A comprehensive analysis of candidate genes and pathways in pancreatic cancer.

The study aimed to dissect the molecular mechanism of pancreatic cancer by a range of bioinformatics approaches. Three microarray datasets (GSE32676, GSE21654, and GSE14245) were downloaded from Gene Expression Omnibus database. Differentially expressed genes (DEGs) with logarithm of fold change (|logFC|) >0.585 and p value <0.05 were identified between pancreatic cancer samples and normal controls. Transcription factors (TFs) were selected from the DEGs based on TRASFAC database. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were performed for the DEGs using The Database for Annotation, Visualization and Integrated Discovery (p value <0.05), followed by construction of protein-protein interaction (PPI) network using Search Tool for the Retrieval of Interacting Genes software. Latent pathway identification analysis was applied to analyze the DEGs-related pathways crosstalk and the pathways with high weight value were included in the pathway crosstalk network using Cytoscape. Sixty-five DEGs were screened out. CCAAT/enhancer-binding protein delta (CEBPD), FBJ osteosarcoma oncogene B (FOSB), Stratifin (SFN), Krüppel-like factor 5 (KLF5), Pentraxin 3 (PTX3), and nuclear receptor subfamily 4, group A, member 3 (NR4A3) were important TFs. Interleukin-6 (IL-6) was the hub node of the PPI network. DEGs were significantly enriched in NOD-like receptor signaling pathway which was primarily interacted with inflammation and immune related pathways (cytosolic DNA-sensing, hematopoietic cell lineage, intestinal immune network for IgA production and chemokine pathways). The study suggested CEBPD, FOSB, SFN, KLF5, PTX3, NR4A3, IL-6, and NOD-like receptor pathways were involved in pancreatic cancer.

Matrine inhibits proliferation and induces apoptosis of human colon cancer LoVo cells by inactivating Akt pathway.

The present study has investigated the anti-tumor activity and the underlying mechanisms of matrine on human colon cancer LoVo cells. Matrine inhibited the proliferation of the cells in dose- and time-dependent manners. The concentration required for 50 % inhibition (IC50) was 1.15, 0.738, and 0.414 mg/ml, when cell were incubated with matrine for 24, 48, and 72 h, respectively. Matrine induced cell cycle arrest at G1 phase by downregulating cyclin D1 and upregulating p27 and p21. Matrine induced cell apoptosis by reducing the ratio of Bcl-2/Bax and increasing the activation of caspase-9 in a dose-dependent manner. Matrine displayed its anti-tumor activity by inactivating Akt, the upstream factor of the above proteins. Matrine significantly reduced the protein levels of pAkt, and increased the protein levels of other downstream factors, pBad and pGSK-3ß. Specific inhibition of pAkt induced cell apoptosis, and synergized with matrine to inhibit the proliferation of LoVo cells; whereas activation of Akt neutralized the inhibitory effect of matrine on cell proliferation. The present study has demonstrated that matrine inhibits proliferation and induces apoptosis of human colon cancer LoVo cells by inactivating Akt pathway, indicating matrine may be a potential anti-cancer agent for colon cancer.

The effects of artemisinin on the proliferation and apoptosis of vascular smooth muscle cells of rats.

OBJECTIVE: The aim of this study was to evaluate the effect of artemisinin on the proliferation and apoptosis of rat vascular smooth muscle cells (VSMCs). METHOD: Primary rat VSMCs were treated with various doses of artemisinin. Cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the messenger RNA and protein expressions of proliferating cell nuclear antigen were determined by reverse-transcription polymerase chain reaction and immunohistochemistry. Apoptosis was measured using annexin V and propidium iodide double staining evaluated by flow cytometry. Protein expression of Bax, Bcl2, and cyclin-dependent kinase 4 was determined by Western blot. RESULTS: After 72 h of treatment, artemisinin significantly inhibited VSMC proliferation in a dose-dependent manner. Treatment with 1 mM artemisinin for 72 h significantly reduced the expression of proliferating cell nuclear antigen messenger RNA. On the other hand, the same treatment increased the apoptosis of VSMCs, the activation of caspase-3, the Bax protein expression, and the Bax/Bcl2 ratio. CONCLUSION: The results suggest that artemisinin can effectively inhibit VSMC proliferation and induce VSMC apoptosis.

Identification of Mutation Landscape and Immune Cell Component for Liver Hepatocellular Carcinoma Highlights Potential Therapeutic Targets and Prognostic Markers.

