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1.  Ginsenoside Rg3 inhibit hepatocellular carcinoma growth via intrinsic apoptotic pathway 
AIM: To investigate the anti-tumor function of ginsenoside Rg3 on hepatocellular carcinoma (HCC) in vitro and in vivo, and its mechanism.
METHODS: Hep1-6 and HepG2 cells were treated by Rg3 in different concentrations (0, 50, 100 and 200 μg/mL) in vitro. After incubation for 0, 6, 12, 24 and 48 h, cell viability was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. Apoptosis was identified by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling. Caspase-3 activity was measured by chromophore p-nitroanilide and flow cytometry. Bcl-2 family proteins were ascertained by Western-blotting. Mitochondria membrane potential was detected by 5, 5’, 6’ 6’ - tetrachloro-1, 1’, 3, 3’ - tetraethylbenzimidazolylcarbocyanine iodide. Forty liver tumor-bearing C57Bl6 mice were divided randomly into 4 groups for intra-tumor injection of saline, ginsenoside Rg3, cyclophosphamide (CTX) and ginsenoside Rg3 + CTX combination.
RESULTS: The survival time was followed up to 102 d. The mice in the Rg3 + CTX group showed significant increased survival time compared with those in the control group (P < 0.05). Rg3 could inhibit HCC cell proliferation and induce cell apoptosis in vitro in the concentration and time dependent manner. It also induced mitochondria membrane potential to decrease. Caspase-3 activation can be blocked by the inhibitor z-DEVD-FMK. Bax was up-regulated while Bcl-2 and Bcl-XL were down-regulated after Rg3 treatment.
CONCLUSION: Our data suggested that Rg3 alone or combined with CTX inhibited tumor growth in vivo and prolonged mouse survival time by inducing HCC cell apoptosis via intrinsic pathway by expression alterations of Bcl-2 family proteins.
PMCID: PMC3180017  PMID: 21987607
Ginsenoside Rg3; Apoptosis; Hepatocellular Carcinoma; Bcl-2 family proteins; Cyclophosphamide
2.  Application of three-dimensionally area-selective atomic layer deposition for selectively coating the vertical surfaces of standing nanopillars 
Scientific Reports  2014;4:4458.
We describe a strategy for selectively coating the vertical surfaces of standing nanopillars using area-selective atomic layer deposition (ALD). Hydrophobic self-assembled monolayers (SAMs) are utilised to selectively inhibit the coating of oxides on the modified horizontal regions to ensure that only the vertical surfaces of vertical standing nanorods are coated using ALD processes. This method makes it possible to fabricate vertical nanodevices using a simple process of depositing oxide layer on a vertical surface, and can also be applied to the area-selective surface passivation of other standing structures.
PMCID: PMC3964512  PMID: 24662775
3.  2DEGs at Perovskite Interfaces between KTaO3 or KNbO3 and Stannates 
PLoS ONE  2014;9(3):e91423.
We report density functional studies of electron rich interfaces between KTaO3 or KNbO3 and CaSnO3 or ZnSnO3 and in particular the nature of the interfacial electron gasses that can be formed. We find that depending on the details these may occur on either the transition metal or stannate sides of the interface and in the later case can be shifted away from the interface by ferroelectricity. We also present calculations for bulk KNbO3, KTaO3, CaSnO3, BaSnO3 and ZnSnO3, showing the different transport and optical properties that may be expected on the two sides of such interfaces. The results suggest that these interfaces may display a wide range of behaviors depending on conditions, and in particular the interplay with ferroelectricity suggests that electrical control of these properties may be possible.
PMCID: PMC3953397  PMID: 24626191
4.  Discovery of Plant Phenolic Compounds That Act as Type III Secretion System Inhibitors or Inducers of the Fire Blight Pathogen, Erwinia amylovora 
Applied and Environmental Microbiology  2013;79(18):5424-5436.
Erwinia amylovora causes a devastating disease called fire blight in rosaceous plants. The type III secretion system (T3SS) is one of the important virulence factors utilized by E. amylovora in order to successfully infect its hosts. By using a green fluorescent protein (GFP) reporter construct combined with a high-throughput flow cytometry assay, a library of phenolic compounds and their derivatives was studied for their ability to alter the expression of the T3SS. Based on the effectiveness of the compounds on the expression of the T3SS pilus, the T3SS inhibitors 4-methoxy-cinnamic acid (TMCA) and benzoic acid (BA) and one T3SS inducer, trans-2-(4-hydroxyphenyl)-ethenylsulfonate (EHPES), were chosen for further study. Both the T3SS inhibitors (TMCA and BA) and the T3SS inducer (EHPES) were found to alter the expression of T3SS through the HrpS-HrpL pathway. Additionally, TMCA altered T3SS expression through the rsmBEa-RsmAEa system. Finally, we found that TMCA and BA weakened the hypersensitive response (HR) in tobacco by suppressing the T3SS of E. amylovora. In our study, we identified phenolic compounds that specifically targeted the T3SS. The T3SS inhibitor may offer an alternative approach to antimicrobial therapy by targeting virulence factors of bacterial pathogens.
