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Pinus koraiensis polysaccharides (PKP) were extracted by hot water from P. koraiensis pine cones. Five polysaccharide fractions named PKP-A, PKP-B, PKP-C, PKP-D and PKP-E were successfully separated at final ethanol concentrations of 30%, 50%, 60%, 70% and 80%, respectively. HPLC, FT-IR, GC-MS and automatic amino-acid analysis were applied to investigate their chemical characteristics. Monosaccharide component analysis indicated that the five fractions were all composed of D-ribose, L-rhamnose, L-arabinose, D-xylose, D-mannose, D-glucose and D-galactose, but their molar ratios were quite different. HPLC results revealed that the polysaccharides precipitated by higher concentrations of ethanol solution had lower molecular masses. Moreover, the antioxidant activities of the five fractions were studied on the basis of hydroxyl radical and ABTS radical scavenging tests. The five graded polysaccharide fractions exhibited good inhibitory power, and MTT tests in vitro showed the IC50 of PKP-A and PKP-E were 1,072.5 and 2,070.0 μg · mL-1, respectively. These results demonstrated that the PKP could be a potential source of natural antioxidants or dietary supplements.

Kaempferol is a natural flavonoid that possesses anti-proliferative and apoptosis-inducing activities in several cancer cell lines. In the present study, we investigated the anti-metastatic activity of kaempferol and its molecular mechanism(s) of action in human osteosarcoma cells. Kaempferol displayed inhibitory effects on the invasion and adhesion of U-2 osteosarcoma (OS) cells in a concentration-dependent manner by Matrigel Transwell assay and cell adhesion assay. Kaempferol also inhibited the migration of U-2 OS cells in a concentration-dependent manner at different treatment time points by wound-healing assay. Additional experiments showed that kaempferol treatment reduced the enzymatic activities and protein levels of matrix metalloproteinase (MMP)-2, MMP-9 and urokinase plasminogen activator (uPA) by gelatin and casein-plasminogen zymography assays and western blot analyses. Kaempferol also downregulated the mRNA levels of MMP-2 and MMP-9 by quantitative PCR analyses. Furthermore, kaempferol was able to reduce the protein phosphorylation of ERK, p38 and JNK by western blotting. By electrophoretic mobility-shift assay (EMSA), we demonstrated that kaempferol decreased the DNA binding activity of AP-1, an action likely to result in the reduced expression of MMP-2, MMP-9 and uPA. Collectively, our data showed that kaempferol attenuated the MAPK signaling pathways including ERK, JNK and p38 and resulted in the decreased DNA binding ability of AP-1, and hence, the downregulation in the expression and enzymatic activities of MMP-2, MMP-9 and uPA, contributing to the inhibition of metastasis of U-2 OS cells. Our results suggest a potential role of kaempferol in the therapy of tumor metastasis of OS.

Radish floral bud abortion (FBA) is an adverse biological phenomenon that occurs during reproduction. Although FBA is a frequent occurrence, its molecular mechanism remains unknown. A transcript-derived fragment (TDF72), which was obtained by cDNA amplified fragment length polymorphism (cDNA-AFLP), was up-regulated in the aborted buds and exhibited 89% sequence homology with the AtγVPE gene. In this study, TDF72 was used to clarify the role of VPE in FBA by isolation of the VPE gene RsVPE1 from radish flower buds. The full-length genomic DNA was 2346 bp including nine exons and eight introns. The full-length cDNA was 1825 bp, containing a complete open reading frame (ORF) of 1470 bp, which encoded a predicted protein containing 489 amino acid residues, with a calculated molecular mass of 53.735 kDa. Expression analysis demonstrated that RsVPE1 was expressed in all tested organs of radish at different levels. Highest expression was detected in aborted flower buds, suggesting that RsVPE1 has a role in FBA. In order to analyze the role of RsVPE1 in FBA, RsVPE1 was overexpressed in transgenic Arabidopsis thaliana plants. Aborted flower buds appeared in transgenic plants subjected to heat stress. In addition, RsVPE1 expression in the transgenic plants reached a maximum when subjected to heat stress for 24 h and increased by 2.1-fold to 2.8-fold in three homozygous transgenic lines. These results indicated that RsVPE1 led to FBA when its expression levels exceeded a particular threshold, and provided evidence for the involvement of RsVPE1 in promoting FBA under heat stress.

Kaempferol belongs to the flavonoid family and has been used in traditional folk medicine. Here, we investigated the antitumor effects of kaempferol on cell cycle arrest and autophagic cell death in SK-HEP-1 human hepatic cancer cells. Kaempferol decreased cell viability as determined by MTT assays and induced a G2/M phase cell cycle arrest in a concentration-dependent manner. Kaempferol did not induce DNA fragmentation, apoptotic bodies or caspase-3 activity in SK-HEP-1 cells as determined by DNA gel electrophoresis, DAPI staining and caspase-3 activity assays, respectively. In contrast, kaempferol is involved in the autophagic process. Double-membrane vacuoles, lysosomal compartments, acidic vesicular organelles and cleavage of microtubule-associated protein 1 light chain 3 (LC3) were observed by transmission electron microscopy, LysoΤracker red staining, GFP-fluorescent LC3 assays and acridine orange staining, respectively. In SK-HEP-1 cells, kaempferol increased the protein levels of p-AMPK, LC3-II, Atg 5, Atg 7, Atg 12 and beclin 1 as well as inhibited the protein levels of CDK1, cyclin B, p-AKT and p-mTOR. Taken together, CDK1/cyclin B expression and the AMPK and AKT signaling pathways contributed to kaempferol-induced G2/M cell cycle arrest and autophagic cell death in SK-HEP-1 human hepatic cancer cells. These results suggest that kaempferol may be useful for long-term cancer prevention.

Baicalin is one of the major compounds in the traditional Chinese medicinal herb from Scutellaria baicalensis Georgi. We investigated the molecular mechanisms of cell autophagy induced by baicalin in human bladder cancer T24 cells. Baicalin inhibited cell survival as shown by MTT assay and increased cell death by trypan blue exclusion assay in a concentration-dependent manner. Baicalin did not induce apoptotic cell death in T24 cells by TUNEL and caspase-3 activity assay. Baicalin induced the acidic vesicular organelle cell autophagy marker, manifested by acridine orange (AO) and monodansylcadaverine (MDC) staining and cleavage of microtubule-associated protein 1 light chain 3 (LC3). The protein expression levels of the Atg 5, Atg 7, Atg 12, Beclin-1 and LC3-II were upregulated in T24 cells after baicalin treatment. Inhibition of autophagy by 3-methyl-adenine (an inhibitor of class III phosphatidylinositol-3 kinase; 3-MA) reduced the cleavage of LC3 in T24 cells after baicalin treatment. Furthermore, protein expression levels of phospho-AKT (Ser473) and enzyme activity of AKT were downregulated in T24 cells after baicalin treatment. In conclusion, baicalin triggered cell autophagy through the AKT signaling pathway in T24 cells.

Rationale and Objectives: The scoring algorithm of the 12-item Short-Form Health Survey (SF-12) was revised in the second version (SF-12v2), but information on its psychometric properties is lacking. This study determined whether the SF-12v2 was a valid and equivalent substitute for the SF-36v2 Health Survey (version 2) for the Chinese., Methods: A total of 2410 Chinese adults in Hong Kong completed the SF-36 Health Survey by telephone. The SF-12v2 data were extracted from the SF-36 data. Internal consistency was assessed by Cronbach's alpha, and test-retest reliabilities were evaluated by intraclass correlation. Criterion validity and equivalence were assessed using the SF-36v2 scores as a gold standard. Construct validity and sensitivity were assessed by known-group comparison., Results: Internal consistency and test-retest reliabilities were good (range 0.67-0.82) for all except three scales. The SF-12v2 summary scores explained >80% of the total variances of the SF-36v2 summary scores. Construct validity and sensitivity were confirmed by significantly lower SF-12v2 scores in people with chronic diseases than those without. Effect size differences were less than 0.3 and relative validities were greater than 0.7 between SF-12v2 and SF-36v2 scores for different groups., Conclusion: The SF-12v2 was valid, reliable and sensitive for the Chinese. It is an equivalent substitute for the SF-36v2 for the summary scales., (© 2011 Blackwell Publishing Ltd.)

