C.J. Chen

The circadian clock gene Bmal1 acts as a potential anti-oncogene in pancreatic cancer by activating the p53 tumor suppressor pathway.

Disruption of the circadian clock has been shown to be associated with tumor development. This study aimed to investigate the role of the core circadian gene Bmal1 in pancreatic cancer (PC). We first found that the levels of Bmal1 were downregulated in PC samples and were closely correlated with the clinicopathological features of patients. To dissect the underlying mechanism, we performed a RNA-seq assay followed by systematic gene function and pathway enrichment analyses. We detected an anti-apoptotic and pro-proliferative transcriptome profile after Bmal1 knockdown in PC cells. Further in vitro and in vivo studies confirmed that Bmal1 overexpression significantly inhibited cell proliferation and invasion and induced G2/M cell cycle arrest, whereas Bmal1 knockdown promoted PC growth, as demonstrated in Bmal1-manipulated AsPC-1 and BxPC-3 cell lines. Our mechanistic studies indicated that Bmal1 could directly bind to the p53 gene promoter and thereby transcriptionally activate the downstream tumor suppressor pathway in a p53-dependent manner. In sum, our findings suggest that Bmal1 acts as an anti-oncogene in PC and represents a potential biomarker for its diagnosis.

Transcriptional repression of SOCS3 mediated by IL-6/STAT3 signaling via DNMT1 promotes pancreatic cancer growth and metastasis

BackgroundPrevious studies have investigated the sustained aberrantly activated Interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) signaling pathway is crucial for pancreatic cancer growth and metastasis. Suppressor of cytokine signaling 3 (SOCS3), as a key negative feedback regulator of this signaling pathway, is usually down-regulated in various cancers. In the present study, we aim at exploring the biological function and the underlying molecular regulation mechanisms of SOCS3 in pancreatic cancer.MethodsThe expression of SOCS3 and other genes in pancreatic cancer was examined by Quantitative real-time PCR, western blotting and immunohistochemical staining. The interaction between pSTAT3 and DNA Methyltransferase 1 (DNMT1) was investigated by co-immunoprecipitation assay. Luciferase reporter assay was used to investigate the transcriptional regulation of pSTAT3 and DNMT1 on the SOCS3 gene. The effects of SOCS3 on the biological behavior of pancreatic cancer cells were assessed both in vitro and vivo. Furthermore, we performed a comprehensive analysis of the expression of SOCS3 in a pancreatic cancer tissue microarray (TMA) and correlated our findings with pathological parameters and outcomes of the patients.ResultsWe showed that SOCS3 expression was decreased in phosphorylated STAT3 (pSTAT3)-positive tumors and was negatively correlated with pSTAT3 in pancreatic cancer cells. We also found that IL-6/STAT3 promoted SOCS3 promoter hypermethylation by increasing DNMT1 activity; silencing DNMT1 or 5-aza-2-deoxycytidine (5-AZA) treatment could reverse the down-regulation of SOCS3 mediated by IL-6. Using co-immunoprecipitation and luciferase reporter assays, we found that STAT3 recruited DNMT1 to the promoter region of SOCS3 and inhibited its transcriptional activity. Overexpression of SOCS3 significantly inhibited cell proliferation, which may be due to the increase in G1-S phase arrest; overexpression of SOCS3 also inhibited cell migration and invasion as well as tumorigenicity in nude mice. Pancreatic cancer tissue microarray analysis showed that high SOCS3 expression was a good prognostic factor and negatively correlated with tumor volume and metastasis.ConclusionWe demonstrated that activated IL-6/STAT3 signaling could induce SOCS3 methylation via DNMT1, which led to pancreatic cancer growth and metastasis. These data also provided a mechanistic link between sustained aberrantly activated IL-6/STAT3 signaling and SOCS3 down-regulation in pancreatic cancer. Thus, inhibitors of STAT3 or DNMT1 may become novel strategies for treating pancreatic cancer.

Imbalance of Wnt/Dkk negative feedback promotes persistent activation of pancreatic stellate cells in chronic pancreatitis.