PMCID: PMC3754148  PMID: 23770912
5.  The role of quality of care and attitude towards disability in the relationship between severity of disability and quality of life: findings from a cross-sectional survey among people with physical disability in China 
People with physical disability (PWPD) is the largest subgroup of people with disability (PWD) in China, but few studies have been conducted among this vulnerable population. The objective of this study was to investigate the level of quality of life (QoL), self-perceived quality of care and support (QOCS), severity of disability and personal attitude towards disability among people with physical disability in China, as well as to identify how QoL can be affected by severity of disability through QOCS and personal attitude towards disability among PWPD.
A cross-sectional study was conducted among 1,853 PWPD in Guangzhou, China. Data were collected on participants’ QoL, QOCS, personal attitude towards disability and severity of disability. Structural equation modeling was used to examine the effects of the other variables on QoL.
Even with a mild disability (mean score:1.72), relatively low levels of QoL (mean score: 2.65- 3.22) and QOCS (mean score: 2.95 to 3.28), as well as unfavorable personal attitude towards disability (mean score: 2.75 to 3.36) were identified among PWPD. According to SEM, we found that the influence of severity of physical disability on QoL is not only exerted directly, but is also indirectly through QOCS and their personal attitudes towards disability, with QOCS playing a more important mediating role than PWPD’s attitudes towards their own disability.
Unfavorable health status was identified among PWPD in China. Focusing on improvement of assistance and care services has the potential to substantially improve PWPD’s QoL. Further research should focus on understanding the needs and their current state of health care of PWPD in China thus being able to develop better interventions for them.
PMCID: PMC3941947  PMID: 24559096
Quality of life; Quality of care and support; Attitude towards disability; Severity of disability; People with physical disability; Structural equation modeling
6.  New insights into prostate cancer stem cells 
Cell Cycle  2013;12(4):579-586.
Prostate cancer (PCa) remains one of the most prevalent malignancies affecting men in the western world. The etiology for PCa development and molecular mechanisms underlying castration-resistant progression are incompletely understood. Emerging evidence from many tumor systems has shown the existence of distinct subpopulations of stem like-cancer cells termed cancer stem cells (CSCs), which may be involved in tumor initiation, progression, metastasis and therapy resistance. Prostate cancer stem cells (PCSCs) have also been identified using different experimental strategies in distinct model systems. In this brief review, we summarize our current knowledge of normal prostate stem/progenitor cells, highlight recent progress on PCSCs, expound on the potential cell-of-origin for PCa and discuss the involvement of PCSCs in PCa progression and castration resistance. Elucidation of the phenotypic and functional properties and molecular regulation of PCSCs will help us better understand PCa biology and may lead to development of novel therapeutics targeting castration-resistant PCa cells.
PMCID: PMC3594258  PMID: 23370446
prostate cancer; cancer stem cells; prostate cancer stem cells; differentiation; therapy resistance
7.  Efficacy and Safety of Therapies for Acute Ischemic Stroke in China: A Network Meta-Analysis of 13289 Patients from 145 Randomized Controlled Trials 
PLoS ONE  2014;9(2):e88440.
Many of these therapies have been compared against placebos, but have not been directly compared against each other. To evaluate the efficacy and safety of several commonly used drugs for AIS directly or indirectly.
A systematic literature review was performed to identify randomized controlled trials (RCTs) published prior to April 2013 for AIS therapies. The primary outcome measures were the National Institutes of Health Stroke Scale (NIHSS) scores and the clinical effective rate. A fixed-effects meta-analysis and meta-regression are performed; lastly, performed a mixed treatment comparison was performed through the Bayesian methods.
Outcome of Efficacy of therapies for acute ischemic stroke are as followed: All of the therapies mentioned above yielded results a more effective result than placebo, Sodium ozagrel (RR 3.86, 95%CI 3.18–4.61); Sodium ozagrel + edaravone (RR 9.60, 95%CI 7.04–13.06); Edaravone (RR 4.07, 95%CI 3.30–5.01); Edaravone + Kininogenase (RR 15.33, 95%CI 10.03–23.05). The significant difference in efficacy between edaravone monotherapy and Sodium ozagrel + edaravone was evident (RR 0.43, 95%CI 0.08–0.61) and was also significant between efficacy of edaravone + Kininogenase and Sodium ozagrel (RR 4.00, 95%CI 2.47–6.24). The differences between the risk and benefit were not significant when comparing Sodium ozagrel and edaravone or edaravone + Kininogenase and Sodium ozagrel + Edaravone for AIS. Outcome of the defect of neurological function: Placebo served a significant difference in treating the defects of neurological function compared with Sodium ozagrel (WMD = −3.11, 95%CI −4.43 to −1.79), Sodium ozagrel + edaravone (WMD = −6.25, 95%CI −7.96 to −4.54) and Edaravone + Kininogenase (WMD =  −3.47, 95%CI −5.73 to −1.21).