This research focused on a Chinese herb medicine, Solanum lyratum Thunb (Solanaceae) by ethanol extracts (SLE) for investigating the molecular anticancer mechanism in vitro for exploring the means of cell death through the effects on mitochondrial function. We found that SLE induced cytotoxic effects in human osteosacroma U-2 OS cells, and these effects include cell morphological changes, a decrease of the percentage of viable cells and induction of apoptosis. The results suggest that cell death induced by SLE is closely related to apoptosis based on the observations of DAPI staining and sub-G1 phase in U-2 OS cells. Flow cytometric assays also showed that SLE promoted the production of reactive oxygen species and nitric oxide but decreased the levels of mitochondrial membrane potential and promoted the activations of caspase-8 and -9 in U-2 OS cells. SLE inhibited the level of Bcl-2 but promoted the Bax level, and both proteins led to the release of cytochrome c from mitochondria to cytosol and activation of caspase-9 and -3, resulting in the apoptotic death which is mediated through the mitochondrial pathway. Taken together, SLE was demonstrated to be effective in killing U-2 OS osteosacroma cells via the ROS-promoted and mitochondria- and caspase-dependent apoptotic pathways.

The histone methyltransferase EZH2 (enhancer of zeste homolog 2) plays critical roles in prostate cancer (PCa) development and is a potential target for PCa treatment. Triptolide possesses anti-tumor activity, but it is unknown whether its therapeutic effect relates with EZH2 in PCa. Here we described EZH2 as a target for Triptolide in PCa cells. Our data showed that Triptolide suppressed PCa cell growth and reduced the expression of EZH2. Overexpression of EZH2 attenuated the Triptolide induced cell growth inhibition. Moreover, Triptolide treatment of PC-3 cells resulted in elevated mRNA levels of target genes (ADRB2, CDH1, CDKN2A and DAB2IP) negatively regulated by EZH2 as well as reduced mRNA levelsan of EZH2 positively regulated gene (cyclin D1). Our findings suggest the PCa cell growth inhibition mediated by Triptolide might be associated with downregulation of EZH2 expression and the subsequent modulation of target genes.

Allyl isothiocyanate (AITC), a member of the isothiocyanate (ITC) family found in a constituent of cruciferous vegetables, possesses anticancer activity and induces apoptosis in various types of human cancer cell lines. However, no available information showed antitumor effects in human breast adenocarcinoma cells. The current study was focused on exploring the mechanisms underlying AITC-induced apoptosis in MDA-MB-468 human breast cancer cells in vitro. We found that AITC reduced the cell number and viability using trypan blue staining with the Countess Automated Cell Counter and the MTT assay, respectively. AITC also was found to induce apoptotic cell morphological changes by a contrast-phase microscope and cell cycle arrest at G(2)/M phase by flow cytometric assay in MDA-MB-468 cells. Intrinsic apoptosis-associated factors such as caspase-9 and caspase-3 activities were performed, and reactive oxygen species (ROS) production, loss of mitochondrial membrane potential (∆Ψm) occurred in AITC-treated MDA-MB-468 cells. AITC also stimulated mitochondria-related signaling, including p-Bcl-2 (Ser-70), cytochrome c and Apaf-1 in MDA-MB-468 cells. We found that the p-ERK signal was upregulated in AITC-treated cells. Importantly, NAC (a ROS scavenger) and U0126 (an ERK inhibitor) abolished AITC-reduced viability in MDA-MB-468 cells. AITC downregulated CDK1 activity and altered the expression of G(2)/M phase-modulated associated protein levels by western blotting in MDA-MB-468 cells. In summary, our findings demonstrated that AITC-promoted G2/M phase and AITC-triggered apoptosis correlate with the activation of phosphorylation of ERK in MDA-MB-468 cells. AITC is a potential agent for applcation in the treatment of human breast cancer.

The traditional method of gas chromatography-mass spectrometry for monosaccharide component analysis with pretreatment of acetylation is described with slight modifications and verified in detail in this paper. It was then successfully applied to the quantitative analysis of component monosaccharides in polysaccharides extracted from the pine cones. The results demonstrated that the three pine cone polysaccharides all consisted of ribose, rhamnose, arabinose, xylose, mannose, glucose and galactose in different molar ratios. According to the recovery experiment, the described method was proved accurate and practical for the analysis of pine cone polysaccharides, meeting the need in the field of chemical analysis of Pinus plants. Furthermore; the chemical characteristics, such as neutral sugar, uronic acids, amino acids, molecular weights, and antioxidant activities of the polysaccharides were investigated by chemical and instrumental methods. The results showed that the chemical compositions of the polysaccharides differed from each other, especially in the content of neutral sugar and uronic acid. In the antioxidant assays, the polysaccharide fractions exhibited effective scavenging activities on ABTS radical and hydroxyl radical, with their antioxidant capabilities decreasing in the order of PKP > PAP > PSP. Therefore, although the polysaccharide fractions had little effect on superoxide radical scavenging, they still have potential to be developed as natural antioxidant agents in functional foods or medicine.

Two HPLC methods with diode array detection (HPLC-DAD) and electrospray ionization-mass spectrometry (HPLC-ESI/MS), respectively, were developed to investigate the differences of chemical constituents and their metabolism in gastrointestinal tract in vitro between two decoctions of crude and processed Glycyrrhizae radix. Total of eleven constituents (liquiritin apioside, liquiritin, licuraside, isoliquiritin, ononin, glycyrrhizin, liquiritigenin-7,4'-diglucoside, licorice saponin A3, 22β-acetoxylglycyrrhizic acid, licorice saponin G2, and yunganoside E2) were identified in the two decoctions, whereas lower contents of these constituents were usually found in the decoction of processed Glycyrrhizae Radix. [corrected] Furthermore, these constituents were metabolized into their respective aglycons in human intestinal bacteria juice, and the metabolism ratios were all higher in processed Glycyrrhizae Radix [corrected] decoction. No change was found in artificial gastric or intestinal juice. This study revealed that the processing can alter the contents of main constituents in crude G. radix and their metabolism in gastrointestinal tract, in which intestinal bacteria play an important role in the metabolism of licorice constituents.

Vitexin, a lignan compound, has been shown to exert apoptotic actions on human breast cancer cell lines and to have anti-inflammatory activities. Nevertheless, there is currently no study addressing the effects of vitexin on the induction of apoptosis in U937 human leukemia cells. The aim of this study was to determine the anticancer effects and molecular mechanisms of vitexin on U937 leukemia cells. We showed that vitexin can potently induce programmed cell death in U937 leukemia cell growth as well as morphological changes that were examined by MTT assay and phase contrast microscopy, respectively. The DNA content and the levels of mitochondrial membrane potential (∆Ψm) were determined by flow cytometric analysis. The cell cycle arrest-regulated and apoptosis-associated protein levels were measured by western blotting. Vitexin-triggered apoptosis was accompanied by a decrease of the level of ∆Ψm and the percentage of viability and provoked apoptosis in U937 cells. The downregulation of the protein level for Bcl-2 with the simultaneous upregulation of caspase-3 and -9 protein expression in U937 cells were observed after treatment with vitexin. Therefore, our data provide a potential mechanism for the chemopreventive activity of vitexin, and we suggest that vitexin may serve as a therapeutic agent for the treatment of human leukemia.

Lyme disease is the most common tick-borne disease in the United States and has a multitude of systemic effects. Infrequently, however, Lyme disease is seen to cause liver dysfunction. Dermatologists should be aware that early, disseminated borreliosis can present with multiple erythema migrans plaques and hepatitis.

Bufalin is the major component of Chan-Su (a traditional Chinese medicine, TCM) extracts from the venom of Bufo bufo gargarizan. In the present study, we investigated the pharmacological mechanisms of cell cycle arrest and autophagic cell death induced by bufalin in SK-HEP-1 human hepatocellular carcinoma cells in vitro. Bufalin inhibited cell survival by MTT assay and increased cell death by trypan blue exclusion assay in a concentration-dependent manner. In addition, bufalin induced G2/M phase arrest by reducing CDK1 activity. Bufalin triggered DNA fragmentation and apoptotic cell death in SK-HEP-1 cells by DNA gel electrophoresis, TUNEL and caspase-3 activity assay, while bufalin induced autophagic cell death by double-membrane vacuoles (transmission electron microscopy, TEM), acidic vesicular organelles (acridine orange staining) and cleavage of microtubule-associated protein 1 light chain 3 (LC3). Protein expression levels of cyclin A and B, CDK1, phospho-CDK1 (Thr161), Cdc25c, phospho-Cdc25c (Ser198), phospho-AKT (Thr308), phospho-AKT (Ser473), phospho‑mTOR (Ser2481) were downregulated. In contrast, protein expression levels of the Chk1, Wee1, LC3-II, Beclin-1, Atg 5, Atg 7 and Atg 12 were upregulated in SK-HEP-1 cells after bufalin treatment. Inhibition of autophagy by 3-methyladenine (an inhibitor of class III phosphatidylinositol-3 kinase; 3-MA) or bafilomycin A1 (an inhibitor of the vacuolar proton pump of lysosomes and endosomes) reduced the effect of bufalin on cell viability and enhanced the effect of bufalin on apoptosis. In conclusion, bufalin triggered autophagic cell death and G2/M phase arrest through the AKT/mTOR signaling pathway in SK-HEP-1 cells. Our findings showed that bufalin may be potentially efficacious in the treatment of human hepatocellular carcinoma.

