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Poor prognosis of digestive system cancers is mainly owing to lack of accurate and timely diagnosis. The exploration of novel tumor biomarkers from extracellular vesicle (EV) might be helpful to clinical diagnosis for digestive system cancers.Several public databases were first used for a preliminary screening of candidate genes. The RNA levels of these candidate genes were then detected in cancer cell lines and the patients serum-derived EVs by PCR Array or digital PCR, respectively.We found that 4 EV-RNAs, ANLN, ITGA6, KRT18, and MMP9, had a lower level in gastrointestinal cancer patients than in benign gastrointestinal diseases patients and healthy controls, while 3 EV-RNAs, ANLN, ITGA6, and KRT18, had a lower level in pancreatic cancer patients than in benign pancreatic diseases patients or healthy individuals. And EV-RNA of MMP9 had a relatively higher level in advanced pancreatic cancer patients than in early-stage patients. Moreover, ROC analysis demonstrated that the determination of the above EV-RNAs could increase the ability of traditional tumor biomarkers to distinguish benign and malignant diseases.The serum-derived EV-RNAs of ANLN, ITGA6, KRT18, and MMP9 could be served as novel, non-invasive biomarkers for digestive system cancers.

Introduction: Inflammation in early life is a risk factor for the development of neuropsychiatric diseases later in adolescence and adulthood, yet the underlying mechanism remains elusive. In the present study, we performed an integrated proteomic and phosphoproteomic analysis of the hippocampus to identify potential molecular mechanisms of early life inflammation-induced cognitive impairment. Methods: Both female and male mice received a single intraperitoneal injection of 100 μg/kg lipopolysaccharide (LPS) on postnatal day 10 (P10). Behavioral tests, including open field, elevated plus-maze, and Y-maze tests, were performed on P39, P40, and P41, respectively. After behavioral tests, male mice were sacrificed. The whole brain tissues and the hippocampi were harvested on P42 for proteomic, phosphoproteomic, Western blot, and Golgi staining. Results: Early life LPS exposure induced cognitive impairment in male mice but not in female mice, as assessed by the Y-maze test. Therefore, following biochemical tests were conducted on male mice. By proteomic analysis, 13 proteins in LPS group exhibited differential expression. Among these, 9 proteins were upregulated and 4 proteins were downregulated. For phosphoproteomic analysis, a total of 518 phosphopeptides were identified, of which 316 phosphopeptides were upregulated and 202 phosphopeptides were downregulated in the LPS group compared with the control group. Furthermore, KEGG analysis indicated that early life LPS exposure affected the glutamatergic synapse and neuroactive ligand-receptor interaction, which were associated with synaptic function and energy metabolism. Increased level of brain protein i3 (Bri3), decreased levels of PSD-95 and mGLUR5, and dendritic spine loss after early life LPS exposure further confirmed the findings of proteomic and phosphoproteomic analysis. Conclusions: Our findings demonstrated that neuroinflammation and impaired synapse may be involved in early life inflammation-induced cognitive impairment. Future studies are required to confirm our preliminary results.

Background Recently, some genomic mutations in exosomal DNA have been found to be related to disease progress and clinical outcomes of patients in several cancers. Unfortunately, the methods for exosome isolation and exosomal DNA analysis are still lack of relevant research to ensure their optimal performance and the comparability. Here we aim to establish a protocol for cancer-related mutation detection on exosomal DNA in clinical application. Methods Taking KRAS mutation in pancreatic cancer as an example, we tested whether the types of blood samples, the potential factors in the courses of exosome isolation and exosomal DNA preparation, as well as the detail in mutation detection by droplet digital PCR (ddPCR) could influence the exosomal DNA analysis. Results We found that the concentration of exosomal DNA from serum was higher than that from plasma, whereas the mutant allele fraction (MAF) of KRAS in serum-derived exosomal DNA was obviously lower. The membrane-based method for exosome isolation showed no evident difference in both exosomal DNA yield and KRAS MAF from the classical ultracentrifugation method. DNase I pretreatment on exosomes could remove the wild-type DNA outside of exosomes and increase the KRAS MAF. PBS might interfere with the effect of DNase I and should not be recommended as resuspension buffer for exosomes if the subsequent experiments would be done with exosomal DNA. Besides, the denaturation of exosomal DNA before droplet generation during ddPCR could effectively improve the total KRAS copy number and mutation-positive droplet number. Conclusion This study provides some methodological evidences for the selection of the optimal experimental conditions in exosomal DNA analysis. We also suggest a protocol for mutation detection on exosomal DNA, which might be suitable for the clinical testing and could be helpful to the comparison of results from different laboratories.

Background: Clinical management of metastatic gastric cancer (mGC) remains a major challenge due to a lack of specific biomarkers and effective therapeutic targets. Recently, accumulating evidence has suggested that exosomes play an essential role in cancer metastasis and can be an excellent reservoir of novel biomarkers and candidate therapeutic targets for cancer. Therefore, in this study, we aimed to reveal the proteomic profile of mGC-derived exosomes. Methods: Exosomes were isolated from pooled serum samples of 20 mGC patients and 40 healthy controls (HC) by ultracentrifugation. Next, quantitative proteomic analyses were applied to analyze the protein profiles of the exosomes, and bioinformatic analyses were conducted on the proteomic data. Finally, the expression of exosomal protein candidates was selectively validated in individual subjects by western blot analysis. Results: We isolated exosomes from serum samples. The size of the serum derived exosomes ranged from 30 to 150 nm in diameter. The exosomal markers CD9 and CD81 were observed in the serum exosomes. However, the exosomal negative marker calnexin, an endoplasmic reticulum protein, was not detected in exosomes. Overall, 443 exosomal proteins, including 110 differentially expressed proteins (DEPs) were identified by quantitative proteomics analyses. The bioinformatics analyses indicated that the upregulated proteins were enriched in the process of protein metabolic, whereas the downregulated proteins were largely involved in cell-cell adhesion organization. Surprisingly, 10 highly vital proteins (UBA52, PSMA1, PSMA5, PSMB6, PSMA7, PSMA4, PSMA3, PSMB1, PSMA6, and FGA) were filtered from DEPs, most of which are proteasome subunits. Moreover, the validation data confirmed that PSMA3 and PSMA6 were explicitly enriched in the serum derived exosomes from patients with mGC. Conclusion: The present study provided a comprehensive description of the serum exosome proteome of mGC patients, which could be an excellent resource for further studies of mGC.