It provides that the efficacy of edaravone monotherapy in treatment was not more effective than Sodium ozagrel + edaravone.The efficacy of edaravone + Kininogenase monotherapy in treatment was more effective than Sodium ozagrel. Edaravone + Kininogenase and Sodium ozagrel + Edaravone appeared the most effective treatments. And Sodium ozagrel, Sodium ozagrel + edaravone, Edaravone + Kininogenase can improve the nerve dysfunction.
PMCID: PMC3923787  PMID: 24551100
8.  Fibroblast Activation Protein (FAP) Is Essential for the Migration of Bone Marrow Mesenchymal Stem Cells through RhoA Activation 
PLoS ONE  2014;9(2):e88772.
The ability of human bone marrow mesenchymal stem cells (BM-MSCs) to migrate and localize specifically to injured tissues is central in developing therapeutic strategies for tissue repair and regeneration. Fibroblast activation protein (FAP) is a cell surface serine protease expressed at sites of tissue remodeling during embryonic development. It is also expressed in BM-MSCs, but not in normal tissues or cells. The function of FAP in BM-MSCs is not known.
Principal Findings
We found that depletion of FAP proteins significantly inhibited the migration of BM-MSCs in a transwell chemotaxis assay. Such impaired migration ability of BM-MSCs could be rescued by re-expressing FAP in these cells. We then demonstrated that depletion of FAP activated intracellular RhoA GTPase. Consistently, inhibition of RhoA activity using a RhoA inhibitor rescued its migration ability. Inhibition of FAP activity with an FAP-specific inhibitor did not affect the activation of RhoA or the migration of BM-MSCs. Furthermore, the inflammatory cytokines interleukin-1beta (IL-1β) and transforming growth factor-beta (TGF-β) upregulated FAP expression, which coincided with better BM-MSC migration.
Our results indicate FAP plays an important role in the migration of BM-MSCs through modulation of RhoA GTPase activity. The peptidase activity of FAP is not essential for such migration. Cytokines IL-1β and TGF-β upregulate the expression level of FAP and thus enhance BM-MSC migration.
PMCID: PMC3923824  PMID: 24551161
9.  M1/M2 macrophages and associated mechanisms in congenital bicuspid aortic valve stenosis 
The aim of this study was to observe macrophage infiltration in congenital bicuspid aortic valve (CBAV) stenosis. M1/M2 macrophage distribution, inflammatory cytokine expression and the role of M1 macrophages during CBAV stenosis were also explored. The experimental and control groups comprised 30 severely stenotic CBAVs and 30 severely stenotic tricuspid aortic valves (TAVs), respectively. Histological and morphological changes were assessed using hematoxylin-eosin (HE) staining and mRNA levels of vascular endothelial growth factor (VEGF) were examined using the quantitative polymerase chain reaction. Nonspecific, M1 and M2 macrophages were monitored using cluster of differentiation (CD)68, inducible nitric oxide synthase (iNOS) and CD163 staining, respectively. Endothelial nitric oxide synthase (eNOS), interleukin (IL)-10, arginase (Arg)-1 and macrophage colony-stimulating factor (M-CSF) were also examined using immunohistochemical staining. Of note, HE staining revealed a higher cell density and neovascularization was more common in CBAVs than TAVs. At the mRNA level, VEGF expression was two-fold higher in CBAVs relative to that in TAVs (P=0.02). Furthermore, CD68 and iNOS were significantly higher in CBAVs compared with TAVs (P=0.029 and 0.021, respectively), while CD163 expression was lower in CBAVs (P=0.033). In addition, eNOS expression was higher and Arg-1, IL-10 and M-CSF expression were lower in CBAVs compared with TAVs (all P<0.0001). The present study suggested that CBAVs are associated with a higher total and M1 macrophage density and a lower M2 macrophage density than TAVs, and that M1 macrophage infiltration may contribute to calcification of CBAVs.
PMCID: PMC3965126  PMID: 24669254
congenital bicuspid aortic valve; tricuspid aortic valve; macrophage; inflammation; neovascularization
10.  Myocardial Infarction-induced N-terminal Fragment of Cardiac Myosin-binding Protein C (cMyBP-C) Impairs Myofilament Function in Human Myocardium* 
The Journal of Biological Chemistry  2014;289(13):8818-8827.