Arsenic trioxide (As₂O₃) is used clinically to treat acute promyelocytic leukemia (APL) and has activity in vitro for induction of apoptosis in several solid tumor cell lines. To investigate the potential therapeutic application of As₂O₃ for leukemia, we analyzed the effects of As₂O₃ on the WEHI-3 cells-induced orthotopic leukemia animal model in vivo in this study. We established the WEHI-3 cells leukemia mice through the injection of murine WEHI-3 cells into BALB/c mice, and they were then treated with As₂O₃ (0.9 and 4.5 mg kg⁻¹ ; p.o.) and/or combined with all-trans-retinoic acid (ATRA), (30 mg kg⁻¹ ; i.p.). The results indicated that (1) As₂O₃ alone or As₂O₃ combined with ATRA promoted the total survival rate of leukemia mice and these effects are dose-dependent; (2) As₂O₃ did not affect the body weight but decreased the spleen weight; however, it did not affect liver weight; (3) As₂O₃ alone or As₂O₃ combined with ATRA increased the levels of CD3 and CD19, indicating that the differentiation of T and B cells were promoted; and (4) As₂O₃ alone or As₂O₃ combined with ATRA did not change the levels of Mac-3 and CD11b markers, indicating that the differentiation of the precursor of macrophage were not inhibited. Based on these observations, As₂O₃ alone or As₂O₃ combined with ATRA have efficacious antileukemia activity in WEHI-3 cells leukemia in vivo., (Copyright © 2010 Wiley Periodicals, Inc.)

Most of the chemotherapy treatments for bladder cancer aim to kill the cancer cells, but a high recurrence rate after medical treatments is still occurred. Bufalin from the skin and parotid venom glands of toad has been shown to induce apoptotic cell death in many types of cancer cell lines. However, there is no report addressing that bufalin induced cell death in human bladder cancer cells. The purpose of this study was investigated the mechanisms of bufalin-induced apoptosis in a human bladder cancer cell line (T24). We demonstrated the effects of bufalin on the cell growth and apoptosis in T24 cells by using DAPI/TUNEL double staining, a PI exclusion and flow cytometric analysis. The effects of bufalin on the production of reactive oxygen species (ROS), the level of mitochondrial membrane potential (ΔΨ(m)), and DNA content including sub-G1 (apoptosis) in T24 cells were also determined by flow cytometry. Western blot analysis was used to examine the expression of G(0)/G(1) phase-regulated and apoptosis-associated protein levels in bufalin-treated T24 cells. The results indicated that bufalin significantly decreased the percentage of viability, induced the G(0)/G(1) phase arrest and triggered apoptosis in T24 cells. The down-regulation of the protein levels for cyclin D, CDK4, cyclin E, CDK2, phospho-Rb, phospho-AKT and Bcl-2 with the simultaneous up-regulation of the cytochrome c, Apaf-1, AIF, caspase-3, -7 and -9 and Bax protein expressions and caspase activities were observed in T24 cells after bufalin treatment. Based on our results, bufalin induces apoptotic cell death in T24 cells through suppressing AKT activity and anti-apoptotic Bcl-2 protein as well as inducing pro-apoptotic Bax protein. The levels of caspase-3, -7 and -9 are also mediated apoptosis in bufalin-treated T24 cells. Therefore, bufalin might be used as a therapeutic agent for the treatment of human bladder cancer in the future., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Emilia sonchifolia (L.) DC (Compositae), an herbaceous plant found in Taiwan and India, is used as folk medicine. The clinical applications include inflammation, rheumatism, cough, cuts fever, dysentery, analgesic, and antibacteria. The activities of Emilia sonchifolia extract (ESE) on colorectal cancer cell death have not been fully investigated. The purpose of this study explored the induction of apoptosis and its molecular mechanisms in ESE-treated HCT 116 human colorectal cancer cells in vitro. The methanolic ESE was characterized, and γ-humulene was formed as the major constituent (63.86%). ESE induced cell growth inhibition in a concentration- and time-dependent response by MTT assay. Apoptotic cells (DNA fragmentation, an apoptotic catachrestic) were found after ESE treatment by TUNEL assay and DNA gel electrophoresis. Alternatively, ESE stimulated the activities of caspase-3, -8, and -9 and their specific caspase inhibitors protected against ESE-induced cytotoxicity. ESE promoted the mitochondria-dependent and death-receptor-associated protein levels. Also, ESE increased ROS production and upregulated the levels of ATM, p53, and Fas in HCT 116 cells. Strikingly, p53 siRNA reversed ESE-reduced viability involved in p53-mediated ATM/Fas signaling in HCT 116 cells. In summary, our result is the first report suggesting that ESE may be potentially efficacious in the treatment of colorectal cancer.

Cucurbitacin E, a tetracyclic triterpenes compound extracted from cucurbitaceous plants, has been shown to exhibit anticancer and anti-inflammatory activities. The purpose of this study was to elucidate whether cucurbitacin E promotes cell cycle arrest and induces apoptosis in T24 cells and further to explore the underlying molecular mechanisms. The effects of cucurbitacin E on T24 cell's growth and accompanied morphological changes were examined by MTT assay and a phase-contrast microscope. DNA content, mitochondrial membrane potential (ΔΨ(m)) and annexin V/PI staining were determined by flow cytometry. The protein levels were measured by Western blotting. Our results demonstrated that cucurbitacin E-induced G(2)/M arrest was associated with a marked increase in the levels of p53, p21 and a decrease in phospho-signal transducer and activator of transcription 3 (STAT3), cyclin-dependent kinase 1 (CDK1) and cyclin B. Cucurbitacin E-triggered apoptosis was accompanied with up-regulation of Fas/CD95, truncated BID (t-BID) and a loss of ΔΨ(m), resulting in the releases of cytochrome c, apoptotic protease activating factor 1 (Apaf-1) and apoptosis-inducing factor (AIF), and sequential activation of caspase-8, caspase-9, and caspase-3. Our findings provided the first evidence that STAT3/p53/p21 signaling, Fas/CD95 and mitochondria-dependent pathways play critical roles in cucurbitacin E-induced G(2)/M phase arrest and apoptosis of T24 cells.

Background: Zoledronic acid is widely used as adjuvant chemotherapy for the treatment of breast cancer. However, previous trials reported inconsistent findings regarding their clinical efficacy and safety. We carried out a comprehensive systematic review and meta-analysis to evaluate the effects of zoledronic acid on disease-free survival (DFS), overall survival (OS), and drug-related toxicities., Methodology and Principal Findings: We systematically searched Medline, EmBase, the Cochrane Central Register of Controlled Trials, reference lists of articles and proceedings of major meetings for relevant literatures with a time limit of Dec. 1, 2011. Randomized controlled trials evaluated the effects of zoledronic acid on OS, DFS, and RFS compared with control were eligible for inclusion in our research. Of 175 identified studies, we collected data from 7 randomized controlled trials of zoledronic acid that had OS, DFS, and RFS reported as one of the endpoint. Overall, we noted that patients receiving zoledronic acid therapy had significant longer OS than the group with non-zoledronic acid therapy (HR, 0.85, 95%CI, 0.73 to 1.00, P = 0.047). Furthermore, zoledronic acid therapy also had a clear effect on frature events (RR, 0.66, 95%CI, 0.52 to 0.84, P<0.001). Subgroup analysis indicated that zoledronic acid therapy showed a great beneficial effect on disease recurrence in patients with early-stage breast cancer, however, it also significantly increased the harm of disease recurrence in patients with advanced breast cancer. Bone pain, neutropenic fever, pyrexia, rash were more frequent in the zoledronic acid therapy group., Conclusion/significance: Treatment with zoledronic acid had a clear effect on fracture events, and it might contribute an important role for overall survival.

