, and Rho activity assays were adopted to address the molecular mechanisms of S1P mediated YAP activation. RESULTS: Serum and exogenous S1P significantly increased the expression of YAP target genes in HSCs. Pharmacologic inhibitors and siRNA-mediated knockdowns of S1P receptors showed S1P receptor 2 (S1PR2) as the primary mediator for S1P-induced CTGF expression in HSCs. Results using siRNA-mediated knockdown, Verteporfin, and Phospho-Tag immunoblots showed that S1P-S1PR2 signaling effectively suppressed the Hippo kinases cascade, thereby activating YAP. Furthermore, S1P increased RhoA activities in cells and ROCK inhibitors effectively blocked CTGF induction. Cytoskeletal-perturbing reagents were shown to greatly modulate CTGF induction, suggesting the important role of actin cytoskeleton in S1P-induced YAP activation. Exogeneous S1P treatment was enough to increase the expression of COL1A1 and α-SMA, that were blocked by YAP specific inhibitor. CONCLUSIONS: Our data demonstrate that S1P/S1PR2-Src-RhoA-ROCK axis leads to Hippo-YAP activation, resulting in the up-regulation of CTGF, COL1A1 and α-SMA expression in HSCs. Therefore, S1PR2 may represent a potential therapeutic target for hepatic fibrosis.

Integrated 1D epitaxial mirror twin boundaries for ultrascaled 2D MoS2 field-effect transistors.

In atomically thin van der Waals materials, grain boundaries-the line defects between adjacent crystal grains with tilted in-plane rotations-are omnipresent. When the tilting angles are arbitrary, the grain boundaries form inhomogeneous sublattices, giving rise to local electronic states that are not controlled. Here we report on epitaxial realizations of deterministic MoS2 mirror twin boundaries (MTBs) at which two adjoining crystals are reflection mirroring by an exactly 60° rotation by position-controlled epitaxy. We showed that these epitaxial MTBs are one-dimensionally metallic to a circuit length scale. By utilizing the ultimate one-dimensional (1D) feature (width ~0.4 nm and length up to a few tens of micrometres), we incorporated the epitaxial MTBs as a 1D gate to build integrated two-dimensional field-effect transistors (FETs). The critical role of the 1D MTB gate was verified to scale the depletion channel length down to 3.9 nm, resulting in a substantially lowered channel off-current at lower gate voltages. With that, in both individual and array FETs, we demonstrated state-of-the-art performances for low-power logics. The 1D epitaxial MTB gates in this work suggest a novel synthetic pathway for the integration of two-dimensional FETs-that are immune to high gate capacitance-towards ultimate scaling.

Deep-ultraviolet electroluminescence and photocurrent generation in graphene/hBN/graphene heterostructures.

Hexagonal boron nitride (hBN) is a van der Waals semiconductor with a wide bandgap of ~ 5.96 eV. Despite the indirect bandgap characteristics of hBN, charge carriers excited by high energy electrons or photons efficiently emit luminescence at deep-ultraviolet (DUV) frequencies via strong electron-phonon interaction, suggesting potential DUV light emitting device applications. However, electroluminescence from hBN has not been demonstrated at DUV frequencies so far. In this study, we report DUV electroluminescence and photocurrent generation in graphene/hBN/graphene heterostructures at room temperature. Tunneling carrier injection from graphene electrodes into the band edges of hBN enables prominent electroluminescence at DUV frequencies. On the other hand, under DUV laser illumination and external bias voltage, graphene electrodes efficiently collect photo-excited carriers in hBN, which generates high photocurrent. Laser excitation micro-spectroscopy shows that the radiative recombination and photocarrier excitation processes in the heterostructures mainly originate from the pristine structure and the stacking faults in hBN. Our work provides a pathway toward efficient DUV light emitting and detection devices based on hBN.

Proline-rich tyrosine kinase 2 mediates transforming growth factor-beta-induced hepatic stellate cell activation and liver fibrosis.