Background: Myocardial infarction (MI) leads to proteolytic cleavage of cMyBP-C (hC0C1f) and decreased contractility.
Results: hC0C1f can incorporate into the human cardiac sarcomere, depressing force generation and increasing tension cost.
Conclusion: Interaction between hC0C1f and both actin and α-tropomyosin causes disruption of intact cMyBP-C function.
Significance: Proteolytic cleavage of cMyBP-C is sufficient to cause contractile dysfunction following MI.
Myocardial infarction (MI) is associated with depressed cardiac contractile function and progression to heart failure. Cardiac myosin-binding protein C, a cardiac-specific myofilament protein, is proteolyzed post-MI in humans, which results in an N-terminal fragment, C0-C1f. The presence of C0-C1f in cultured cardiomyocytes results in decreased Ca2+ transients and cell shortening, abnormalities sufficient for the induction of heart failure in a mouse model. However, the underlying mechanisms remain unclear. Here, we investigate the association between C0-C1f and altered contractility in human cardiac myofilaments in vitro. To accomplish this, we generated recombinant human C0-C1f (hC0C1f) and incorporated it into permeabilized human left ventricular myocardium. Mechanical properties were studied at short (2 μm) and long (2.3 μm) sarcomere length (SL). Our data demonstrate that the presence of hC0C1f in the sarcomere had the greatest effect at short, but not long, SL, decreasing maximal force and myofilament Ca2+ sensitivity. Moreover, hC0C1f led to increased cooperative activation, cross-bridge cycling kinetics, and tension cost, with greater effects at short SL. We further established that the effects of hC0C1f occur through direct interaction with actin and α-tropomyosin. Our data demonstrate that the presence of hC0C1f in the sarcomere is sufficient to induce depressed myofilament function and Ca2+ sensitivity in otherwise healthy human donor myocardium. Decreased cardiac function post-MI may result, in part, from the ability of hC0C1f to bind actin and α-tropomyosin, suggesting that cleaved C0-C1f could act as a poison polypeptide and disrupt the interaction of native cardiac myosin-binding protein C with the thin filament.
PMCID: PMC3979389  PMID: 24509847
Contractile Protein; Heart Failure; Myocardial Infarction; Protein Degradation; Protein-Protein Interactions
11.  Heterogeneous Effect of Two Selectin Gene Polymorphisms on Coronary Artery Disease Risk: A Meta-Analysis 
PLoS ONE  2014;9(2):e88152.
The selectins play important roles in the inflammatory process of coronary artery disease (CAD) and myocardial infarction (MI). Previous studies have shown ambiguous findings regarding a possible association between the selectin genes and CAD. The E-selectin Ser128Arg polymorphism and the P-selectin Thr715Pro polymorphism have been investigated widely but with inconsistent results. We performed a comprehensive meta-analysis to shed light on this issue.
Data were extracted by searches of MEDLINE, Embase, CNKI, Wanfang, Google Scholar, PORTA, GeNii, CiNii, J-STAGE, Nurimedia and Koreanstudies Information Service System [Kiss] up to October 2013, in which 10 studies on the Ser128Arg polymorphism with 3369 cases and 2577 controls and 10 studies on the Thr715Pro polymorphism with 5886 cases and 18345 controls. A random-effects model was used to calculate the combined odds ratios. The between-study heterogeneity and publication bias were addressed.
The 128Arg carriers had a significant increased risk of CAD (allele comparison: P = 0.02, OR = 1.33, 95%CI 1.04–1.69, Pheterogeneity = 0.01); The 715Pro conferred a non-significant risk reduction relative to the 715Thr (allele comparison: P = 0.40, OR = 0.94, 95%CI 0.82–1.08, Pheterogeneity = 0.03).Subgroup analyses demonstrated that the 128Arg carriers had a significant increased risk of CAD among Asians (allele comparison: P = 0.001, OR = 2.07, 95%CI 1.33–3.24, Pheterogeneity = 0.77) but not among Caucasians (allele comparison: P = 0.33, OR = 1.13, 95%CI 0.88–1.45, Pheterogeneity = 0.08). Carrier status for the 715Pro was significantly associated with reduced risk of MI (allele comparison: P = 0.04, OR =  0.81, 95%CI 0.67–0.99, Pheterogeneity = 0.14). The asymmetric funnel plot and the Egger's test (P = 0.041) suggested the presence of publication bias for the Ser128Arg polymorphism.
Our results suggested there is an increase in the risk of CAD conferred by the Ser128Arg polymorphism and the thr715Pro polymorphism may be a protective factor of MI.