Cytarabine (Ara-c) has been an important agent in acute myeloid leukemia (AML) treatment for more than 40 years. While, the mechanisms underlying low dose cytarabine (LD Ara-c) is poorly understood. In this study, we investigated the therapeutic effect of LD Ara-C in vitro. U937 and HEL cell lines were treated with increasing dose of Ara-C and showed growth inhibition rates in a time and dose-dependent manner. Treatment with LD Ara-C (50nM) induced a time-dependent increase in expression of microtubule-associated protein light chain 3 (LC3) and beclin1, but degradation of sequestosome1 (p62) in both U937 and HEL cells. Characteristic of autophagosomes appeared after 24h treatment. Meanwhile, deregulation of Akt-mTOR pathway was also detected. When cultured in presence of autophagy inhibitors, autophagy and differentiation was reversed, and cell growth inhibition was also attenuated. Similar phenomenon could also be seen when beclin1 expression was down-regulated. Taken together, we concluded that LD Ara-C can induce autophagy in AML cells and appeared to play an important role in differentiation and death. Down-regulation of Akt-mTOR pathway is involved in these processes. We suggest that cytarabine-induced autophagy is not a pro-survival mechanism, but accounts for its antineoplastic effects.

Interferon regulatory factor-1 (IRF-1) is a member of the interferon regulatory factor family. It acts as a transcriptional activator and plays a critical role in antiviral defense, immune response, cell growth regulation, apoptosis and cell differentiation. Deletions, mutations or aberrant splicing of IRF-1 would result in its functional inactivation, and closely related to the tumorigenesis. In this work, we identified an IRF-1 splicing transcript (IRF-1-s) in all-trans retinoic acid (ATRA)-treated acute promyelocytic leukemia (APL) cell line NB4 cells. It lost the exon 8 and 9 of the full length IRF-1, expressed in numerous cell types and could be induced to expression by ATRA in NB4 cells. It turned out similar biological activity as full length IRF-1 to enhance the transcription of interferon stimulated response element (ISRE)-containing target genes. Identification of IRF-1-s in NB4 cells would be benefit for our further exploring the signaling pathway of ATRA and interferons, as well as the mechanisms of differentiation of APL cells.

Background Whether muscle weakness is a cause, or result, of degenerative spondylolisthesis is not currently well understood. Little biomechanical evidence is available to offer an explanation for the mechanism behind exercise therapy. Therefore, the aim of this study is to investigate the effects of back muscle weakness on degenerative spondylolisthesis and to tease out the biomechanical mechanism of exercise therapy. Methods A nonlinear 3-D finite element model of L3–L5 was constructed. Forces representing global back muscles and global abdominal muscles, follower loads and an upper body weight were applied. The force of the global back muscles was reduced to 75%, 50% and 25% to simulate different degrees of back muscle weakness. An additional boundary condition which represented the loads from other muscles after exercise therapy was set up to keep the spine in a neutral standing position. Shear forces, intradiscal pressure, facet joint forces and von Mises equivalent stresses in the annuli were calculated. Findings The intervertebral rotations of L3–L4 and L4–L5 were within the range of in vitro experimental data. The calculated intradiscal pressure of L4–L5 for standing was 0.57 MPa, which is similar to previous in vivo data. With the back muscles were reduced to 75%, 50% and 25% force, the shear force moved increasingly in a ventral direction. Due to the additional stabilizing force and moment provided by boundary conditions, the shear force varied less than 15%. Interpretation Reducing the force of global back muscles might lead to, or aggravate, degenerative spondylolisthesis with forward slipping from biomechanical point of view. Exercise therapy may improve the spinal biomechanical environment. However, the intrinsic correlation between back muscle weakness and degenerative spondylolisthesis needs more clinical in vivo study and biomechanical analysis.

Background: The research on circulating tumor DNA (ctDNA) in pancreatic cancer (PC) has emerged recently. Although the detection rate of the KRAS mutation in ctDNA was relatively consistent with that in tumor tissue, whether the KRAS mutant allele fraction (MAF) differed was still not reported. So far, the clinical application of ctDNA detection in PC remains inconclusive. Methods: Plasma samples were collected from 110 PC and 52 pancreatic benign (PB) disease patients. The detection of KRAS mutation in ctDNA was performed using droplet digital PCR and compared with that in matched tumor tissue. We assessed the utility of KRAS MAFs in ctDNA and tissue for pancreatic malignancy assessment. Results: We found that KRAS MAF in ctDNA of PC patients was higher than that of PB patients, and was obviously associated with tumor staging and distant metastasis. However, KRAS MAF in ctDNA was significantly different from that in matched tissue. KRAS MAF in tumor tissue had no significant correlation with the clinical status. In addition, a ROC curve analysis revealed that mutant KRAS ctDNA combined with CA19-9 could increase the sensitivity rate of early-stage PC prediction, compared with CA19-9 test alone. Conclusion: The MAF of KRAS in ctDNA was related to the clinical stage of PC (p = 0.001). Mutant KRAS ctDNA could improve the sensitivity in early diagnosis of PC as a complement to CA19-9. Our study suggested that KRAS mutation in ctDNA could be a valuable circulating biomarker for the malignancy assessment in PC.