Hepatic fibrogenesis is characterized by activation of hepatic stellate cells (HSCs) and accumulation of extracellular matrix (ECM). The impact of ECM on TGF-ß-mediated fibrogenic signaling pathway in HSCs has remained obscure. We studied the role of non-receptor tyrosine kinase focal adhesion kinase (FAK) family members in TGF-ß-signaling in HSCs. We used a CCl4-induced liver fibrosis mice model to evaluate the effect of FAK family kinase inhibitors on liver fibrosis. RT-PCR and Western blot were used to measure the expression of its target genes; α-SMA, collagen, Nox4, TGF-ß1, Smad7, and CTGF. Pharmacological inhibitors, siRNA-mediated knock-down, and plasmid-based overexpression were adopted to modulate the function and the expression level of proteins. Association of PYK2 activation with liver fibrosis was confirmed in liver samples from CCl4-treated mice and patients with significant fibrosis or cirrhosis. TGF-ß treatment up-regulated expression of α-SMA, type I collagen, NOX4, CTGF, TGF-ß1, and Smad7 in LX-2 cells. Inhibition of FAK family members suppressed TGF-ß-mediated fibrogenic signaling. SiRNA experiments demonstrated that TGF-ß1 and Smad7 were upregulated via Smad-dependent pathway through FAK activation. In addition, CTGF induction was Smad-independent and PYK2-dependent. Furthermore, RhoA activation was essential for TGF-ß-mediated CTGF induction, evidenced by using ROCK inhibitor and dominant negative RhoA expression. We identified that TGF-ß1-induced activation of PYK2-Src-RhoA triad leads to YAP/TAZ activation for CTGF induction in liver fibrosis. These findings provide new insights into the role of focal adhesion molecules in liver fibrogenesis, and targeting PYK2 may be an attractive target for developing novel therapeutic strategies for the treatment of liver fibrosis.

The therapeutic potential of induced hepatocyte-like cells generated by direct reprogramming on hepatic fibrosis.

BACKGROUND: Until now, there is no effective anti-fibrotic therapy available for liver cirrhosis. Stem cell therapies have been studied for the treatment of hepatic fibrosis. However, the use of embryonic stem cells or induced pluripotent stem cells (iPSC) has limitations such as ethical concern or malignancy potential. Induced hepatocyte-like cells (iHEPs) generated by direct reprogramming technology may overcome these limitations. METHODS: In this study, we generated iHEPs by direct reprogramming from mouse embryonic fibroblast (MEF) either using specific transcription factors such as c-Myc and Klf-4 (type A), or adding small molecules to HNF1α (type B). RESULTS: We investigated the effect of iHEPs on acute liver injury and chronic hepatic fibrosis animal models induced by CCl4 intra-peritoneal injection in BALB/C nude mice. In acute liver injury model, serum AST/ALT levels peaked at 24 h after CCl4 injection. Intra-splenic transplantation of iHEPs significantly attenuated CCl4-induced acute liver injury. GFP-labeled iHEPs (type A) migrated to the liver after intra-splenic transplantation that was confirmed by Western blotting and immunofluorescence staining. We found that GFP and albumin were co-localized in migrated iHEPs in the liver suggesting migrated iHEPs were functional. In chronic hepatic fibrosis mice experiment, transplantation of either type A or type B iHEPs significantly attenuated liver fibrosis induced by CCl4 injection for 10 weeks. CONCLUSIONS: Our study suggests that iHEPs may be used as a novel therapeutic strategy for the treatment of hepatic fibrosis.

KML001 enhances anticancer activity of gemcitabine against pancreatic cancer cells.

BACKGROUND/AIM: Gemcitabine is a drug commonly used to treat pancreatic cancer but chemoresistance to it is a common clinical issue. KML001 (sodium meta-arsenite) has demonstrated certain antitumor activity. The objective of the study was to evaluate the influence of KML001 on the anticancer activity of gemcitabine against pancreatic cancer cells. MATERIALS AND METHODS: Cell proliferation, migration, and invasion were assessed, as well as the expression of nuclear factor-kappa B (NF-κB) p65, epidermal growth factor receptor (EGFR), matrix metalloproteinase-2 (MMP2), and vascular endothelial growth factor-C (VEGFC) in pancreatic cancer cells. RESULTS: Treatment with a combination of KML001 and gemcitabine resulted in significant inhibition of cell proliferation, migration, and invasion, and significantly reduced EGFR and MMP2 expression compared to gemcitabine treatment-alone. CONCLUSION: Combination treatment of gemcitabine and KML001 could be an effective chemotherapeutic treatment for pancreatic cancer.

KML001 inhibits cell proliferation and invasion in pancreatic cancer cells through suppression of NF-κB and VEGF-C.

Pancreatic cancer is an aggressive malignancy with poor prognosis and the efficacy of chemotherapy is limited. KML001 (sodium meta-arsenite) has been demonstrated to have anticancer activity against some solid cancer cells. The aim of the present study was to determine the effect of KML001 on cell proliferation, migration, and invasion of pancreatic cancer cells. The Dojindo Cell Counting Kit-8 assay was used to determine the inhibition of pancreatic cancer cell proliferation by drugs. Cell migration and invasion were examined using 24-well inserts and Matrigel™-coated invasion chambers. The activity of nuclear factor-kappa B (NF-κB) p65, vascular endothelial growth factor-C (VEGF-C), and matrix metalloproteinase-9 (MMP-9) were measured by enzyme-linked immunosorbent assay (ELISA). KML001 inhibited the proliferation of pancreatic cancer cells in a dose- and time-dependent manner. KML001 also inhibited the migration and invasion of pancreatic cancer cells in a dose-dependent manner. KML001 significantly decreased NF-κB p65 and VEGF-C activities in the pancreatic cancer cells. KML001 inhibited cell proliferation, migration, and invasion in pancreatic cancer cells. Suppression of NF-κB and VEGF-C activation may partly be associated with the anticancer activity of KML001. These results suggest that KML001 could be a novel potential therapeutic agent for treatment of pancreatic cancer.