PMCID: PMC3912165  PMID: 24498435
12.  TNFR2 is critical for the stabilization of the CD4+FoxP3+ regulatory T cell phenotype in the inflammatory environment 
Several lines of evidence indicate the instability of CD4+FoxP3+ regulatory T cells (Tregs). We have therefore investigated means of promoting the stability of Tregs. In this study, we found that the proportion of Tregs in mouse strains deficient in TNFR2 or its ligands was reduced in the thymus and peripheral lymphoid tissues, suggesting a potential role of TNFR2 in promoting the sustained expression of FoxP3. We observed that upon in vitro activation with plate-bound anti-CD3 Ab and soluble anti-CD28 Ab, FoxP3 expression by highly purified mouse Tregs was markedly down-regulated. Importantly, TNF partially abrogated this effect of TCR stimulation and stabilized FoxP3 expression. This effect of TNF was blocked by anti-TNFR2 Ab, but not by anti-TNFR1 Ab. Furthermore, TNF was not able to maintain FoxP3 expression by TNFR2-deficient Tregs. In mouse colitis model induced by transfer of naïve CD4 cells into Rag1−/− mice, the disease could be inhibited by co-transfer of WT Tregs, but not by co-transfer of TNFR2-deficient Tregs. Furthermore, in the lamina propria of the colitis model, the majority of WT Tregs maintained FoxP3 expression. In contrast, increased number of TNFR2-deficient Tregs lost FoxP3 expression. Thus, our data clearly show that TNFR2 is critical for the phenotypic and functional stability of Treg in the inflammatory environment. This effect of TNF should be taken into account when designing future therapy of autoimmunity and GVHD by using TNF inhibitors.
PMCID: PMC3552130  PMID: 23277487
T cell; cytokine receptor; tolerance/suppression/anergy
13.  MicroRNA 181 Suppresses Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) Infection by Targeting PRRSV Receptor CD163 
Journal of Virology  2013;87(15):8808-8812.
We previously showed that microRNA 181 (miR-181) can inhibit PRRSV replication by directly targeting its genomic RNA. Here, we report that miR-181 can downregulate the PRRSV receptor CD163 in blood monocytes and porcine alveolar macrophages (PAMs) through targeting the 3′ untranslated region (UTR) of CD163 mRNA. Downregulation of CD163 leads to the inhibition of PRRSV entry into PAMs and subsequently suppresses PRRSV infection. Our findings indicate that delivery of miR-181 can be used as antiviral therapy against PRRSV infection.
PMCID: PMC3719804  PMID: 23740977
14.  Draft Genome Sequence of Sphingobium sp. Strain C100, a Polycyclic Aromatic Hydrocarbon-Degrading Bacterium from the Deep-Sea Sediment of the Arctic Ocean 
Genome Announcements  2014;2(1):e01210-13.
Sphingobium sp. strain C100 was isolated from a polycyclic aromatic hydrocarbon (PAH)-degrading consortium from the deep-sea sediment of the Arctic Ocean. It can degrade two- to four-ring PAHs at 25°C. Here we present the draft genome sequence of this strain, which is 4,776,810 bp with a G+C content of 63.9%.
PMCID: PMC3907727  PMID: 24482512
15.  Relationship of EMG/SMG features and muscle strength level: an exploratory study on tibialis anterior muscles during plantar-flexion among hemiplegia patients 
Improvement in muscle strength is an important aim for the rehabilitation of hemiplegia patients. Presently, the rehabilitation prescription depends on the evaluation results of muscle strength, which are routinely estimated by experienced physicians and therefore not finely quantitative. Widely-used quantification methods for disability, such as Barthel Index (BI) and motor component of Functional Independent Measure (M-FIM), yet have limitations in their application, since both of them differentiated disability better in lower than higher disability, and they are subjective and recorded in wide scales. In this paper, to explore finely quantitative measures for evaluation of muscle strength level (MSL), we start with the study on quantified electromyography (EMG) and sonomyography (SMG) features of tibialis anterior (TA) muscles among hemiplegia patients.
12 hemiplegia subjects volunteered to perform several sets of plantar-flexion movements in the study, and their EMG signals and SMG signals were recorded on TA independently to avoid interference. EMG data were filtered and then the root-mean-square (RMS) was computed. SMG signals, specifically speaking, the muscle thickness of TA, were manually measured by two experienced operators using ultrasonography. Reproducibility of the SMG assessment on TA between operators was evaluated by non-parametric test (independent sample T test). Possible relationship between muscle thickness changes (TC) of TA and muscle strength level of hemiplegia patients was estimated.