To establish a method for detecting the exosomal PML-RARA fusion gene expression by droplet digital PCR (ddPCR).By using Taqman probe-based ddPCR technique, the method that able to detect both long and short isoforms of PML-RARA fusion gene transcripts was established. RNA from PML-RARA negative cell line HL-60 as negative control was used to set the limit of blank (LOB), while the RNA from PML-RARA positive cell line NB4 and the recombinant plasmid pSG5-PML-RARA(S) were used to set the limit of detection (LOD) for long and short PML-RARA transcripts, respectively. Furtherly, the expression of exosomal PML-RARA fusion gene in NB4 cell culture supernatant and serum of patients with acute promyelocytic leukemia (APL) was analyzed by ddPCR technique.The LOB of ddPCR assay for long and short PML-RARA transcripts were 0.0725 and 0.083 copies per microliter of PCR reaction system, respectively, while the LOD of long and short PML-RARA transcripts were 0.19 and 0.21 copies per microliter of PCR reaction system, respectively. In addition, the expression of exosomal PML-RARA fusion gene derived from both NB4 cell culture supernatant and serum of APL patients was successfully detected.A ddPCR-based technique for detecting fusion gene transcripts has been established, which can be used to analyze absolute quantification in the minimal quantity of PML-RARA transcripts derived from exosomes. It suggests the possibility of this technique to non-invasively and dynamicly monitore the exosomal PML-RARA transcripts from APL patients' serum.微滴式数字PCR技术检测外泌体PML-RARA融合基因的表达.利用微滴式数字PCR技术检测外泌体中PML-RARA融合基因表达水平.使用基于Taqman探针法的ddPCR技术，建立可检测长型和短型PML-RARA融合基因转录本的方法。以PML-RARA阴性细胞株HL-60细胞的RNA逆转录合成的cDNA为阴性对照建立空白检测限（LOB）；以表达长型PML-RARA融合基因的NB4细胞RNA逆转录合成的cDNA和含短型PML-RARA cDNA的重组质粒为模板分别确定两者的最低检测限（LOD）；并检测NB4细胞培养上清来源的外泌体和急性早幼粒白血病患者血清来源的外泌体中PML-RARA融合基因的表达情况.采用ddPCR技术针对长型和短型PML-RARA转录本测得的LOB分别为每微升PCR反应体系中含0.0725拷贝和0.083拷贝；测得的LOD分别为每微升PCR反应体系中含0.19拷贝和0.21拷贝；使用该检测方法，在NB4细胞培养上清来源的外泌体和急性早幼粒细胞白血病患者血清来源的外泌体中均可检测到PML-RARA 融合基因的表达.建立了一种基于ddPCR技术检测融合基因转录本的方法，可对外泌体中的微量PML-RARA转录本进行绝对定量，为无创动态监测急性早幼粒细胞白血病患者血清外泌体中PML-RARA融合基因的表达水平提供了可能.

In civil engineering, imaging can be a useful non-destructive testing (NDT) tool for revealing defects in a concrete structure. To date, there is no effective method for imaging the defects inside in situ concrete structures. In this study, a new elastic wave-based NDT method has been developed and then applied to reveal the defects inside concrete specimens using an imaging method. This method integrates the point-source/point-receiver method with the synthetic aperture focusing technique (SAFT) to achieve a scanning effect when using a phase array system. This method operates with a large functioning depth because the transient elastic wave method possesses a high-energy feature, whereas the traditional ultrasonic method does not. As an alternative to using displacement responses, this method uses velocity responses to construct the SAFT image. Based on the numerical simulation and the experimental results, using the proposed method allowed a high image quality to be produced and successfully revealed d...

This paper discusses the feasibility of detecting defects inside reinforced concrete (RC) structures by using multi-directional Synthetic Aperture Focusing Technique (SAFT) to create an image which contains the comprehensive information about the geometry of possible defects. This is achieved by scanning the structure in multiple directions and processing the response data with SAFT and proposed scheme so that the shape and the position of a defect or defects can be completely described. The demonstration was carried out in numerical and experimental aspects. Furthermore the correlation between the numerical and experimental results was analyzed in this paper.

To explore the growth inhibitory effect of quercetin on imatinib-resistant chronic myeloid leukemia cell lines and to clarify its involved mechanisms.The cell viability was detected by trypan blue Staining, percentage of apoptotic cells and cell cycle distribution were detected by flow cytometry, the protein expression was detected by Western blot.Both inhibitory effect of proliferation and apoptosis-inducing effect were similar between the imatinib-resistant and -sensitive cell lines treated with 25 µmol/L quercetin for 24 hours and with arrest of cell cycle at GThe growth of imatinib-resistant cells can be inhibited by quercetin, and the apoptosis of cells can be induced by quercetin, which may be related to cell cycle arrest in G

It has been found that sertraline, a widely used antidepressant drug, possessed antitumor roles in a variety of cancers including liver cancer, colorectal cancer and lymphoma. In this study, we provided evidences that sertraline had potent antiproliferative activity not only in acute myeloid leukemia (AML) cell lines but also in the fresh leukemia cells from AML patients, and could induce cell death through both apoptosis and autophagy pathways. Moreover, we found that inhibiting autophagy pathway could partially attenuate sertraline-induced apoptosis and cell growth inhibition, indicating that sertraline-induced autophagy process could facilitate AML cell apoptosis to some degree. However, blocking apoptosis pathway seemed no obvious effects on sertraline-caused autophagy as well as cell growth inhibition. Our results suggested a potential application value of sertraline in the treatment of AML patients, furnishing some perspectives for novel therapeutic strategies in leukemia.