The present study was to explore the biological responses of the newly compound, MJ-29 in murine myelomonocytic leukemia WEHI-3 cells in vitro and in vivo fates. We focused on the in vitro effects of MJ-29 on ER stress and mitochondria-dependent apoptotic death in WEHI-3 cells, and to hypothesize that MJ-29 might fully impair the orthotopic leukemic mice. Our results indicated that a concentration-dependent decrease of cell viability was shown in MJ-29-treated cells. DNA content was examined utilizing flow cytometry, whereas apoptotic populations were determined using annexin V/PI, DAPI staining and TUNEL assay. Increasing vital factors of mitochondrial dysfunction by MJ-29 were further investigated. Thus, MJ-29-provaked apoptosis of WEHI-3 cells is mediated through the intrinsic pathway. Importantly, intracellular Ca(2+) release and ER stress-associated signaling also contributed to MJ-29-triggered cell apoptosis. We found that MJ-29 stimulated the protein levels of calpain 1, CHOP and p-eIF2α pathways in WEHI-3 cells. In in vivo experiments, intraperitoneal administration of MJ-29 significantly improved the total survival rate, enhanced body weight and attenuated enlarged spleen and liver tissues in leukemic mice. The infiltration of immature myeloblastic cells into splenic red pulp was reduced in MJ-29-treated leukemic mice. Moreover, MJ-29 increased the differentiations of T and B cells but decreased that of macrophages and monocytes. Additionally, MJ-29-stimulated immune responses might be involved in anti-leukemic activity in vivo. Based on these observations, MJ-29 suppresses WEHI-3 cells in vitro and in vivo, and it is proposed that this potent and selective agent could be a new chemotherapeutic candidate for anti-leukemia in the future.

Objective: To evaluate the feasibility of the tissue engineered submandibular gland constructed in vivo based on submandibular gland cells and silk fibroin-chitosan (SFCS)., Methods: Submandibular gland cells were obtained and purified. The second generation cells labeled by 5'-BrdU were seeded on SFCS(5 mm × 5 mm × 5 mm). Submandibular gland cells seeded on SFCS was implanted beneath the skin on the back. At 3, 7, 14 d post-implantation, implant sites were examined local and systemic responses. After paraffin embedding, serial sections 6 mm thick were cut and stained with either hematoxylin and eosin or brdu tissue stain for immunohistochemical studies and examined the responses of tissue. Scanning electron microscope was used to observe the growth behavior submandibular gland cells on SFCS scaffolds., Results: General observation: at the 3, 7, 14 d after in vivo implantation, capsule formed in the surface of insert. Histological observation: in experimental group, submandibular gland cells proliferate on the SFCS scaffold fused to form unit 14 d after implantation. Brdu immunohistochemical observation: the results of labelled cells were positive by immunohistochemical method at each time point. Cytokeratin-8 (CK-8) immunohistochemical observation: the results of labelled cells were positive by immunohistochemical method at each time point. With time, the positive cells gradually increased. Scanning electron microscope: the shape of the SFCS scaffold was mesh. At earlier, submandibular gland cells presence in disorder at attach to the SFCS. At the 14 d submandibular gland cells proliferate on the SFCS scaffold and form functional unit., Conclusions: Such constructed tissue engineered submandibular gland based on submandibular gland cells and SFCS is promising.

Human osteogenic sarcoma is the most common primary bone tumor. Despite of the success of frontline therapy, about 40% of patients have disease progression and further therapy is palliative and toxic. In this study, we developed a novel quinazoline HMJ-30 to investigate the cell growth inhibition and apoptotic responses in U-2 OS human osteogenic sarcoma cells. Our results demonstrated that HMJ-30 significantly reduced cell viabilities of U-2 OS, HOS, and 143B cells in a dose-dependent manner, but it exhibited low cytotoxicity in normal hFOB cells. HMJ-30 induced DNA damage and apoptosis in U-2 OS cells as revealed by morphologic changes, comet assay and DAPI staining. Immuno-staining, colorimetric assays, and Western blotting analyses indicated that activities of caspase-8, caspase-9, and caspase-3 and the levels of Bcl-2 family-related proteins (Bcl-2, Mcl-1, Bax, BAD, and t-Bid) were altered in HMJ-30-treated U-2 OS cells. Pretreatment of cells with caspase-8, -9, and -3 specific inhibitors significantly reduced the cell growth inhibition. HMJ-30-induced apoptosis was mediated through both death-receptor and mitochondria-dependent apoptotic pathways in U-2 OS cells. HMJ-30 induced early phosphorylation of p53(Ser18) was through the activation of ataxia telangiectasia mutated (ATM) in U-2 OS cells. The cell growth inhibition by HMJ-30 was substantially attenuated either by the pre-incubation of U-2 OS cells with N-acetylcysteine (NAC, an antioxidant) and caffeine (an ATM kinase inhibitor) or by p53 knockdown via RNAi. In conclusion, ROS dependent-ATM/p53 signaling pathway is involved in HMJ-30-induced apoptosis in U-2 OS cells., (Copyright © 2011 Orthopaedic Research Society.)

Our earlier studies showed that DATS induced apoptosis in human colon cancer HT29 and colo 205 cell lines in vitro. However, there is no report to show that DATS induced apoptosis in vitro and inhibited CT26 cancer cells in vivo on a murine allograft animal model. In vitro studies, the results indicated that DATS induced morphological changes and induction of apoptosis in CT26 cells. In vivo studies, CT26 cancer cells were implanted into BALB/c mice and groups of mice were treated with vehicle, DATS (10 and 50 mg/kg of body weight). DATS were injected once per four days intraperitoneally (i.p.), with treatment starting 4 weeks prior to cells inoculation. Treatment with vehicle or with 10 and 50 mg/kg of DATS resulted in a reduction in tumor volume and weight. Tumor volume and total hemoglobin in allograft mice treated with 50 mg/kg DATS were significantly smaller than that in the control group. These findings indicated that DATS inhibits tumor growth in an allograft animal model. Thus, DATS may represent a colon cancer preventive agent and can be used in the future., (Copyright © 2011 Elsevier GmbH. All rights reserved.)

Cantharidin (CTD) is a traditional Chinese medicine and an effective component isolated from blister beetle, and it has been demonstrated to have anticancer, antibiotic, antivirus activities and immune-regulated functions. It has been reported that CTD induces cell cycle arrest and apoptosis in many cancer cell types. However, there are no reports showing that CTD would induce cell cycle arrest and apoptosis in human colorectal cancer colo 205 cells. In this study, we studied colo 205 cells which were treated with CTD and demonstrated its molecular mechanisms in apoptosis. CTD induced growth inhibition, G2/M phase arrest and apoptosis in colo 205 cells. The IC50 is 20.53 µM in CTD-treated colo 205 cells. DAPI/TUNEL double staining and Annexin V assays were used to confirm the apoptotic cell death in colo 205 cells after CTD exposure. CTD caused G2/M arrest, down-regulated CDK1 activity, decreased Cyclin A, Cyclin B, CDK1 and increased CHK1 and p21 protein levels. Colorimetric assays also indicated that CTD triggered activities of casapse-8, -9 and -3 in colo 205 cells. Moreover, CTD increased ROS production and decreased the level of mitochondrial membrane potential (ΔΨm) in colo 205 cells. Consequently, CTD-induced growth inhibition was significantly attenuated by N-acetylcysteine (NAC, a scavenger). CTD stimulated the protein levels of Fas/CD95, the caspase-3 active form, cytochrome c and Bax, but suppressed the protein levels of pro-caspase-8, pro-caspase-9 and Bcl-2, determined by Western blot analysis. Based on our observations, we suggest that CTD is able to induce G2/M phase arrest and apoptosis in colo 205 cells through inhibition of CDK1 activity and caspase-dependent signaling pathways.

Triptolide, the main active component obtained from Tripterygium wilfordii Hook. f, has been reported to present potent immunosuppressive and anti-inflammatory biological activities. It has been previously shown that due to the cytotoxicity of triptolide it has a limited use in the clinic. Although numerous studies have shown that triptolide induced apoptosis in many human cancer cells there is no report to show triptolide-induced apoptosis in human adrenal cancer cells. We treated the human adrenal cancer NCI-H295 cells with triptolide in vitro and investigated its cytotoxic effects. The cytotoxicity was examined and quantitated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay and the viability of inhibition and apoptosis was determined by flow cytometric assay, using propidium iodide (PI) staining for apoptosis. Flow cytometric assay also used for the determination of reactive oxygen species (ROS) production and the levels of mitochondrial membrane potential (ΔΨm), and the caspase-3 and -9 activation in NCI-H295 cells. Western blotting was used for examining the changes of apoptotic associated proteins. The results indicated that triptolide induced cytotoxicity (decreased the percentage of viable cells) and induced sub-G1 phase (apoptosis) occurring in NCI-H295 cells and those effects are dose-dependent. Results also showed that triptolide promoted the production of ROS and decreased the levels of ΔΨm in examined NCI-H295 cells. The results showed that triptolide promoted the levels of cytochrome c, Apaf-1, AIF, Endo G, caspase-9 and -3 which were analyzed by Western blotting. These results suggest that triptolide is able to induce apoptosis on NCI-H295 cells through the mitochondrial-dependent signal pathway.