Mean of EMG RMS between subjects is found linearly correlated with MSL (R2 = 0.903). And mean of TA muscle TC amplitudes is also linearly correlated with MSL among dysfunctional legs (R2 = 0.949). Moreover, rectified TC amplitudes (dysfunctional leg/ healthy leg, DLHL) and rectified EMG signals (DLHL) are found in linear correlation with MSL, with R2 = 0.756 and R2 = 0.676 respectively. Meanwhile, the preliminary results demonstrate that patients’ peak values of TC are generally proportional to their personal EMG peak values in 12 dysfunctional legs and 12 healthy legs (R2 = 0.521).
It’s concluded that SMG could be a promising option to quantitatively estimate MSL for hemiplegia patients during rehabilitation besides EMG. However, after this exploratory study, they should be further investigated on a larger number of subjects.
PMCID: PMC3923562  PMID: 24461052
Sonomyography; Tibialis anterior muscle; Surface electromyography; Muscle strength level
16.  Draft Genome Sequence of Marinomonas sp. Strain D104, a Polycyclic Aromatic Hydrocarbon-Degrading Bacterium from the Deep-Sea Sediment of the Arctic Ocean 
Genome Announcements  2014;2(1):e01211-13.
Marinomonas sp. strain D104 was isolated from a polycyclic aromatic hydrocarbon-degrading consortium enriched from deep-sea sediment from the Arctic Ocean. The draft genome sequence of D104 (approximately 3.83 Mbp) contains 62 contigs and 3,576 protein-encoding genes, with a G+C content of 44.8%.
PMCID: PMC3900904  PMID: 24459272
17.  Generation of “Virtual” Control Groups for Single Arm Prostate Cancer Adjuvant Trials 
PLoS ONE  2014;9(1):e85010.
It is difficult to construct a control group for trials of adjuvant therapy (Rx) of prostate cancer after radical prostatectomy (RP) due to ethical issues and patient acceptance. We utilized 8 curve-fitting models to estimate the time to 60%, 65%, … 95% chance of progression free survival (PFS) based on the data derived from Kattan post-RP nomogram. The 8 models were systematically applied to a training set of 153 post-RP cases without adjuvant Rx to develop 8 subsets of cases (reference case sets) whose observed PFS times were most accurately predicted by each model. To prepare a virtual control group for a single-arm adjuvant Rx trial, we first select the optimal model for the trial cases based on the minimum weighted Euclidean distance between the trial case set and the reference case set in terms of clinical features, and then compare the virtual PFS times calculated by the optimum model with the observed PFSs of the trial cases by the logrank test. The method was validated using an independent dataset of 155 post-RP patients without adjuvant Rx. We then applied the method to patients on a Phase II trial of adjuvant chemo-hormonal Rx post RP, which indicated that the adjuvant Rx is highly effective in prolonging PFS after RP in patients at high risk for prostate cancer recurrence. The method can accurately generate control groups for single-arm, post-RP adjuvant Rx trials for prostate cancer, facilitating development of new therapeutic strategies.
PMCID: PMC3897405  PMID: 24465467
18.  Chemo-Immunotherapy with Oxaliplatin and Interleukin-7 Inhibits Colon Cancer Metastasis in Mice 
PLoS ONE  2014;9(1):e85789.
Combination of immunotherapy and chemotherapy has shown promise for cancer. Interleukin-7 (IL-7) can potentially enhance immune responses against tumor, while oxaliplatin (OXP), a platinum-based drug, can promote a favorable immune microenvironment and stimulate anticancer immune responses. We evaluated the anti-tumor activity of IL-7 combining OXP against a murine colon carcinoma in vitro and in vivo and studied the tumor immune microenvironment to investigate whether the combined treatment affects on the local immune cell populations. Utilizing lung and abdomen metastasis models by inoculation of CT26 mice colon cancer cells, we evaluated the anti-tumor efficacy of combining IL-7 and OXP in mice models. Tumor immune microenvironment was evaluated by flow cytometric analysis and immunohistochemical staining. Our study showed that the in vivo administration of IL-7 combined with OXP markedly inhibited the growth of tumors in lung and abdomen metastasis models of colon cancer. IL-7 alone had no effect on tumor growth in mice and IL-7 did not alter cell sensitivity to OXP in culture. The antitumor effect of combining IL-7 and OXP correlated with a marked increase in the number of tumor-infiltrating activated CD8+ T cells and a marked decrease in the number of regulatory T (Treg) cells in spleen. Our data suggest that OXP plus IL-7 treatment inhibits tumor cell growth by immunoregulation rather than direct cytotoxicity. Our findings justify further evaluation of combining IL-7 and chemotherapy as a novel experimental cancer therapy.
PMCID: PMC3897491  PMID: 24465710
19.  Correction: Prognostic Value of Cancer Stem Cell Marker CD133 Expression in Gastric Cancer: A Systematic Review 
PLoS ONE  2014;9(1):10.1371/annotation/80c50c32-16b8-4099-98e4-af72154192b3.