Using elastic-wave-based NDT methods to detect defects in RC structures is difficult because of the interference of rebars. In this study, the interference effect on the quality of SAFT images was quantitatively explored using a series of numerical simulations. Furthermore, two concrete specimens were fabricated for experimental verification. The experimental results exhibited strong agreement with those of quantitative analyses used in numerical simulations. These results indicated that using SAFT images to expose the defects under a layer of rebars is feasible. Moreover, a useful guideline for selecting the impacting source to produce a high-quality SAFT image is proposed.

Depression symptoms are now recognized to be the predominant cause of disability for bipolar disorder (BD) patients. The treatment strategies for the depressed phase of BD remain more anecdotal than data-based. Olanzapine has been investigated as an alternative to antidepressants and a mood stabilizer for acute bipolar depression. The purpose of this study was to assess the efficacy of olanzapine monotherapy for bipolar I depression. Sixty-eight patients with bipolar I depression were randomly assigned to treatment with olanzapine (mean final dose 14.4 mg/day) (n = 34) or placebo (n = 34) in a double-blind parallel-group study design. Planned assessments included Montgomery-Asberg Depression Rating Scale (MADRS), Young Mania Rating Scale (YMRS), Clinical Global Impressions-Severity of Illness scale (CGI-S), Clinical Global Impressions-Improvement scale (CGI-I), Hamilton Depression scale (HAMD), Hamilton Anxiety scale (HAMA), and Treatment Emergent Symptom Scale (TESS). Of the 68 patients who were randomly assigned, 57 (83.8 %) completed treatments. Improvements in MADRS total score, CGI-S, CGI-I, and HAMD in the olanzapine group were significantly greater relative to those in the placebo group during the 8-week follow-up period (p

We previously showed that Rig-G, an antiproliferative protein induced by interferon, can sequester CSN5 protein in the cytoplasm. Here, we report that Rig-G can destroy the functions of CSN5-containing COP9 signalosome (CSN), a highly conserved multiprotein complex implicated in protein deneddylation, deubiquitination, and phosphorylation. By damaging integrity and stability of the CSN complex, Rig-G can dramatically reduce the cellular content of CSN complex and inhibit its regulatory roles in assembly and activation of cullin-RING ubiquitin E3 ligases (CRL). Furthermore, Rig-G can cause excessive activation of CRL through inhibition of CSN-mediated deneddylation, largely decreasing protein levels of Cul1 and βTrCP, two important subunits of SCF (Skp1-Cul1-F-box protein)-E3 ligase. Rig-G can also attenuate the ability of CSN to recruit USP15 and impair CSN-associated deubiquitination. Increased autoubiquitination of βTrCP and concomitant accumulation of target substrates (such as IκBα) are observed in Rig-G-expressing cells. Taken together, our findings reveal for the first time the negative regulation of Rig-G on SCF-E3 ligase activities through disrupting CSN complex, not only contributing to further investigation on biological functions of Rig-G, but also leading to better understanding of the CSN complex as a potential target in tumor diagnosis and treatment.

Despite advances in the treatment of T‑cell acute lymphoblastic leukemia (T‑ALL), the outcome of T‑ALL treatment remains unsatisfactory, therefore, more effective treatment is urgently required. The present study examined the cytotoxicities of bortezomib in combination with daunorubicin against human Jurkat and Molt‑4 T‑ALL cells and primary T‑ALL cells. Compared with treatment alone, co‑exposure of cells to bortezomib and daunorubicin resulted in a significant increase in cell death in the Jurkat cells, as evidenced by the increased percentage of Annexin V‑positive cells, the formation of apoptotic bodies. In addition, the administration sequence of bortezomib and daunorubicin had an effect on cell viability. Treatment with bortezomib followed by daunorubicin treatment was more effective, compared with treatment with daunorubicin followed by bortezomib. Co-treatment with bortezomib and daunorubicin markedly enhanced the activation of caspase‑3, ‑8 and ‑9, which was reversed by the pan‑caspase inhibitor, Z‑VAD‑FMK. In addition, cotreatment with bortezomib and daunorubicin enhanced the collapse of mitochondrial transmembrane potential and upregulated the proapoptotic protein, B‑cell lymphoma 2 (Bcl‑2)‑interacting mediator of cell death (Bim), but not Bcl‑2 or Bcl‑extra large. Consistent with this, it was demonstrated that cotreatment of bortezomib and daunorubicin efficiently induced apoptosis in primary T‑ALL cells, and cell death was associated with the collapse of mitochondrial transmembrane potential and the upregulation of Bim. Taken together, these findings indicated that the combination of bortezomib and daunorubicin significantly enhanced their apoptosis‑inducing effect in T‑ALL cells, which may warrant further investigation in preclinical and clinical investigations.

Using nondestructive testing techniques to evaluate the length or depth of an existing foundation is an important issue with potential high application values. One of these is to evaluate whether the foundation is broken after severe earthquakes. In this aspect, academic research related to nondestructive evaluation for caisson foundations is rarely reported. The objective of this paper is to study the feasibility of using Sonic Echo method to evaluate the depth of caisson foundations. Two types of caissons, simple cylindrical caisson and compound caisson with chambers, were studied for their responses to the Sonic Echo tests. The study was carried out in numerical simulation with finite element method and experimental way with in-situ tests. A bridge system which spans over Sofong Brook in Taiwan was selected for the tests in situ. The bridge system is still under construction and therefore the effect of different construction stages on the testing results may be studied. In this paper, the parameters to be varied for the studies include the testing locations and the existence of chamber plates, the bottom plate and the top plate. Finally some preliminary conclusions can be reached for a successful test.