Liver hepatocellular carcinoma (LIHC) is a primary malignancy, and there is a lack of effective treatment for advanced patients. Although numerous studies exist to reveal the carcinogenic mechanism of LIHC, few studies have integrated multi-omics data to systematically analyze pathogenesis and reveal potential therapeutic targets. Here, we integrated genomic variation data and RNA-seq profiles obtained by high-throughput sequencing to define high- and low-genomic instability samples. The mutational landscape was reported, and the advanced patients of LIHC were characterized by high-genomic instability. We found that the tumor microenvironment underwent metabolic reprograming driven by mutations accumulate to satisfy tumor proliferation and invasion. Further, the co-expression network identifies three mutant long non-coding RNAs as potential therapeutic targets, which can promote tumor progression by participating in specific carcinogenic mechanisms. Then, five potential prognostic markers (RP11-502I4.3, SPINK5, CHRM3, SLC5A12, and RP11-467L13.7) were identified by examining the association of genes and patient survival. By characterizing the immune landscape of LIHC, loss of immunogenicity was revealed as a key factor of immune checkpoint suppression. Macrophages were found to be significantly associated with patient risk scores, and high levels of macrophages accelerated patient mortality. In summary, the mutation-driven mechanism and immune landscape of LIHC revealed by this study will serve precision medicine.

Combining kallistatin gene therapy and meloxicam to treat hepatocellular carcinoma in mice.

Hepatocellular carcinoma (HCC) is one of the most common cancer-related causes of death, and conventional treatments offer unsatisfactory response. We have previously reported that kallistatin gene therapy suppressed the growth of HCC tumors by its anti-angiogenic activity, and meloxicam, a selective COX-2 inhibitor, inhibited proliferation and induced apoptosis of human HCC cells in vitro. The aim of this study was to determine whether combining kallistatin gene therapy and meloxicam could offer a better therapeutic effect to combat HCC in mice. A kallistatin expression plasmid was constructed and its expression was detected after intratumoral gene transfer. Both kallistatin gene therapy and meloxicam suppressed the growth of subcutaneous human HepG2 tumors established in BALB/c nude mice, and the combinational therapy showed a stronger effect in suppressing tumor growth, tumor angiogenesis and cell proliferation, and increasing cell apoptosis, than the respective monotherapies. Gene transfer of kallistatin inhibited tumor angiogenesis, and slightly inhibited cell proliferation and increased cell apoptosis in situ, but had no effect on expression of vascular endothelial growth factor, basic fibroblast growth factor, proliferating cell nuclear antigen, Bcl-2, Bax, or activation of caspase-3. Meloxicam therapy inhibited cell proliferation, induced cell apoptosis, reduced expression of proliferating cell nuclear antigen, increased activation of caspase-3, and upregulated Bax. Meloxicam also slightly inhibited tumor angiogenesis with no effect on the expression of vascular endothelial growth factor or basic fibroblast growth factor. Combining two novel anticancer agents, kallistatin targeting tumoral vascularization and meloxicam targeting cell proliferation and apoptosis, warrants investigation as a therapeutic strategy to combat HCC.

The IL-6/STAT3 pathway regulates adhesion molecules and cytoskeleton of endothelial cells in thromboangiitis obliterans.

Thromboangiitis obliterans (TAO) (also known as Buerger's disease) is an inflammatory vascular disease that predominantly affects small- and medium-sized blood vessels of extremities. Endothelial cells play critical roles in the initiation and progression of this disease, but the underlying mechanisms remain unclear. In the present study, we demonstrate that patients with TAO had significantly higher levels of interleukin-6 (IL-6), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1) in their plasmas, and the involved arterial tissues expressed higher levels of phosphorylated signal transducer and activator of transcription 3 (p-STAT3), ICAM-1 and VCAM-1. In exploring the molecular mechanisms with human aortic endothelial cells (HAECs), we found that recombinant IL-6 activated the STAT3 pathway, leading to the upregulation and overproduction of ICAM-1 and VCAM-1. RhoA (Ras homolog family member A), eNOS (endothelial nitric oxide synthase) and MMP-9 (matrix metalloproteinase-9) participated in this cellular signaling, and their interaction regulated the expression of ICAM-1 and VCAM-1. The activated STAT3 pathway by IL-6 also modulated the cytoskeleton of HAECs by regulating phosphorylation of focal adhesion kinase (FAK) and acetylation of α-tubulin through interplaying with RhoA. In summary, the present results indicate that activation of the IL-6/STAT3 pathway contributes to the pathogenesis of TAO by regulating cellular adhesion molecules and cytoskeleton of vascular endothelial cells, suggesting that targeting this pathway may provide a potential approach for the management of TAO.