Emodin is one of major compounds in rhubarb (Rheum palmatum L.), a plant used as herbal medicine in Chinese population. Although many reports have shown that emodin exhibits anticancer activity in many tumor cell types, there is no available information addressing emodin-affected apoptotic responses in the murine leukemia cell line (WEHI-3) and modulation of the immune response in leukemia mice. We investigated that emodin induced cytotoxic effects in vitro and affected WEHI-3 cells in vivo. This study showed that emodin decreased viability and induced DNA fragmentation in WEHI-3 cells. Cells after exposure to emodin for 24 h have shown chromatin condensation and DNA damage. Emodin stimulated the productions of ROS and Ca(2+) and reduced the level of ΔΨ(m) by flow cytometry. Our results from Western blotting suggest that emodin triggered apoptosis of WEHI-3 cells through the endoplasmic reticulum (ER) stress, caspase cascade-dependent and -independent mitochondrial pathways. In in vivo study, emodin enhanced the levels of B cells and monocytes, and it also reduced the weights of liver and spleen compared with leukemia mice. Emodin promoted phagocytic activity by monocytes and macrophages in comparison to the leukemia mice group. In conclusions, emodin induced apoptotic death in murine leukemia WEHI-3 cells and enhanced phagocytosis in the leukemia animal model.

Objective: To observe the effects of traditional Chinese medicine mask combined with skin care in the treatment of facial pigmentation after burns., Methods: Forty-one patients with facial pigmentation after burns hospitalized from January 2009 to June 2010 were enrolled and divided into treatment group (n = 26, odd number) and control group (n = 15, even number) according to the visiting order. After cleaning, massaging, and steam spraying to faces, patients in treatment and control groups were respectively treated with traditional Chinese medicine mask developed by physicians in our hospital and common commercial beauty mask. Masks were removed when they became completely dry. The treatment was repeated every other day. Ten times made up a course of treatment. Three consecutive courses were carried out on each patient. Before treatment and at the end of each course, the skin color differences were measured with Vancouver Pigmented Scar Scale; image gray scale value of pigmented skin was measured with image analysis technique. Satisfaction degree acknowledged by both patients and physicians were recorded at the end of each course; adverse effects were recorded; and the overall efficacy between two groups was compared. Data were processed with chi-square test or t test., Results: Skin color differences, image gray scale value of patients in treatment group were close to those in control group before treatment (with t value respectively 0.800 and 0.694, P values all above 0.05). Skin color differences, image gray scale value, and satisfaction degree acknowledged by both patients and physicians in treatment group were better than those in control group at the end of each course. At the end of the third course of treatment, the skin color differences in treatment and control groups scored 0.5 ± 0.4 and 1.1 ± 0.6, respectively, with image gray scale value of 55 ± 5 and 66 ± 6, respectively, which were statistically different from each other (with t value respectively 3.389 and 5.102, P values all below 0.01). The overall efficacy of the treatment group was 92.3%, which was much better than that of the control group (53.3%, χ(2) = 6.31, P < 0.05). No allergy caused by the traditional Chinese medicine mask was observed during the treatment., Conclusions: The traditional Chinese medicine mask combined with skin care is effective for the treatment of facial pigmentation after burns.

Kaempferol is a natural flavonoid. Previous studies have reported that kaempferol has anti-proliferation activities and induces apoptosis in many cancer cell lines. However, there are no reports on human osteosarcoma. In this study, we investigate the anti-cancer effects and molecular mechanisms of kaempferol in human osteosarcoma cells. Our results demonstrate that kaempferol significantly reduces cell viabilities of U-2 OS, HOB and 143B cells, especially U-2 OS cells in a dose-dependent manner, but exerts low cytotoxicity on human fetal osteoblast progenitor hFOB cells. Comet assay, DAPI staining and DNA gel electrophoresis confirm the effects of DNA damage and apoptosis in U-2 OS cells. Flow cytometry detects the increase of cytoplasmic Ca(2+) levels and the decrease of mitochondria membrane potential. Western blotting and fluorogenic enzymatic assay show that kaempferol treatment influences the time-dependent expression of proteins involved in the endoplasmic reticulum stress pathway and mitochondrial signaling pathway. In addition, pretreating cells with caspase inhibitors, BAPTA or calpeptin before exposure to kaempferol increases cell viabilities. The anti-cancer effects of kaempferol in vivo are evaluated in BALB/c(nu/nu) mice inoculated with U-2 OS cells, and the results indicate inhibition of tumor growth. In conclusion, kaempferol inhibits human osteosarcoma cells in vivo and in vitro.

The Xiaogushan cave site is one of the most important prehistoric sites in North China. The stone and bone artifacts found in the cave are similar to European contemporaneous artifacts. Cave deposits consist of five layers that have been dated from 46,353 ± 1179 to 4229 ± 135 cal. yr BP, using radiocarbon dating techniques on charcoal and bone samples collected from Layers 2-5. In this paper, optically stimulated luminescence (OSL) techniques were applied to date six samples taken from Layers 1-3. The luminescence properties of the fine-grained and coarse-grained quartz extracts indicate that the materials are suitable for OSL dating using a single-aliquot regeneration-dose (SAR) protocol. The OSL ages obtained are broadly consistent with the stratigraphy and the associated calibrated radiocarbon ages. The dating results show that the cave was first occupied by humans about 70 ka. The human occupation of the cave may be related to climate change. An occupation hiatus is inferred to between ∼ 17 to ∼ 10 ka. The stone and bone artifacts found in Layers 2 and 3 may indicate the Middle to Upper Paleolithic transitions in the region., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

CHM-1 [2-(2-fluorophenyl)-6,7-methylenedioxyquinolin-4-one] is a quinolone derivative that has been reported to induce apoptosis and inhibit invasion of cancer cells. However, there is no available information to address the effects of CHM-1 on leukemia cells in vivo. Therefore, the present study examined the effects of CHM-1 using a mouse model of leukemia. We established leukemia in mice by injecting WEHI-3 cells into BALB/c mice. Mice were then treated with or without CHM-1 (5 and 10 mg/kg). CHM-1 promoted the total survival rate of leukemic mice and these effects were dose-dependent. CHM-1 increased body weight and decreased spleen weight, but did not affect liver weight. The levels of cell markers Mac-3 and CD11b were reduced by CHM-1, indicating that the differentiation of macrophage precursor cells was inhibited. Levels of CD3 and CD19 were induced by CHM-1, suggesting that the differentiation of precursors of T and B cells was promoted in PBMC. Results of the present study indicate that CHM-1 has an inhibitory effect on leukemia induced in mice in vivo and warrants further study as to the mechanisms and effects in other types of cancer.

Pipoxolan HCl (5,5-diphenyl-2-(2'-piperidino-ethyl)-1,3-dioxolane-4-one hydrochloride) is a compound containing a dioxolan moiety that was reported to induce apoptosis in cancer cells. In this study, we investigated the anti-leukemia effects of pipoxolan on U937 leukemia cells both in vivo and in vitro. Cell viability, reactive oxygen species (ROS) production, mitochondrial membrane potential, apoptosis and caspases-9 and -3 activity were examined following treatment of U937 leukemia cells with 10 µM pipoxolan by flow cytometry and caspase-activity assay. The apoptosis-associated Bcl-2 family proteins, Bax, Bcl-2 and Bcl-xL, were examined by Western blotting. We found that pipoxolan inhibited U937 cell proliferation in a dose- and time-dependent manner. Morphological assessment and cell cycle analysis indicated that pipoxolan induced the apoptosis of the U937 cells. Pipoxolan (10 µM) increased ROS production and decreased mitochondrial membrane potential 1 h after pipoxolan treatment. Pre-treatment of pipoxolan-treated cells with N-acetyl-L-cysteine (a ROS chelator) inhibited the increase in ROS production. After treatment with 10 µM pipoxolan for 24 h, there was an increase in pro-apoptotic Bax and a decrease in anti-apoptotic Bcl-2 and Bcl-xL proteins. In vivo, we found that pipoxolan significantly suppressed tumor growth in BALB/cnu-/nu- mice inoculated with U937 cells. Taken together, the data from our studies indicate that pipoxolan possesses potent anti-leukemia activity and is a potential novel alternative cancer therapeutic agent for human leukemia.