Objectives To evaluate the efficacy of metformin for treatment of antipsychotic-induced weight gain. Methods Seventy-two patients with first-episode schizophrenia who gained more than 7% of their predrug weight were randomly assigned to receive 1000 mg/d of metformin or placebo in addition to their ongoing treatment for 12 weeks using a double-blind study design. The primary outcome was change in body weight. The secondary outcomes included changes in body mass index, fasting glucose and insulin, and insulin resistance index. Results Of the 72 patients who were randomly assigned, 66 (91.6%) completed treatments. The body weight, body mass index, fasting insulin and insulin resistance index decreased significantly in the metformin group, but increased in the placebo group during the 12-week follow-up period. Significantly more patients in the metformin group lost their baseline weight by more than 7%, which was the cutoff for clinically meaningful weight loss. Metformin was tolerated well by majority patients. Conclusion Metformin was effective and safe in attenuating antipsychotic-induced weight gain and insulin resistance in first-episode schizophrenia patients. Patients displayed good adherence to metformin.

Background To investigate the validity of gastric cancers with nodes metastasis at Level II stations limited to No. 7 being classified as level-based n1 stage disease and the impact of this revision on lymph node staging. Methods Clinicopathologic features and prognosis of 1,606 node positive gastric cancers were retrospectively reviewed. Four patient groups were classified according to the status of node involvement: Group A, 734 patients with node metastasis at Level I stations; Group B, 317 patients with nodes metastasis at Level II stations limited to No. 7; Group C, 501 patients with nodes metastasis at Level II stations besides No. 7; and Group D, 54 patients with nodes metastasis at Level III stations. Results Although the extent of node metastasis for patients in Group B was more severe than that for patients in Group A, clinicopathologic features (especially pT stage) were not significantly different. Although overall survival for patients in Group B was significantly worse than that for patients in Group A, no significant differences in prognosis could be observed when stratified by pN or rN category. A revised level-based n category was established by considering cancers in Group B as level-based n1 stage disease. Multivariate analysis confirmed rN category and the revised level-based n category independently predicted patients' survival. A novel N category was established by combining rN category and the revised level-based n category. Further analysis revealed the novel N category had better homogeneity, discriminatory ability, and monotonicity of gradients than the other node categories, indicating the novel N system might be the most valuable node staging system for prognostic assessment. Conclusion It might be more suitable for cancers in Group B being classified as level-based n1 stage disease. And we recommend the anatomical location of metastatic lymph nodes also being considered in the categorization of lymph node metastasis. J. Surg. Oncol. 2012; 105:805–812. © 2011 Wiley Periodicals, Inc.

Background To clarify the biologic behavior of the early signet-ring cell cancers (SRCs) by comparing the clinicopathologic features and the incidence of lymph node metastasis between different histologic types of early gastric cancer (EGC) and to propose the indications of endoscopic surgery for SRCs. Methods Clinicopathologic features and the incidence of lymph node metastasis of 422 EGCs were retrospectively reviewed and compared according to the histologic type. Results Clinicopathologic features, incidence of node metastasis, prognosis, as well as the incidence of recurrence for SRCs, were similar to those of differentiated cancers (DCs), however, significantly different from those of undifferentiated cancers (UDCs). Tumor size, histologic type, lymphatic and/or blood vessel invasion (LBVI), and depth of invasion were independent factors predicting node metastasis for EGCs. For DCs and SRCs with mucosal invasion and ≤2 cm in diameter without LBVI, no metastatic lymph node was detected (95% CI, 0–5.0). Also, for DCs and SRCs with mucosal invasion and >2 cm in diameter without LBVI, or with submucosal invasion and ≤2 cm in diameter without LBVI, no metastatic lymph node was detected (95% CI, 0–3.0). Conclusion Clinicopathologic features of SRCs were similar with DCs, but different from other UDCs. Consequently, the treatment strategy for SRCs might be similar with that for DCs. According to the incidence of node metastasis, we propose SRCs with mucosal invasion without LBVI, or with submucosal invasion and ≤2 cm in diameter without LBVI, might be suitable for endoscopic surgery.

We previously reported that IRF-9/STAT2 functional interaction could drive the expression of retinoic acid-induced gene G (RIG-G), independently of STAT1 and the classical JAK-STAT pathway, providing a novel alternative pathway for interferons (IFN) to mediate their multiple biological properties. In addition, we also found that IRF-1 could regulate RIG-G induction as well as the expression of IRF-9 and STAT2 in some cases. But the mechanisms by which IRF-1 exerted its action remained to be elucidated. Here, we showed that STAT1 could significantly enhance the effects of the IRF-9/STAT2 complex or IRF-1 on RIG-G induction through an activated JAK-STAT pathway, though it was not essential for RIG-G expression. In STAT1-deficient U3A cells, IRF-1 could induce RIG-G expression via the IFN-stimulated response elements in the RIG-G gene promoter, but it failed to upregulate IRF-9 and STAT2 unless the U3A cells were reconstituted by exogenous STAT1. In STAT1-expressing cells, IRF-1 indirectly activated RIG-G expression through an IRF-9/STAT2-dependent manner. Taken together, we concluded that the expression of RIG-G was independent on the classical JAK-STAT pathway, but could be greatly increased by it. This work will be of great benefit to us for a better understanding of the mechanisms on RIG-G gene expression regulation.