The role of persistent activation of pancreatic stellate cells (PSCs) in the fibrosis associated with chronic pancreatitis (CP) is increasingly being recognized. Recent studies have shown that Wnt signaling is involved in the development of fibrosis in multiple organs, however, the role of specific Wnts in pancreatic fibrosis remains unknown. We investigated the role of Wnt signaling during PSC activation in CP and the effect of β-catenin inhibition and Dickkopf-related protein 1 (Dkk1) restoration on the phenotype of PSCs. CP was induced in mice by repetitive caerulein injection and mouse PSCs were isolated and activated in vitro. The expression of Wnts, β-catenin, secreted frizzled-related proteins (sFRPs) and Dkks was analyzed by quantitative RT-PCR and western blotting. The canonical Wnt signaling pathway was examined by immunofluorescence and western blot detection of nuclear β-catenin expression. The effect of recombinant mouse Dkk-1 (rmDkk-1) on cell proliferation and apoptosis was assessed by flow cytometry, immunofluorescence, immunocytochemistry and Cell Counting Kit-8 (CCK-8) analysis. The expression of β-catenin, collagen1α1, TGFβRII, PDGFRβ and α-SMA in PSCs treated with different concentrations of rmDkk-1 or siRNA against β-catenin was determined by quantitative RT-PCR and western blotting. Wnt2 was the only Wnt whose expression was significantly upregulated in response to PSC activation, and Wnt2 and β-catenin protein levels were significantly increased in the pancreas of CP mice, whereas Dkk-1 expression was evidently decreased. Nuclear β-catenin levels were markedly increased in activated PSCs, and rmDkk-1 suppressed the nuclear translocation of β-catenin and the proliferation and extracellular matrix production of PSCs through the downregulation of PDGFRβ and TGFβRII. Upregulation of Dkk-1 expression increased apoptosis in cultured PSCs. These results indicate that Wnt signaling may mediate the profibrotic effect of PSC activation, and Wnt2/Dkk-1 could be potential therapeutic targets for CP.

The inhibitory effects of xanthohumol, a prenylated chalcone derived from hops, on cell growth and tumorigenesis in human pancreatic cancer.

Pancreatic cancer (PC) is one of the most lethal human malignancies worldwide. Here, we demonstrated that xanthohumol (XN), the most abundant prenylated chalcone isolated from hops, inhibited the growth of cultured PC cells and their subcutaneous xenograft tumors. XN treatment was found to induce cell cycle arrest and apoptosis of PC cells (PANC-1, BxPC-3) by inhibiting phosphorylation of signal transducer and activator of transcription 3 (STAT3) and expression of its downstream targeted genes cyclinD1, survivin, and Bcl-xL at the messenger RNA level, which involved in regulation of apoptosis and the cell cycle. Overall, our results suggested that XN presents a promising candidate therapeutic agent against human PC and the STAT3 signaling pathway is its key molecular target.

Pancreatic acinar cells-derived cyclophilin A promotes pancreatic damage by activating NF-κB pathway in experimental pancreatitis

Inflammation triggered by necrotic acinar cells contributes to the pathophysiology of acute pancreatitis (AP), but its precise mechanism remains unclear. Recent studies have shown that Cyclophilin A (CypA) released from necrotic cells is involved in the pathogenesis of several inflammatory diseases. We therefore investigated the role of CypA in experimental AP induced by administration of sodium taurocholate (STC). CypA was markedly upregulated and widely expressed in disrupted acinar cells, infiltrated inflammatory cells, and tubular complexes. In vitro, it was released from damaged acinar cells by cholecystokinin (CCK) induction. rCypA (recombinant CypA) aggravated CCK-induced acinar cell necrosis, promoted nuclear factor (NF)-κB p65 activation, and increased cytokine production. In conclusion, CypA promotes pancreatic damage by upregulating expression of inflammatory cytokines of acinar cells via the NF-κB pathway.