PMCID: PMC3894292
20.  Effective Chemoimmunotherapy with Anti-TGFβ Antibody and Cyclophosphamide in a Mouse Model of Breast Cancer 
PLoS ONE  2014;9(1):e85398.
TGFβ is reportedly responsible for accumulation of CD4+Foxp3+ regulatory T cells (Tregs) in tumor. Thus, we treated mouse 4T1 mammary carcinoma with 1D11, a neutralizing anti-TGFβ (1,2,3) antibody. The treatment delayed tumor growth, but unexpectedly increased the proportion of Tregs in tumor. In vitro, 1D11 enhanced while TGFβ potently inhibited the proliferation of Tregs. To enhance the anti-tumor effects, 1D11 was administered with cyclophosphamide which was reported to eliminate intratumoral Tregs. This combination resulted in long term tumor-free survival of up to 80% of mice, and the tumor-free mice were more resistant to re-challenge with tumor. To examine the phenotype of tumor infiltrating immune cells, 4T1-tumor bearing mice were treated with 1D11 and a lower dose of cyclophosphamide. This treatment markedly inhibited tumor growth, and was accompanied by massive infiltration of IFNγ-producing T cells. Furthermore, this combination markedly decreased the number of splenic CD11b+Gr1+ cells, and increased their expression levels of MHC II and CD80. In a spontaneous 4T1 lung metastasis model with resection of primary tumor, this combination therapy markedly increased the survival of mice, indicating it was effective in reducing lethal metastasis burden. Taken together, our data show that anti-TGFβ antibody and cyclophosphamide represents an effective chemoimmunotherapeutic combination.
PMCID: PMC3887137  PMID: 24416401
21.  Digestive tract reconstruction pattern as a determining factor in postgastrectomy quality of life 
Postgastrectomy quality of life (QoL) is affected by various symptoms, and compared with the preoperative baseline QoL, is typically impaired for the first 6 mo after surgery. Thereafter, improvement to a stable QoL is observed at approximately 12 mo postoperatively. We consider the digestive tract reconstruction pattern to be a determining factor in postgastrectomy QoL among gastric cancer patients, and believe it requires further discussion. Proximal gastrectomy is associated with the worst postoperative QoL among gastrectomy procedures and should be performed cautiously. The trend of better QoL provided by the pouch procedure of total gastrectomy requires further robust support. Whether the use of Billroth-I gastroduodenostomy or Roux-en-Y gastrojejunostomy for distal gastrectomy is optimal remains controversial, but Roux-en-Y gastrojejunostomy is likely to be preferable.
PMCID: PMC3886029  PMID: 24415891
Gastric cancer; Gastrectomy; Quality of life; Reconstruction; Digestive tract
22.  Top-down Targeted Proteomics for Deep Sequencing of Tropomyosin Isoforms 
Journal of proteome research  2012;12(1):187-198.
Tropomyosins (Tm) constitute a family of ubiquitous and highly conserved actin-binding proteins, playing essential roles in a variety of biological processes. Tm isoforms produced by multiple Tm encoding genes and alternatively expressed exons along with post-translational modifications (PTMs) regulate Tm function. Therefore, to gain a better understanding of the functional role of Tm, it is essential to fully characterize Tm isoforms. Herein, we developed a top-down high-resolution mass spectrometry (MS) based targeted proteomics method for comprehensive characterization of Tm isoforms. α–Tm was identified to be the predominant isoform in swine cardiac muscle. We further characterized its sequence and localized the PTMs such as acetylation and phosphorylation as well as amino acid polymorphisms. Interestingly, we discovered a “novel” Tm isoform that does not match with any of the currently available swine Tm sequences. A deep sequencing of this isoform by top-down MS revealed an exact match with mouse β–Tm sequence, suggesting that this “novel” isoform is swine β–Tm which is 100% conserved between swine and mouse. Taken together, we demonstrated that top-down targeted proteomics provides a powerful tool for deep sequencing of Tm isoforms from genetic variations together with complete mapping of the PTM sites.
PMCID: PMC3596867  PMID: 23256820
Tropomyosin; Muscle Contraction; Actin Filament; Mass Spectrometry; Electron Capture Dissociation
23.  The in vitro study of apoptosis in NB4 cell induced by citral 
Cytotechnology  2012;65(1):49-57.