Retinoic acid–induced gene G (RIG-G), a gene originally identified in all-trans retinoic acid–treated NB4 acute promyelocytic leukemia cells, is also induced by IFNα in various hematopoietic and solid tumor cells. Our previous work showed that RIG-G possessed a potent antiproliferative activity. However, the mechanism for the transcriptional regulation of RIG-G gene remains unknown. Here, we report that signal transducer and activator of transcription (STAT) 2 together with IFN regulatory factor (IRF)-9 can effectively drive the transcription of RIG-G gene by their functional interaction through a STAT1-independent manner, even without the tyrosine phosphorylation of STAT2. The complex IRF-9/STAT2 is both necessary and sufficient for RIG-G gene expression. In addition, IRF-1 is also able to induce RIG-G gene expression through an IRF-9/STAT2–dependent or IRF-9/STAT2–independent mechanism. Moreover, the induction of RIG-G by retinoic acid in NB4 cells resulted, to some extent, from an IFNα autocrine pathway, a finding that suggests a novel mechanism for the signal cross-talk between IFNα and retinoic acid. Taken together, our results provide for the first time the evidence of the biological significance of IRF-9/STAT2 complex, and furnish an alternative pathway modulating the expression of IFN-stimulated genes, contributing to the diversity of IFN signaling to mediate their multiple biological properties in normal and tumor cells. [Cancer Res 2009;69(8):3673–80]

Seawater drowning-induced acute lung injury (ALI) is a serious clinical condition characterized by increased alveolar-capillary permeability, excessive inflammatory responses, and refractory hypoxemia. However, current therapeutic options are largely supportive; thus, it is of great interest to search for alternative agents to treat seawater aspiration-induced ALI. Erythropoietin (EPO) is a multifunctional agent with antiinflammatory, antioxidative, and antiapoptotic properties. However, the effects of EPO on seawater aspiration-induced ALI remain unclear. In the present study, male rats were randomly assigned to the naive group, normal saline group, seawater group, or seawater + EPO group. EPO was administered intraperitoneally at 48 and 24 h before seawater aspiration. Arterial blood gas analysis was performed with a gas analyzer at baseline, 30 min, 1 h, 4 h, and 24 h after seawater aspiration, respectively. Histological scores, computed tomography scan, nuclear factor kappa B p65, inducible nitric oxide synthase, caspase-3, tumor necrosis factor-alpha, interleukin (IL)-1β, IL-6, IL-10, wet-to-dry weight ratio, myeloperoxidase activity, malondialdehyde, and superoxide dismutase in the lung were determined 30 min after seawater aspiration. Our results showed that EPO pretreatment alleviated seawater aspiration-induced ALI, as indicated by increased arterial partial oxygen tension and decreased lung histological scores. Furthermore, EPO pretreatment attenuated seawater aspiration-induced increase in the expressions of pulmonary nuclear factor kappa B p65, inducible nitric oxide synthase, caspase-3, tumor necrosis factor-alpha, IL-1β, myeloperoxidase activity, and malondialdehyde when compared with the seawater group. Collectively, our study suggested that EPO pretreatment attenuates seawater aspiration-induced ALI by down-regulation of pulmonary pro-inflammatory cytokines, oxidative stress, and apoptosis.

To investigate the biological effects of sertraline, one of psychotropic drugs, on actue myeloid leukemia cell line Kasumi-1.Cells were treated by different concentrations of sertraline for different times. The effects of sertraline were evaluated by cell growth, cell morphology, cell cycle distribution and markers of cell apoptosis, respectively. Western blot was used to detect the expression change of related proteins.Sertraline could inhibit cell proliferation and induce apoptosis. After treatment with 15 µmol/L and 20 µmol/L sertraline for 24 h, the inhibitory rate of Kasumi-1 cell proliferation was (19.00 ± 7.37)% and (47.90 ± 11.19)%, respectively. Meanwhile, compared with the control group, the percentage of Annexin V positive cells in Kasumi-1 cells treated with sertraline for 24 h raised obviously from (9.71 ± 2.12)% to (20.54 ± 2.52)% and (45.37 ± 7.88)% (P0.01), respectively. The cells were arrested in G0/G1 and G2/M phase. In addition, it was found that sertraline could also down-regulate the level of translationally controlled tumor protein (TCTP) in Kasumi-1 cells.Sertraline can significantly induce the apoptosis of Kasumi-1 cells, that probably is associated with the down-regulation of TCTP protein expression.

RIG-G (retinoic acid-induced gene G) was originally identified in ATRA (all-trans retinoic acid)-treated NB4 acute promyelocytic leukemia (APL) cells. It was induced to expression by ATRA along with the differentiation of the cells. However, little is known about its role(s). Here, we established a RIG-G stably expression transformant of NB4 cells. By using the transformant, we showed that expression of RIG-G in NB4 cells not only arrested the cells at G1/G0 transition phase and inhibited their proliferation, but also markedly drive the maturation of NB4 cells in the presence of very low concentration of ATRA (10(-9)mol/L). What's more, by detecting the expression of RIG-G in fresh primary bone marrow mononuclear cells of APL patients in different morbid states, we found high RIG-G expression level in complete remission patients, while low level in untreated or relapsed patients. These results indicated that RIG-G level was high in maturated cells and low in blast cells, and suggested that RIG-G might play a role in the differentiation of bone marrow hemocytes in vivo.

Sepsis-associated encephalopathy (SAE) is a frequent complication in critically ill patients and is associated with long-term cognitive impairments. However, the pathophysiology underlying SAE is poorly understood and the pharmacologic treatment is lacking. The purpose of the present study was to investigate the effects of erythropoietin (EPO) on cognitive impairments in an animal model of SAE induced by peripheral administration of lipopolysaccharide (LPS). Mice were randomly divided into the sham + vehicle, sham + EPO, LPS + vehicle, and LPS + EPO groups. EPO was administrated 30 min after the LPS administration and daily afterward for 2 days. Behavioral tests were performed on days 6 and 7 with open field and fear conditioning tests, respectively. The survival rate was estimated by the Kaplan–Meier method. The levels of proinflammatory responses, oxidative stress, and apoptosis-related markers were measured in the hippocampus at the indicated time points. The synaptic morphometry changes in the CA1 region were observed with transmission electron microscopy. Our results showed that LPS administration resulted in high mortality rate and cognitive impairments, which were accompanied by increased expressions of interleukin-1β, malondialdehyde, cleaved caspase-3, and abnormal synaptic morphometry changes in the hippocampus. Notably, EPO treatment reversed the cognitive impairments and rescued the brain pathology induced by LPS administration. In conclusion, our data suggested that treatment with EPO reduced the mortality rate and ameliorated cognitive impairments in an animal model of SAE.