We investigated the signaling pathways associated with microtubule interaction and apoptosis in U937 cells in vitro and in the U937 xenograft model in vivo by using 6-pyrrolidinyl-2-(2-hydroxyphenyl)-4-quinazolinone (MJ-29). MJ-29 induced growth inhibition and cell death of leukemia cell lines (U937, HL-60, K562, and KG-1) in a dose- and time-dependent manner but did not obviously impair the viability of normal cells (peripheral blood mononuclear cells and human umbilical vein endothelial cells). MJ-29 interacted with alpha- and beta-tubulin, inhibited tubulin polymerization both in vitro and in vivo, and disrupted microtubule organization. MJ-29 caused mitotic arrest by activating cyclin-dependent kinase 1 (CDK1)/cyclin B complex activity. MJ-29-induced growth inhibition and activation of CDK1 activity were significantly attenuated by roscovitine (CDK inhibitor) and CDK1 small interfering RNA (siRNA). Furthermore, MJ-29-induced Bcl-2 phosphorylation was also significantly attenuated by CDK1 siRNA. MJ-29 caused an increase in the protein levels of cytosolic cytochrome c, apoptotic protease-activating factor-1, procaspase-9, and apoptosis-inducing factor. MJ-29-promoted activation of caspase-9 and caspase-3 during apoptosis was significantly attenuated by caspase-9 and caspase-3 inhibitors. It is noteworthy that in BALB/c(nu/nu) mice bearing U937 xenograft tumors MJ-29 inhibited tumor growth in vivo. The terminal deoxynucleotidyl transferase-mediated d-UTP nick end-labeling-positive apoptotic cells of tumor sections significantly increased in MJ-29-treated mice compared with the control group. In conclusion, our results suggest that MJ-29 induces mitotic arrest and apoptosis in U937 cells via CDK1-mediated Bcl-2 phosphorylation and inhibits the in vivo tumor growth of U937 xenograft mice.

It is generally believed that liposomes modified with polyethylene glycol (PEG) have no or lower immunogenicity. However, based on many recent literatures, when the PEGylated liposomes were repeatedly applied to the same animal, the immune responses occurred. The first injection of PEGylated liposomes resulted in a reduction in the circulation time and an increase in hepatic and splenic accumulation of the second dose of PEGylated liposomes in a time-interval, which was called "accelerated blood clearance (ABC)" phenomenon. Such immunogenicity of PEGylated liposomes presents a barrier in the research of liposomal formulations and their use in the clinics. This review focused on the definition, the method of verification, the development of the reason for ABC phenomenon, influencing factors of ABC phenomenon, and discussed if other PEGylated nanocarriers also induce ABC phenomenon.

Aims: Cytology fails to detect neoplastic cells in 40-50% of cases of malignant pleural effusion, a condition that frequently accompanies lung adenocarcinoma. Published reports of diagnostic sensitivity of various tumour markers are inconsistent, and optimal cut-off points have not been determined. This study aimed to evaluate the ability of three markers to discriminate lung adenocarcinoma-associated malignant pleural effusion (LAC-MPE) from benign effusion., Methods: Pleural effusion samples were collected from 41 patients with LAC-MPE, and from 93 with various benign conditions. The diagnostic sensitivity and specificity for Her-2/neu, Cyfra 21-1, and carcinoembryonic antigen (CEA) were evaluated. Cut-off points for these markers are optimally set at 3.6 microg/L, 60 microg/L, and 6.0 microg/L, respectively., Results: Her-2/neu, Cyfra 21-1, and CEA vary in their diagnostic accuracy to differentiate LAC-MPE from benign pleural effusion: 79.85%, 88.81%, and 94.03%, respectively. CEA combined with Cyfra 21-1 increases diagnostic sensitivity to 97.6%, with a specificity of 91.4%., Conclusions: With appropriate cut-off points, CEA currently provides the best diagnostic accuracy. Combining CEA with Cyfra 21-1 increases diagnostic sensitivity to nearly 100%. The results of the present study may help clinicians decide whether to obtain a cytological/histological specimen by invasive means to investigate a possible diagnosis of malignancy.

The 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) inhibitors from hawthorn fruit ( Crataegus pinnatifida Bge.) were isolated and evaluated for their antihyperlipidemic effect induced by high-fat diet in mice. After being further purified with silica and polyamide column chromatography from the fractions (fractions A, F, H, and G) with a high inhibitory rate (IR) to HMGR, 24 chromatographic fractions were obtained, including 8 active fractions with a high IR to HMGR. However, the total inhibitory activity of 24 fractions was decreased by about 70%. From eight active fractions, four compounds were obtained by recrystallization and identified as quercetin (a), hyperoside (b), rutin (c), and chlorogenic acid (d), the contents of which in hawthorn EtOH extract were 0.16, 0.32, 1.45, and 0.95%, respectively. The IR values of compounds a-d to HMGR were 6.28, 9.64, 23.53, and 10.56% at the corresponding concentrations of 0.16, 0.32, 1.45, and 0.95 mg/mL, respectively. It was discovered that the IR of a mixture (2.85 mg/mL) matching the original percentage of compounds a-d in hawthorn EtOH extract was up to 79.5%, much higher than that of the single compound and the total IR of these four compounds (50.01%). The in vivo results also revealed that the mixture had a more significant lipid-lowering efficacy than the monomers. Structure-activity relationship revealed the inhibitory activity and lowering-lipid ability of compounds a-c decreased with increasing glycoside numbers. It was concluded that there were synergetic effects on inhibiting HMGR and lowering lipid among compounds a-d, and the weak hydrophilic ability benefits the inhibition to HMGR and lowering-lipid efficacy.

Background: High seroprevalence of human herpesvirus type 8 (HHV-8) in patients with cirrhosis has been reported to be associated with thrombocytopenia. Severe cirrhosis is always complicated with ascites. HHV-8 DNA levels in effusion from patients with primary effusion lymphoma has been reported to be significantly greater than in blood. The status of HHV-8 antibody and DNA in cirrhotic ascites is unclear., Aims: To assess the status of HHV-8 antibody and DNA in cirrhotic ascites compared to that in cirrhotic plasma., Methods: Plasma and ascites samples were collected from 85 patients with cirrhosis. HHV-8 antibody and DNA were detected by immunofluorescence assay and PCR, respectively., Results: Male patients seropositive for HHV-8 antibody were significantly younger than seropositive female patients (p=0.0039). The seropositive rate in patients with cirrhosis was not associated with thrombocytopenia (p=0.6860). Both positive rate and titres of antibody in plasma were much greater than in ascites (p<0.0001). More male or Child-Pugh class C than female or class B seropositive patients were positive for ascites. No hepatitis C virus-related ascites were positive for antibody. Neither plasma nor ascites samples from any subject were positive for HHV-8 DNA., Conclusions: In patients with cirrhosis, the seropositive rate for HHV-8 antibody is independent of thrombocytopenia. The positive rate for HHV-8 antibody in cirrhotic ascites seems to be associated with sex, disease severity and disease aetiology.

ITR-284, a potent anti-leukemia agent of carboxamide derivative, has been shown to inhibit the proliferation of leukemia cells. In this study, the underlying molecular mechanisms in vitro and anti-leukemia activity in vivo of ITR-284 were investigated. ITR-284 reduced the cell viability and induced apoptosis in HL-60 and WEHI-3 leukemia cells. Following exposure of cells to 30 nM of ITR-284, there is a time-dependent decrease in the mitochondrial membrane potential (DeltaPsi(m)) and an increase in the reactive oxygen species (ROS). ITR-284 treatment also caused a time-dependent increase of Fas/CD95, cytosolic cytochrome c, cytosolic active form of caspase-8/-9/-3, cytosolic Apaf-1 and Bax, and the decrease of Bcl-2. However, the ITR-284-induced caspase-8/-9 and -3 activities can be blocked by pan-caspase inhibitor (Z-VAD-FMK). In addition, the anti-leukemia effects of ITR-284 in vivo were further evaluated in BALB/c mice inoculated with WEHI-3 cells. Orally treatment with ITR-284 (2 and 10mg/kg/alternate day for 7 times) increased the survival rate and prevented the loss of body weight in leukemia mice. The enlargement of spleen and infiltration of immature myeloblastic cells into spleen red pulp were significantly reduced in ITR-284-treated mice compared with control mice. Moreover, ITR-284 application can enhance the anti-leukemia effect of all-trans retinoic acid (ATRA). These results revealed that ITR-284 acted against both HL-60 and WEHI-3 in vitrovia both intrinsic and extrinsic apoptotic signaling pathways, and exhibited an anti-leukemic effect in a WEHI-3 orthotopic mice model of leukemia.