Citral, 3,7-dimethyl-2,6-octadienal, is a key component of the essential oils extracted from several lemon-scented herbal plants. Besides its antifungal activity, the anticancer effect of citral was studied in recent years. In this study, we investigated the effect of citral on the acute promyelocytic leukemia cell line NB4. Citral treatment had an antiproliferative effect in NB4 cells via the induction of apoptosis assessed by morphology, proliferation assay, DNA electrophoresis, Annexin V-FITC/PI staining and caspase-3 activation. And citral induced apoptosis of NB4 cells in a dose- and time-dependent manner. In addition, citral treatment induced decreased mitochondrial membrane potential, indicating that citral induced apoptosis via the mitochondrial pathway. Bax up-regulation and Bcl-2 down-regulation on mRNA level and NF-κB down-regulation on protein level was found in this study, suggesting that Bcl-2, Bax and NF-κB may be involved in the mechanism of the apoptotic effect of citral on NB4 cells. These data suggest that citral has a potential therapeutic effect on leukemia.
PMCID: PMC3536870  PMID: 22573288
Citral; Apoptosis; Acute promyelocytic leukemia; Bax; NB4
24.  Effects of Parabens on Adipocyte Differentiation 
Toxicological Sciences  2012;131(1):56-70.
Parabens are a group of alkyl esters of p-hydroxybenzoic acid that include methylparaben, ethylparaben, propylparaben, butylparaben, and benzylparaben. Paraben esters and their salts are widely used as preservatives in cosmetics, toiletries, food, and pharmaceuticals. Humans are exposed to parabens through the use of such products from dermal contact, ingestion, and inhalation. However, research on the effects of parabens on health is limited, and the effects of parabens on adipogenesis have not been systematically studied. Here, we report that (1) parabens promote adipogenesis (or adipocyte differentiation) in murine 3T3-L1 cells, as revealed by adipocyte morphology, lipid accumulation, and mRNA expression of adipocyte-specific markers; (2) the adipogenic potency of parabens is increased with increasing length of the linear alkyl chain in the following potency ranking order: methyl- < ethyl- < propyl- < butylparaben. The extension of the linear alkyl chain with an aromatic ring in benzylparaben further augments the adipogenic ability, whereas 4-hydroxybenzoic acid, the common metabolite of all parabens, and the structurally related benzoic acid (without the OH group) are inactive in promoting 3T3-L1 adipocyte differentiation; (3) parabens activate glucocorticoid receptor and/or peroxisome proliferator-activated receptor γ in 3T3-L1 preadipocytes; however, no direct binding to, or modulation of, the ligand binding domain of the glucocorticoid receptor by parabens was detected by glucocorticoid receptor competitor assays; and lastly, (4) parabens, butyl- and benzylparaben in particular, also promote adipose conversion of human adipose–derived multipotent stromal cells. Our results suggest that parabens may contribute to obesity epidemic, and the role of parabens in adipogenesis in vivo needs to be examined further.
PMCID: PMC3621350  PMID: 22956630
paraben; endocrine disrupting compound; adipocyte differentiation; glucocorticoid receptor; peroxisome proliferator–; activated receptor.
25.  Translocation of mixed lineage kinase domain-like protein to plasma membrane leads to necrotic cell death 
Cell Research  2013;24(1):105-121.
Mixed lineage kinase domain-like protein (MLKL) was identified to function downstream of receptor interacting protein 3 (RIP3) in tumor necrosis factor-α (TNF)-induced necrosis (also called necroptosis). However, how MLKL functions to mediate necroptosis is unknown. By reconstitution of MLKL function in MLKL-knockout cells, we showed that the N-terminus of MLKL is required for its function in necroptosis. The oligomerization of MLKL in TNF-treated cells is essential for necroptosis, as artificially forcing MLKL together by using the hormone-binding domain (HBD*) triggers necroptosis. Notably, forcing together the N-terminal domain (ND) but not the C-terminal kinase domain of MLKL causes necroptosis. Further deletion analysis showed that the four-α-helix bundle of MLKL (1-130 amino acids) is sufficient to trigger necroptosis. Both the HBD*-mediated and TNF-induced complexes of MLKL(ND) or MLKL are tetramers, and translocation of these complexes to lipid rafts of the plasma membrane precedes cell death. The homo-oligomerization is required for MLKL translocation and the signal sequence for plasma membrane location is located in the junction of the first and second α-helices of MLKL. The plasma membrane translocation of MLKL or MLKL(ND) leads to sodium influx, and depletion of sodium from the cell culture medium inhibits necroptosis. All of the above phenomena were not seen in apoptosis. Thus, the MLKL oligomerization leads to translocation of MLKL to lipid rafts of plasma membrane, and the plasma membrane MLKL complex acts either by itself or via other proteins to increase the sodium influx, which increases osmotic pressure, eventually leading to membrane rupture.
PMCID: PMC3879712  PMID: 24366341
MLKL; necroptosis; sodium influx; RIP3; plasma membrane translocation

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