TheRIG-Ggene, originally isolated from an acute promyelocytic leukemia cell line NB4, codes for a 60-kDa cytoplasmic protein that is induced byall-transretinoic acid (ATRA) treatment along with the induction of morphological differentiation of NB4 cells. Here, we provide evidence that ectopic expression of Rig-G in U937 cells can lead to a significant accumulation of cells at G1/S transition. Growth arrest seems to occur by modulating several major cell cycle regulatory players. Interestingly, Rig-G alters JAB1 cellular distribution through interacting with this protein and increases the intracellular level of p27 by preventing it from the JAB-1-dependent and ubiquitin/proteasome-mediated degradation. Furthermore, we demonstrate a role of Rig-G for c-myc down-regulation that results in an up-regulation of p21, tightly associated with cell cycle arrest. In addition, our studies reveal that Rig-G is a direct target of STAT1, a key transcription factor in regulating IFN responses, and may be one of the first experimentally proven molecular mediators for the antiproliferative effect of IFN-α. Considering that IFN-α and ATRA synergistically inhibit growth along the intracellular pathways triggered by the two compounds in many cell types, we suggest that Rig-G may also represent one of the key molecular nodes of signaling cross-talk between ATRA and IFN-α.

The involvement of telomerase in cellular immortalization and senescence has often been assessed by means of telomerase expression at the RNA level and quantification of telomerase activity by the telomeric repeat amplification protocol assay. However, these methods either neglected the existence of various telomerase splice variants, or ignored the nonconventional functions of telomerase independent of its ability to elongate and maintain telomere length. Immunodetection of telomerase is now being recognized as a necessary approach to precisely elucidate its roles in oncogenesis and senescence. A few antibodies directed against the catalytic subunit of the human telomerase (hTERT) are currently used but their specificity is not always demonstrated. A survey of the literature showed inconsistencies and led us to comparatively re-evaluate the most frequently used antibodies. Surprisingly, mass spectrometry, two-dimensional gel analysis and immunofluorescent experiments revealed that the most frequently used hTERT immunoprobe, a mouse monoclonal antibody that was claimed to be directed against an hTERT protein epitope, in fact recognizes nucleolin rather than telomerase. Our findings have interesting implications regarding the biology of nucleolin and telomerase in the context of pathophysiological investigations recently carried out.

The theoretical capabilities of the Parallel Seismic (PS) test for determining the length of piles and characterizing possible defects are investigated in this paper. In deriving the theory, a correction factor is proposed in this paper to enhance the accuracy of the prediction. An axisymmetric finite element (FE) model was developed to carry out a series of parametric studies which included the effects of the pile length, the pile-to-borehole distance and the stiffness of surrounding soil. A miniature experiment using an aluminum bar embedded in epoxy prism was also designed and tested in the lab to verify the theory and to demonstrate the use of the correction factor. The results from the experiment and the numerical FE model were also compared to expose the potential of this nondestructive method to in situ application. The model was further modified to simulate the PS test on defective piles with axisymmetric necks and bulges. It can be concluded that the geometric configuration of a pile and the possible significant defect can be characterized with this nondestructive test.

The velocity of elastic wave is usually the essential property that has to be measured in advance, when applying the civil NDT technique based on the theory of transient elastic wave. The accuracy and handiness of the velocity measuring method are often become a key about the practicability of the NDT method. In this paper, based on the transient elastic wave theory, an elastic-wave-velocity measurement system was developed. The interdisciplinary techniques related to civil engineering, electronic engineering, mechanical engineering, and computer science were integrated to make this system more stable and easier to operate on the construction site. The processes during the measurement such as data capturing, signal processing and even data analysis are accomplished by the controlling software automatically. The measurement will be completed in seconds. Therefore, lots of data could be gathered in very short time and then used to estimate the quality of concrete. Besides, the wireless data transfer technology was also adopted in this system. With this function, more experts could monitor the measurement simultaneously through the Internet. In the preliminary experiment, the result shows good correlation between the wave velocity and the compression strength. It demonstrates the potential of this system on the application of NDT technology.

Arsenic trioxide (As(2)O(3)), an effective drug for the treatment of acute promyelocytic leukemia (APL), can induce apoptosis and partial differentiation in APL cells in vitro and in vivo. However, As(2)O(3) also induces apoptosis in cancer cells other than APL with complex mechanisms, which seem to be cell type dependent. In this study, we report that APL cells (NB4 cell line) are arrested at early mitotic phase before the collapse of mitochondrial transmembrane potential (Deltavarphi(m)) and apoptosis after treatment with pharmacological concentrations (1.0-2.0 micro M) of As(2)O(3). We have also made the following new discoveries: (1) 0.5 micro M As(2)O(3) that fails to induce apoptosis has no effects on cell cycle distribution. (2) With inhibition of As(2)O(3)-induced Deltavarphi(m) collapse and apoptosis, dithiothreitol also effectively inhibits As(2)O(3)-induced mitotic arrest, suggesting that both As(2)O(3)-induced apoptosis and mitotic arrest involve proteins with thiol groups. (3) 1.5 mM caffeine that relieves cells from G(2)/M arrest also inhibits As(2)O(3)-induced Deltavarphi(m) collapse and apoptosis, (4) 1.0 micro M As(2)O(3) increases the expression of both cyclin B(1) and hCDC20 whereas it inhibits Tyr15 phosphorylation of p34(cdc2). In conclusion, our results strongly support that there is a tight link between As(2)O(3)-induced apoptosis and mitotic arrest, the latter being one of common mechanisms for As(2)O(3)-induced apoptosis in cancer cells.