Purpose: Chromosome 9 is frequently deleted in high-risk gastrointestinal stromal tumors (GISTs), whereas its specific tumor suppressor genes (TSGs) are less understood. We did an integrative study of MTAP gene at 9p21 to analyze its implication in GISTs., Experimental Design: To search TSGs on chromosome 9, we used ultrahigh-resolution array comparative genomic hybridization to profile DNA copy number alterations of 22 GISTs, with special attention to MTAP gene. MTAP immunoexpression was assessable for 306 independent GISTs on tissue microarrays, with 146 cases analyzed for MTAP homozygous deletion, 181 for mutations of KIT and PDGFRA receptor tyrosine kinase genes, and 7 for MTAP hypermethylation., Results: Array comparative genomic hybridization identified 11 candidate TSGs on 9p and six on 9q. MTAP and/or CDKN2A/CDKN2B at 9p21.3 were deleted in one intermediate-risk (11%) and seven high-risk (70%) GISTs with two cases homozygously codeleted at both loci. MTAP homozygous deletion, present in 25 of 146 cases, was highly associated with larger size and higher mitotic rate, Ki-67 index, and risk level (all P < 0.01) but not with receptor tyrosine kinase genotypes. Whereas MTAP homozygous deletion correlated with MTAP protein loss (P < 0.001), 7 of 30 GISTs without MTAP expression did not show homozygous deletion, including three MTAP-hypermethylated cases. MTAP homozygous deletion was univariately predictive of decreased disease-free survival (P < 0.0001) and remained multivariately independent (P = 0.0369, hazard ratio = 2.166), together with high-risk category (P < 0.0001), Ki-67 index >5% (P = 0.0106), and nongastric location (P = 0.0416)., Conclusions: MTAP homozygous deletion, the predominant mechanism to deplete protein expression, is present in 17% of GISTs. It correlates with important prognosticators and independently predicts worse outcomes, highlighting the role in disease progression.

Aim: To evaluate the efficacy and safety of Traditional Chinese Medicine (TCM) in the treatment of Helicobacter pylori (H. pylori) infection., Methods: We electronically and manually searched electronic databases, references lists and conferences compilations, and included all randomized clinical trials comparing the treatment of H. pylori using TCM with proton pump inhibitor or colloidal bismuth subcitrate-based triple therapy as controls. The Jadad score was used to assess trial quality, H. pylori eradication rate and the incidence of side effects were taken as outcome measurements, and heterogeneity analysis, meta-analysis and funnel plot analysis were conducted., Results: Sixteen trials were included. The Jadad scores of all the trials were not more than 2. Clinical heterogeneity and substantial statistical heterogeneity existed among the trials (P = 0.001, I(2) = 59%) and meta-analysis was not conducted. The average eradication rates following TCM and triple therapy were 72% and 78% and the incidence of side effects were 2% and 29%, respectively. The funnel plot was obviously asymmetric., Conclusion: Available evidence is not convincing enough to show that TCM has the same efficacy as triple therapy in H. pylori treatment. TCM may be safer than triple therapy. TCM should not be recommended as monotherapy in H. pylori infection.

Ezrin, a member of the ezrin-radixin-moesin family, acts as a link between the cell membrane and actin cytoskeleton to integrate cell adhesion-mediated signaling. It implicates tumor progression, metastatic dissemination, and adverse outcomes in several cancer types, including pediatric and adult sarcomas. Although ezrin upregulation was shown by cDNA expression profiling, no study has systematically evaluated the significance of ezrin expression in a large cohort of gastrointestinal stromal tumors (GISTs). Ezrin immunostaining was carried out on tissue microarrays of primary GISTs and assessable in 347 cases, 188 of which were successfully evaluated for mutation variants of KIT and PDGFRA receptor tyrosine kinase (RTK) genes by sequencing with or without screening by denatured high-performance liquid chromatography. These GISTs with known RTK genotypes were dichotomized into two prognostically different groups. The endogenous expression and phosphorylation of ezrin in GIST cell lines were analyzed by western blotting. By immunohistochemistry, ezrin overexpression was present in 66% of GISTs and significantly associated with the non-gastric location (P=0.002) and decreased disease-free survival (P=0.032, univariately). However, it was not related to the National Institute of Health (NIH) risk category, Ki-67 labeling index, RTK genotypes, and other variables. In multivariate analyses, ezrin overexpression remained independently predictive of adverse outcome (P=0.008, risk ratio=2.363), together with Ki-67 labeling index >5% (P<0.001, risk ratio=3.581), high-risk category (P<0.001, risk ratio=2.156), and the non-gastric location (P=0.029, risk ratio=1.899). Despite the variation in the ezrin expression level, phosphorylated ezrin at threonine(567) was only detectable in GIST882 and GIST48 cells, but not in colonic smooth muscle cells. In conclusion, ezrin is frequently overexpressed in GISTs, especially those arising from the non-gastric sites. Given that its impact is independent of the NIH risk category, cell proliferation, and tumor location, ezrin immunoreactivity represents a valuable prognostic adjunct of GISTs, suggesting a causative role in conferring an aggressive phenotype.

The aim of the present study was to further characterize potential clinicopathological predictors for urinary bladder recurrence-free survival (UBRFS) in patients with primary urothelial carcinoma of the upper urinary tract (UUT-UC). The present series included 385 cases of surgically treated primary localized UUT-UC without previous or concurrent urothelial carcinoma of the urinary bladder. Among the 374 patients with follow-up information, clinicopathological features and therapeutic information including whether they received a laparoscopy-assisted nephroureterectomy (LNU) and adjuvant chemotherapy were correlated with UBRFS. After a median follow up of 69 months, 86 patients (23%) developed urinary bladder recurrence. The median time to develop urinary bladder recurrence was 12 months. At the univariate level, an increment in histological grade (P= 0.0321), a prominent papillary configuration (P= 0.0004), LNU (P= 0.0397) and male gender (P= 0.0401) significantly predicted an inferior UBRFS. At the multivariate level, increase of histological grade (P < 0.0001, relative risk (RR) = 3.776), prominent papillary configuration (P < 0.0001, RR = 3.244), and male gender (P= 0.0463, RR = 1.444) independently predicted UBRFS. In conclusion, male patients and those with high-grade and papillary UUT-UC, and who received LNU had higher risks of urinary bladder recurrence. Accordingly, for these patients, more intensive follow up coupled with postoperative intravesical adjuvant therapy to prevent urinary bladder recurrence should also be considered.

(-)-Epigallocatechin-3-gallate (EGCG) is a major constituent of green tea and has been identified as an excellent anticancer agent. Nevertheless, there are no reports to date about the molecular mechanisms and signal pathways of EGCG on the induction of apoptosis in human adrenal NCI-H295 cancer cells. The purpose of this study was to investigate the anticancer effect and molecular mechanisms of EGCG on human adrenal NCI-H295 cancer cells. The results showed that EGCG induced growth inhibition in a dose- and time-dependent manner. Moreover, it exerted low cytotoxicity on Detroit 551 normal human embryonic skin cell. When NCI-H295 cells were treated with 20 microM EGCG, the mitochondrial membrane potential decreased and intracellular free Ca(2+) increased in a time-dependent manner as analysed by flow cytometry. EGCG decreased the protein levels of Bcl-2, Bcl-xl, xIAP, cIAP, Hsp70 and Hsp90, but increased the protein expression of Bad, Bax, Fas/CD95, cytochrome c, Apaf-1, AIF, GADD153, GRP78, and caspase-3, -7,-8 and -9 as observed by Western blotting examination. EGCG promoted caspase-8, -9 and -3 activities in a time-dependent manner. However, pretreatment of cells with inhibitors of caspase-8, -9 and -3 led to a decrease in caspase-8, -9 and -3 activities and an increase in the percentage of viable cells. Based on the above findings, it was confirmed that EGCG may be a drug candidate for the treatment of human adrenal cancer in the future.