Dual inhibition of Akt and c-Met as a second-line therapy following acquired resistance to sorafenib in hepatocellular carcinoma cells.

Sorafenib displays a limited efficacy for advanced hepatocellular carcinoma (HCC). Some patients with HCC initially respond to sorafenib, but eventually succumb to the disease, indicating that the acquired resistance to sorafenib reduces its beneficial effects. No alternative drugs are available after the failure of sorafenib therapy. Therefore, investigation of the mechanisms underlying the acquired resistance and development of second-line treatments for sorafenib-resistant HCC are urgently required. In this study, sorafenib-resistant HCC cells generated from sorafenib-sensitive human HCC cells were shown to overproduce hepatocyte growth factor (HGF) and overexpress c-Met kinase and its phosphorylated form, leading to the activation of Akt and ERK (extracellular signaling-regulated kinase) pathways. Use of specific c-Met inhibitors enhanced the effects of sorafenib by inhibiting the growth of sorafenib-resistant HCC cells. Akt inhibitors, a class of second-line therapeutic drugs under investigation for treating HCC in clinical trials, enhanced the effects of sorafenib, but also activated the c-Met pathway in sorafenib-resistant cells. Dual inhibition of Akt and c-Met by their respective inhibitors, MK2206 and capmatinib, additively or synergistically suppressed sorafenib-resistant HCC cells in vitro and sorafenib-resistant HCC xenografts in mice. The anticancer activities of MK2206 mainly rely on its ability to induce cell apoptosis and autophagic death, while capmatinib treatment leads to cell cycle arrest at phase G1. These results provide strong evidence for further investigation on the clinical utility of dual inhibition of Akt and c-Met, particularly MK2206 and capmatinib, as a second-line therapy for advanced HCC that has acquired resistance to sorafenib.

Pentoxifylline inhibits pulmonary inflammation induced by infrarenal aorticcross-clamping dependent of adenosine receptor A2A.

Infrarenal aortic cross-clamping (IAC) is commonly used during infrarenal vascular operations. Prolonged IAC causes ischemia-reperfusion injury to local tissues, resulting in the release of inflammatory cytokines and acute lung injury (ALI). Pentoxifylline (PTX) is a clinically used drug for chronic occlusive arterial diseases and exerts protective effects against ALI induced by various factors in experimental models. In this study, we evaluated the protective effects of PTX in a rat model of IAC. Wistar rats underwent IAC for 2 h, followed by 4 h reperfusion. PTX alone, or in combination with ZM-241385 (an adenosine receptor A2A antagonist) or CGS-21680 (an A2A agonist), was pre-administered to rats 1 h prior to IAC, and the severity of lung injury and inflammation were examined. Administration of PTX significantly attenuated ALI induced by IAC, evidenced by reduced histological scores and wet lung contents, improved blood gas parameters, decreased cell counts and protein amounts in bronchoalveolar lavage fluids, and inhibition of MPO activity and ICAM-1 expression in lung tissues, and lower plasma levels of TNF-α, IL-6, IL-1ß and soluble ICAM-1. ZM-241385 significantly abrogated, while CGS-21680 slightly enhanced, the effects of PTX in ameliorating ALI and inhibiting pulmonary inflammation. In exploration of the mechanisms, we found that PTX stimulated IL-10 production through the phosphorylation of STAT3, and A2A receptor participated in this regulation. The study indicates PTX plays a protective role in IAC-induced ALI in rats by inhibiting pulmonary inflammation through A2A signaling pathways.

Resveratrol attenuates inflammation in the rat heart subjected to ischemia-reperfusion: Role of the TLR4/NF-κB signaling pathway.