Most acute promyelocytic leukemia (APL) cases have t(15;17)(q22;q21) chromosomal translocation and PML-RARalpha chimeric gene which blocks granulocytic differentiation. The introduction of all-trans-retinoic acid (ATRA) and arsenic compounds, especially arsenic trioxide (As(2)O(3)), has provided good models to study not only differentiation and/or apoptosis therapy but also molecular target-based cancer treatment. In vivo and in vitro investigations have shown that both agents are able to induce differentiation of APL cells: ATRA tends to induce terminal differentiation, while low-dose As(2)O(3) can induce partial differentiation. Significant progress has been made in understanding the molecular mechanisms of APL pathogenesis and differentiation therapy. Pharmacological concentrations (0.1 approximately 1 microM) of ATRA derepresses transcription by releasing CoR from, and recruiting CoA to PML-RARalpha, whereas As(2)O(3) triggers a rapid degradation of PML-RARalpha. In fact, the two drugs act on the same oncoprotein through targeting different moieties and in distinct ways and thereby abrogate its dominant-negative effects on regulatory pathways necessary for granulocytic differentiation. As to apoptosis, it is clear that high-dose As(2)O(3) can induce mitochondria-mediated cell death pathway in a thiol-dependent manner, while the mechanism of ATRA-induced apoptosis needs further elucidation. Transcriptomic and proteomic analysis are also expected to find new molecular targets. It is the hope that what we have learnt from APL will benefit further developments of anti-leukemia therapy.

Acute promyelocytic leukemia (APL) is characterized by the specific chromosome translocation t(15;17) with promyelocytic leukemia-retinoic acid receptor-α (PML-RARA) fusion gene and the ability to undergo terminal differentiation as an effect of all-trans retinoic acid (ATRA). Recently, arsenic trioxide (As2O3) has been identified as an alternative therapy in patients with both ATRA-sensitive and ATRA-resistant APL. At the cellular level, As2O3 triggers apoptosis and a partial differentiation of APL cells in a dose-dependent manner; both effects are observed in vivo among patients with APL and APL animal models. To further explore the mechanism of As2O3-induced differentiation, the combined effects of arsenic and a number of other differentiation inducers on APL cell lines (NB4 and NB4-R1) and some fresh APL cells were examined. The data show that a strong synergy exists between a low concentration of As2O3 (0.25 μM) and the cyclic adenosine monophosphate (cAMP) analogue, 8-CPT-cAMP, in fully inducing differentiation of NB4, NB4-R1, and fresh APL cells. Furthermore, cAMP facilitated the degradation of As2O3-mediated fusion protein PML-RARα, a process considered to play a key role in overcoming the differentiation arrest of APL cells. On the other hand, cAMP could significantly inhibit cell growth by modulating several major players in G1/S transition regulation. Interestingly, H89, an antagonist of protein kinase A, could block the differentiation-inducing effect of As2O3potentiated by cAMP. These results thus support the existence of a novel signaling cross-talk for APL maturation, which may deepen understanding of As2O3-induced differentiation in vivo, and thus furnish insights for new therapeutic strategies.

To elucidate the molecular mechanism of all-trans-retinoic acid (ATRA)–induced differentiation of acute promyelocytic leukemia (APL) cells, the gene expression patterns in the APL cell line NB4 before and after ATRA treatment were analyzed using complementary DNA array, suppression-subtractive hybridization, and differential-display–polymerase chain reaction. A total of 169 genes, including 8 novel ones, were modulated by ATRA. The ATRA-induced gene expression profiles were in high accord with the differentiation and proliferation status of the NB4 cells. The time courses of their modulation were interesting. Among the 100 up-regulated genes, the induction of expression occurred most frequently 12-48 hours after ATRA treatment, while 59 of 69 down-regulated genes found their expression suppressed within 8 hours. The transcriptional regulation of 8 induced and 24 repressed genes was not blocked by cycloheximide, which suggests that these genes may be direct targets of the ATRA signaling pathway. A balanced functional network seemed to emerge, and it formed the foundation of decreased cellular proliferation, maintenance of cell viability, increased protein modulation, and promotion of granulocytic maturation. Several cytosolic signaling pathways, including JAKs/STAT and MAPK, may also be implicated in the symphony of differentiation.

The promyelocytic leukemia zinc finger gene ( PLZF ) is involved in chromosomal translocation t(11;17) associated with acute promyelocytic leukemia. In this work, a 201-kilobase genomic DNA region containing the entire PLZF gene was sequenced. Repeated elements account for 19.83%, and no obvious coding information other than PLZF is present over this region. PLZF contains six exons and five introns, and the exon organization corresponds well with protein domains. There are at least four alternative splicings (AS-I, -II, -III, and -IV) within exon 1. AS-I could be detected in most tissues tested whereas AS-II, -III, and -IV were present in the stomach, testis, and heart, respectively. Although splicing donor and acceptor signals at exon–intron boundaries for AS-I and exons 1–6 were classical (gt–ag), AS-II, -III, and -IV had atypical splicing sites. These alternative splicings, nevertheless, maintained the ORF and may encode isoforms with absence of important functional domains. In mRNA species without AS-I, there is a relatively long 5′ UTR of 6.0 kilobases. A TATA box and several transcription factor binding sites were found in the putative promoter region upstream of the transcription start site. PLZF is a well conserved gene from Caenorhabditis elegans to human. PLZF paralogous sequences are found in human genome. The presence of two MLL/PLZF- like alignments on human chromosome 11q23 and 19 suggests a syntenic replication during evolution. The chromosomal breakpoints and joining sites in the index acute promyelocytic leukemia case with t(11;17) also were characterized, which suggests the involvement of DNA damage-repair mechanism.