Substance P-regulated leukotriene B4 production promotes acute pancreatitis-associated lung injury through neutrophil reverse migration

&NA; Leukotriene B4 (LTB4) is a potent chemoattractant and inflammatory mediator involved in multiple inflammatory diseases. Substance P (SP) has been reported to promote production of LTB4 in itch‐associated response in vivo and in some immune cells in vitro. Here, we investigated the role of LTB4 in acute pancreatitis (AP), AP‐associated acute lung injury (ALI) and the related mechanisms of LTB4 production in AP. In vivo, murine AP model was induced by caerulein and lipopolysaccharide or L‐arginine. The levels of LTB4 and its specific receptor BLT1 were markedly upregulated in both AP models. Blockade of BLT1 by LY293111 attenuated the severity of AP, decreased neutrophil reverse transendothelial cell migration (rTEM) into the circulation and alleviated the severity of ALI. In vitro, treatment of pancreatic acinar cells with SP increased LTB4 production. Furthermore, SP treatment increased phosphorylation of protein kinase C (PKC) &agr; and mitogen activated protein kinases (MAPKs), including extracellular signal‐regulated kinase (ERK), p‐38 MAPK and c‐Jun NH2‐terminal kinase (JNK). Finally, blockade of neurokinin‐1 receptor by CP96345 significantly attenuated the severity of AP and decreased the level of LTB4 when compared to AP group. In summary, these results show that SP regulates the production of LTB4 via PKC&agr;/MAPK pathway, which further promotes AP‐associated ALI through neutrophil rTEM. HighlightsLTB4 increases local inflammation of acute pancreatitis (AP).LTB4 promotes AP‐associated lung injury through neutrophil reverse migration.Substance P regulates LTB4 production in AP via PKC&agr;/MAPK pathway.

Identification of Sox6 as a regulator of pancreatic cancer development

Pancreatic cancer (PC) is an aggressive malignancy associated with a poor prognosis and low responsiveness to chemotherapy and radiotherapy. Most patients with PC have metastatic disease at diagnosis, which partly accounts for the high mortality from this disease. Here, we explored the role of the transcription factor sex‐determining region Y‐box (Sox) 6 in the invasiveness of PC cells. We showed that Sox6 is down‐regulated in patients with PC in association with metastatic disease. Sox6 overexpression suppressed PC cell proliferation and migration in vitro and tumour growth and liver metastasis in vivo. Sox6 inhibited epithelial‐mesenchymal transition (EMT), and Akt signalling. Sox6 was shown to interact with the promoter of Twist1, a helix–loop–helix transcription factor involved in the induction of EMT, and to modulate the expression of Twist1 by recruiting histone deacetylase 1 to the promoter of the Twist1 gene. Twist1 overexpression reversed the effect of Sox6 on inhibiting EMT, confirming that the effect of Sox6 on suppressing tumour invasiveness is mediated by the modulation of Twist1 expression. These results suggest a novel mechanism underlying the aggressive behaviour of PC cells and identify potential therapeutic targets for the treatment of PC.

Thalidomide inhibits proliferation and epithelial‑mesenchymal transition by modulating CD133 expression in pancreatic cancer cells

Pancreatic cancer is a solid malignancy with a high mortality rate, on account of the high incidence of metastasis at the time of detection. The aggressiveness of pancreatic cancer may be partly driven by cancer stem cells (CSCs), which are characterized by the ability to self-renew and recapitulate tumors in the ectopic setting. However, although a number of drugs targeting CSCs are currently under clinical investigation, few effective drugs have been developed. The present study demonstrated that thalidomide inhibited cell proliferation and metastasis in pancreatic cancer cell lines through the inhibition of epithelial mesenchymal transition. The effect of thalidomide was more pronounced in cluster of differentiation 133 (CD133)+ SW1990 cells than in Capan-2 cells, in which CD133 expression was almost undetectable. The results revealed that CD133 is likely to serve a role in the antitumor effect of thalidomide and indicated that thalidomide could be developed as a CSC-specific adjuvant chemotherapy in pancreatic cancer.

Additional file 1: Figure S1. of Transcriptional repression of SOCS3 mediated by IL-6/STAT3 signaling via DNMT1 promotes pancreatic cancer growth and metastasis

(A) Normalized band quantization data of pSTAT3 and SOCS3 in Fig. 1c by Image J between matched pancreatic cancer and pericancerous tissues were used to confirm our claim. (B) Normalized band quantization data of pSTAT3 and SOCS3 in Fig. 1d by Image J were evaluated