It has been previously reported that Toll­like receptor 4 (TLR4)/NF­κB signaling mediates early inflammation during myocardial ischemia and reperfusion. It has additionally been suggested that resveratrol produces cardioprotective and anti­inflammatory effects. The aim of the present study was to investigate whether resveratrol could modulate TLR4/NF­κB signaling, reduce neutrophil accumulation and TNF­α induction in an ischemia/reperfusion injured rat heart model. Rats were randomly exposed to a sham operation, myocardial ischemia and reperfusion (MI/R), MI/R + resveratrol or MI/R + resveratrol + L­NAME. The data showed that following MI/R, the expression of myocardial TLR4 and NF­κB increased significantly in the area of induced ischemia. As compared with MI/R, resveratrol significantly attenuated the expression of TLR4 and NF­κB and reduced the levels of myeloperoxidase, serum and myocardial TNF­α production, myocardial infarct size and myocardial apoptosis induced by MI/R. All the effects of resveratrol were abolished upon application of L­NAME, a nitric oxide (NO) synthase inhibitor. These data provide evidence that resveratrol inhibits TLR4/NF­κB signaling in the rat heart subjected to MI/R, and the anti­inflammatory effect of resveratrol is associated with NO production.

Association of polymorphisms of the receptor for advanced glycation end products gene and susceptibility to sporadic abdominal aortic aneurysm.

Accumulating evidence has suggested that receptor for advanced glycation end products (RAGE) is involved in the development and progression of human abdominal aortic aneurysms (AAAs). However, the association between RAGE gene polymorphisms and AAA has not yet been determined. The present study was aimed at analyzing the potential association between the RAGE gene polymorphisms and AAAs. A cohort of 381 patients and 436 age-matched healthy controls were genotyped to detect the three RAGE polymorphisms (-374 T/A, -429 T/C, and G82S) using SNaPshot. Our study demonstrated a significant difference in the genotype and allele frequencies of the RAGE G82S polymorphism between the AAA patients and the controls. Further stratification by gender and smoking status revealed that the presence of the RAGE 82S allele confers a higher risk for developing AAA in men and smokers. Moreover, AAA patients with the variant 82S allele of RAGE presented with reduced serum soluble RAGE (sRAGE) production, and this decrease was more significant in men and smokers with AAA. Our study provides preliminary evidence that the 82S allele of RAGE is a risk factor for AAA. This new piece of knowledge regarding RAGE may be clinically important for the prevention and therapy of AAAs.

Factors predisposing to severe acute pancreatitis: evaluation and prevention.

AIM: To analyze factors predisposing to the infections associated with severe acute pancreatitis (SAP) and to work out ways for its prevention. METHODS: Total 208 cases of SAP treated in this hospital from Jan. 1980 to Dec. 2001 were retrospectively analyzed. RESULTS: Statistical difference in the incidence of the aforementioned infections was found between the following pairs: between the groups of bloody or non-bloody ascites, paralytic ileus lasting shorter or longer than 5 days, Ranson scores lower or higher than 5, hematocrit lower or higher than 45 %, CT Balthazar scores lower or higher than 7 and between 1980.1-1992.6 or 1992.7-2001.12 admissions (chi(2)>3.84, P<0.05), while no statistical difference was established between the groups of biliogenic and non - biliogenic pancreatitis, serum amylase <200 U/L and > or =200 U/L, serum calcium <2 mmol /L and > or =2 mmol/L or groups of total parenteral nutrition shorter or longer than 7 days (chi(2)<3.84, P>0.05). CONCLUSION: Occurrence of infection in patients with SAP is closely related with bloody ascites, paralytic ileus > or =5 days, Ranson scores > or =5, hematocrit > or =45 % and CT Balthazar Scores > or =7, but not with pathogens, serum calcium and total parenteral nutrition (TPN). Comprehensive prevention of pancreatic infection and practice of individualized therapy contribute to reducing the incidence of infection.

Analysis and prevention of factors predisposing to infections associated with severe acute pancreatitis.

OBJECTIVE: To analyze factors predisposing to the infections associated with severe acute pancreatitis (SAP) and work out ways for their prevention. METHODS: 208 patients with SAP treated at our hospital from January 1986 to December 2001 were retrospectively analyzed. RESULTS: Statistical difference in the incidence of the infections was found between the following pairs: the groups of bloody or non-bloody ascites, paralytic ileus lasting shorter or longer than 5 days, Ranson's scores lower or higher than 5, hematocrit lower or higher than 45%, CT Balthazar scores lower or higher than 7, and between January 1986-June 1992 or July 1992-December 2001 admissions (chi2>7.58, P<0.05), while no statistical difference established between the groups of biliogenic and non-biliogenic pancreatitis, serum amylase <200 U/L and >/=200 U/L, serum calcium <2 mmol/L and >/=2 mmol/L or groups of total parenteral nutrition shorter or longer than 7 days (chi2<1.61, P>0.05). CONCLUSIONS: Occurrence of infection in patients with SAP is closely related with bloody ascites, paralytic ileus (>/=5 days), Ranson's scores (>/=5), hematocrit (>/=45%) and CT Balthazar scores (>/=7), but not with pathogenesis, serum calium or total parenteral nutrition. Comprehensive prevention of pancreatic infection and individualized therapy may reduce the incidence of infection.