Inhibition of SCAMP1 suppresses cell migration and invasion in human pancreatic and gallbladder cancer cells.

Lymph node (LN) metastasis is one of the most important risk factors for the prognosis of pancreatic cancer. This study aimed to identify novel LN metastasis-associated markers and therapeutic targets for pancreatic and gallbladder cancers. DNA microarray analysis was carried out to identify genes differentially expressed between 17 pancreatic cancer tissues with LN metastasis and 17 pancreatic cancer tissues without LN metastasis. The expression of LZIC, FXR, SCAMP1, and SULT1E1 is significantly higher in pancreatic cancer tissues with LN metastasis than in pancreatic cancer tissues without LN metastasis. We recently reported that FXR plays an important role in LN metastasis of pancreatic cancer, and in this study, we selected the secretory carrier membrane protein 1 (SCAMP1) gene. To determine that function of the SCAMP1 gene, we examined the effects of SCAMP1 knockdown on pancreatic and gallbladder cancer proliferation, migration, and invasion using SCAMP1 small interfering RNA (siRNA) and the activity of vascular endothelial growth factor (VEGF) was measured by enzyme-linked immunosorbent assay. SCAMP1 overexpression is associated with LN metastasis in pancreatic cancer patients. The siRNA-mediated downregulation of SCAMP1 resulted in a marked reduction in cell migration and invasion, but not proliferation in MIA-PaCa2, PANC-1, TGBC-1, and TGBC-2 cells. In addition, downregulation of SCAMP1 inhibited VEGF levels of conditioned medium from SCAMP1 siRNA-transfected cells. These results suggest that downregulation of SCAMP1 could be a potential therapeutic target for patients with pancreatic and gallbladder cancer.

Guggulsterone enhances antitumor activity of gemcitabine in gallbladder cancer cells through suppression of NF-κB.

PURPOSE: Patients with gallbladder cancer usually have a poor prognosis, and effective standard chemotherapeutic regimens have not been established. The anticancer activities of guggulsterone have been demonstrated in various cancer cells. The aims of the study were to determine the effect of guggulsterone on gallbladder cancer cells and to investigate whether treatment with guggulsterone influences the antitumor activities of gemcitabine. METHODS: The Dojindo Cell Counting Kit-8 assay was used to determine the inhibition of proliferation by drugs in TGBC1 and TGBC2 cells. Cell migration and invasion were examined using 24-well inserts and Matrigel™-coated invasion chambers. The activities of NF-κB p65, VEGF-C, and MMP-2 were measured by ELISA. RESULTS: Guggulsterone inhibited the proliferation and suppressed migration and invasion of gallbladder cancer cells in a dose-dependent manner. Guggulsterone significantly decreased NF-κB p65, VEGF-C, and MMP-2 activities in the gallbladder cancer cells examined. Gallbladder cancer cells treated with a combination of guggulsterone and gemcitabine demonstrated significant inhibition of cell proliferation and invasion when compared to treatment with gemcitabine alone. In addition, NF-κB p65 activation decreased significantly in cells treated with a combination of guggulsterone and gemcitabine when compared to treatment with gemcitabine alone. CONCLUSIONS: Guggulsterone exhibits anticancer activities and enhances the antitumor activities of gemcitabine through the suppression of NF-κB activation in gallbladder cancer cells. These results suggest that guggulsterone could be a potential therapeutic option for patients with gallbladder cancer.

A critical time window for the survival of neural progenitor cells by HDAC inhibitors in the hippocampus.

Histone deacetylase inhibitors (HDACIs) that modulate gene expression by inhibiting HDAC enzymes may contribute to the survival of immature hippocampal neurons. However, it remains unknown how and when HDACIs regulate the survival of newly generated immature hippocampal neurons. In the present study, if the treatment of valproic acid (VPA) and sodium butyrate (SBt) in the specific time window during the development of newly generated n eurons r esulted in the i ncreased survival of bromodeoxyuridine (BrdU)(+) neurons in the dentate gyrus (DG) of hippocampus in mice was investigated. It was found that the number of BrdU(+) cells, the expressions of anti-apoptotic Bcl-2 family members and pCREB [D1] were increased by HDACIs when HDACIs were treated no later than 2-3 weeks after BrdU labeling. This suggests that epigenetic modification within a specific time window is critical for the survival of newborn hippocampal neurons by inhibiting the apoptotic pathway.