Purpose: To determine whether patient self-report of prior laser treatment can be used as a reliable tool for assessing the presence of severe diabetic retinopathy., Design: This was a retrospective study on two groups of diabetic subjects., Methods: One hundred patients with diabetes were recruited from the general eye and retina clinics at the University of Chicago Hospitals. The patients were asked, "Have you ever received laser treatment for your diabetic eye disease (DED)?" A chart review was then conducted noting if the patient had received either focal laser treatment for diabetic macular edema or panretinal photocoagulation for proliferative diabetic retinopathy. Data from the Wisconsin Epidemiological Study of Diabetic Retinopathy (WESDR) were also analyzed. Participant responses to the question "Have you had laser photocoagulation treatment for your eyes?" were analyzed with documentation of photocoagulation scars determined by grading seven-standard field color fundus photographs., Results: In the University of Chicago group, 96 of 100 (96%) of patients were accurate in reporting whether they had received previous laser treatment for DED (sensitivity 95.8%, specificity 96.1%, and positive predictive value 88.5%). In the WESDR analysis, 2,329 of 2,348 (99%) of participants were accurate in reporting whether they had prior laser treatment for DED (sensitivity 96.0%, specificity 99.5%, and positive predictive value 95.6%)., Conclusions: The high sensitivity and specificity of our results validate the use of patient self-report as a useful tool in assessing past laser treatment for severe diabetic retinopathy. Patient self-report may be a useful surrogate to clinical examination or medical record review to determine the presence of severe diabetic retinopathy.

Rhein, an anthraquinone compound, can be found in the rhizome of rhubarb, a traditional Chinese medicine herb showing antitumor activity. In this study, it was observed that rhein induced S-phase arrest through the inhibition of p53, cyclin A and E and it induced apoptosis through the endoplasmic reticulum stress by the production of reactive oxygen species (ROS) and Ca2+ release, mitochondrial dysfunction, and caspase-8, -9 and -3 activation in human tongue cancer cell line (SCC-4). The most efficient induction of apoptosis was observed at 30 microM for 24 h. Mechanistic analysis demonstrated that rhein induced changes in the ratio of Bax/Bcl-2 based on the decrease of Bcl-2 levels, the loss of mitochondrial membrane potential, cytochrome c release from the mitochondria and the activation of caspase-9 and -3. The data demonstrated that rhein induces apoptosis in SCC-4 cells via caspase, ROS and mitochondrial death pathways.

Background/purpose: The proportion of elderly persons among terminal cancer patients is around 60% in Taiwan. Prediction of the last week of survival can be significant in palliative care units for planning patient management, fulfilling patients' wishes at the end of life, and meeting the needs of relatives. We aimed to find out the predictive factors for the last week of survival in elderly terminal cancer patients., Methods: We prospectively observed items based on previous research that we anticipated might influence survival among 459 consecutive elderly patients. Within 24 hours of admission, we collected data including clinical symptoms and signs, demographic information, and biochemical test results. Multivariate logistic regression analysis was performed to identify factors associated with patient mortality within 7 days., Results: Multivariate analysis indicated that the following factors were predictive of death: higher score of Eastern Cooperative Oncology Group (ECOG) performance status (OR, 2.018; p < 0.001), primary liver cancer (OR, 1.968; p = 0.024), male gender (OR, 2.885; p < 0.001), lower extremity muscle power (OR, 0.722; p = 0.0266), lower systolic blood pressure (OR, 0.985; p = 0.011), higher heart rate (OR, 1.017; p = 0.016), higher hemoglobin (OR, 1.216; p = 0.003), and higher blood urea nitrogen (BUN) (OR, 1.028; p < 0.001)., Conclusion: We propose that the probability of survival of less than a week can be predicted by our formula. This formula, which includes factors of demographic information (male gender, and presence of liver cancer), clinical signs (lower systolic blood pressure, higher heart rate, and lower mean extremity muscle power), and biochemical tests (elevated BUN, and higher hemoglobin), may help improve survival prediction in terminal elderly cancer patients.

The Rho family small GTPases are critically involved in the regulation of spine and synaptic properties, but the underlying mechanisms are poorly defined. We took genetic approaches to create and analyze knockout mice deficient in the expression of the protein kinase PAK1 that is directly associated with and activated by the Rho GTPases. We demonstrated that while these knockout mice were normal in both basal and presynaptic function, they were selectively impaired in long-term potentiation (LTP) at hippocampal CA1 synapses. Consistent with the electrophysiological deficits, the PAK1 knockout mice showed changes in the actin cytoskeleton and the actin binding protein cofilin. These results indicate that PAK1 is critical in hippocampal synaptic plasticity via regulating cofilin activity and the actin cytoskeleton.

Purpose: Most gastrointestinal stromal tumors harbor a mutated KIT or PDGFRA receptor tyrosine kinase (RTK). Heat shock protein 90 (Hsp90) is a chaperone mediating the folding and stabilization of many oncoproteins, including KIT. An Hsp90 inhibitor, 17-AAG, can attenuate KIT activation and proliferation of gastrointestinal stromal tumor cell lines. We further evaluated Hsp90 immunoexpression and the difference between alpha and beta isoforms in gastrointestinal stromal tumor specimens., Experimental Design: Hsp90 immunostain was assessable in 306 cases on tissue microarrays of primary gastrointestinal stromal tumors and correlated with various variables and disease-free survival (DFS). RTK mutation variants, confirmed in 142 cases by sequencing with or without precedent denaturing high pressure liquid chromatography screening, were dichotomized into two prognostically different groups. Differential expression of transcript and protein isoforms was measured by real-time reverse transcription-PCR and Western blotting in 16 and 6 cases, respectively., Results: Hsp90 overexpression (55%) significantly correlated with larger size, nongastric location, higher mitotic count and NIH risk level, Ki-67 overexpression (all P < or = 0.001), and unfavorable RTK genotypes (P = 0.020). It strongly portended inferior DFS univariately (P < 0.0001) and remained independent in multivariate analysis (P = 0.031; risk ratio, 2.44), along with high-risk category, Ki-67 overexpression, and old age. For both mRNA and protein, Hsp90beta was more abundant than Hsp90alpha, whereas the latter was significantly higher in high-risk cases., Conclusions: Hsp90 overexpression represents a poor prognosticator that correlates with several adverse parameters, highlighting its role in disease progression and alternative therapy for high-risk, imatinib-resistant gastrointestinal stromal tumors. Hsp90alpha seems more relevant to the intrinsic aggressiveness of gastrointestinal stromal tumors, albeit less abundant than Hsp90beta.

Objective: To understand mental health status of the students and adults in the earthquake disaster areas, as to providing guidance on intervention for post-disaster psychological crisis., Methods: The Self-Reporting Questionnaire (SRQ-20), an appendix of The Ministry of Health of "emergency psychological crisis intervention guiding principles", was used among 1222 victims in Wenchuan earthquake, Sichuan province. This questionnaire survey was conducted in 729 students and 493 adult victims in a randomized method. Of the 734 questionnaires distributed and received in students, 729 were validly responded with efficiency rate of 99.32%; while for adults, 496 questionnaires distributed, 496 received, 493 were valid with efficiency rate of 99.39%., Results: (1) In students victims, 65% responded that they did feel "uneasy, nervous or worried", followed by "easy to be scared", "feel unhappy", "easy fatigue" and "hard to make a decision". For adult victims, 80.5% replied that they felt "uneasy, nervous or worried", followed by "feel unhappy", "easy fatigue", "difficult sleeping", "easy to be scared". (2) The average of the SRQ score of student victims was 6.58 +/- 3.99, and 6.07 +/- 4.02 for males, 7.03 +/- 3.91 for females. The average of the SRQ score of student victims was 7.36 +/- 3.98, male 6.68 +/- 3.82, female 7.99 +/- 4.03. The significant differences (t(women) = 2.985, P < 0.01; t(total) = 3.332, P < 0.01) was observed in women and the total group of students and adults. (3) The SRQ positive screening rate of students group was 46.50%, the adult group was 52.33%, SRQ >or= 7 points and SRQ < 7 sub-group of students and adults of the total score there were significant differences (t(1) = 39.771, P < 0.01; t(2) = 31.961, P < 0.01). SRQ >or= 7 sub-group of students and adult women in between there was a significant difference (t = 23.641, P < 0.01), SRQ < 7 sub-group of students and adults in general and women there were significant differences (t(1) = 3.092, P < 0.01; t(2) = 2.911, P < 0.01). (4) SRQ scores with gender and age had had a positive relation (r(SRQ total-sex) = 0.118, P = 0.000; r(SRQ total-age) = 0.103, P = 0.000)., Conclusion: The emotional suffering symptoms of students victims and adult victims should be "nervous, unhappy, vulnerable on issues such as fatigue", the adult group had a higher symptom rate than the group of students, the mental health was more serious than that of students, as influenced by some factors including age and gender. Students, being a special population group should be paid more attention to focusing a designated behavior intervention, as a planned manner of intervention.