The CLU gene rs11136000 variant is significantly associated with Alzheimer's disease in Caucasian and Asian populations.

Large-scale genomewide association studies have reported that the CLU rs11136000 polymorphism is significantly associated with Alzheimer's disease (AD) in people of Caucasian ancestry. Recently, this association was investigated in Asian populations (Chinese, Japanese, and Korean). However, these studies reported either a weak association or no association between the rs11136000 polymorphism and AD. We believe that this discrepancy may be caused by the relatively small sample size of the previous studies and the genetic heterogeneity of the rs11136000 polymorphism in AD among different populations. For this study, we searched the PubMed and AlzGene databases. We selected 18 independent studies (6 studies of Asian populations and 12 of populations of Caucasian ancestry) that evaluated the association between the rs11136000 polymorphism and AD using a case-control experimental design. We evaluated the genetic heterogeneity of the rs11136000 polymorphism in Caucasian and Asian populations. We then investigated the rs11136000 polymorphism by a meta-analysis in Asian populations using allele, dominant, and recessive models. We identified a significant association between rs11136000 and AD with the allele model (P = 2.00 × 10(-4)) and the dominant model (P = 5.00 × 10(-3)). Meanwhile, a similar genetic risk of the rs11136000 polymorphism in AD was observed in Asian and Caucasian populations. Further meta-analysis in pooled Asian and Caucasian populations indicated a more significant association with the allele (P = 8.30 × 10(-24)), dominant (P = 4.46 × 10(-17)), and recessive (P = 3.92 × 10(-12)) models. Collectively, our findings from this meta-analysis indicate that the effect of the CLU rs11136000 polymorphism on AD risk in Asian cohorts (Chinese, Japanese, and Korean) is consistent with the protective effect observed in Caucasian AD cohorts.

Serum bilirubin level is negatively correlated with disease progression of peripheral arterial disease: an observational cohort study.

We determined whether low bilirubin level is a risk factor for peripheral arterial disease (PAD). We recruited 318 patients with PAD and 100 healthy volunteers. Patients were divided into 4 groups by the Fontaine classification for PAD, namely, group 1 (grade 1, n = 4); group 2 (grade 2, n = 114), group 3 (grade 3, n = 164), and group 4 (grade 4, n = 36). Total bilirubin (T-BIL), direct bilirubin (D-BIL), and indirect bilirubin (I-BIL) levels were compared using stepwise multiple regressions adjusted for selected factors. After adjusting for gender, age, smoking, and diastolic blood pressure, serum levels of T-BIL, D-BIL, and I-BIL were significantly lower in the PAD group (P < .05). Patients with grade 4 PAD showed significantly (P < .05) lower levels of T-BIL when compared with grade 2 patients. We concluded that serum bilirubin levels are negatively correlated with the severity and progression of PAD.

Therapeutic effects of matrine on primary and metastatic breast cancer.

Matrine, one of the main components extracted from a traditional Chinese herb, Sophora flavescens Ait, has displayed anti-cancer activity in several types of cancer cells. This study aims to evaluate the therapeutic benefits of matrine on primary and metastatic breast cancer. Matrine inhibited the viability of and induced apoptosis in human MCF-7 and mouse 4T1 breast cancer cells in a dose-dependent manner in vitro as shown by MTT assay, flow cytometry and laser scanning confocal microscopy. Administration of matrine inhibited the growth of primary tumors and their metastases to lungs and livers, in a dose-dependent manner, in a highly metastatic model of 4T1 breast cancer established in syngeneic Balb/c mice. Tumors from matrine-treated mice had a smaller proliferation index, shown by immunostaining with an anti-Ki-67 antibody, a greater apoptosis index, shown by TUNEL-staining, and a less microvessel density, shown by immunostaining with an anti-CD31 A antibody, compared to the controls. Western blot analysis of tumoral homogenates indicated that matrine therapy reduced the ratio of Bcl-2/Bax, downregulated the expressions of VEGF and VEGFR-2, and increased the activation of caspase-3 and caspase-9. This study suggests matrine may be a potent agent, from a natural resource, for treating metastatic breast cancer because of its anti-apoptotic, anti-proliferative and anti-angiogenic activities.