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1.  Control of cotton fibre elongation by a homeodomain transcription factor GhHOX3 
Nature Communications  2014;5:5519.
Cotton fibres are unusually long, single-celled epidermal seed trichomes and a model for plant cell growth, but little is known about the regulation of fibre cell elongation. Here we report that a homeodomain-leucine zipper (HD-ZIP) transcription factor, GhHOX3, controls cotton fibre elongation. GhHOX3 genes are localized to the 12th homoeologous chromosome set of allotetraploid cotton cultivars, associated with quantitative trait loci (QTLs) for fibre length. Silencing of GhHOX3 greatly reduces (>80%) fibre length, whereas its overexpression leads to longer fibre. Combined transcriptomic and biochemical analyses identify target genes of GhHOX3 that also contain the L1-box cis-element, including two cell wall loosening protein genes GhRDL1 and GhEXPA1. GhHOX3 interacts with GhHD1, another homeodomain protein, resulting in enhanced transcriptional activity, and with cotton DELLA, GhSLR1, repressor of the growth hormone gibberellin (GA). GhSLR1 interferes with the GhHOX3–GhHD1 interaction and represses target gene transcription. Our results uncover a novel mechanism whereby a homeodomain protein transduces GA signal to promote fibre cell elongation.
Cotton fibre is the most important renewable material for textiles, with a huge economic output. Here the authors show that a homeodomain-leucine zipper transcription factor, GhHOX3, transduces a gibberellin signal that in turn promotes fibre cell elongation.
PMCID: PMC4263147  PMID: 25413731
2.  ABCG2 regulated by MAPK pathways is associated with cancer progression in laryngeal squamous cell carcinoma 
ATP-binding cassette, subfamily G, member 2 (ABCG2) overexpression has been associated with multidrug resistance and cancer progression by promoting proliferation and/or suppressing apoptosis, but how this process happens remains to be determined. In this study, the roles and the mechanisms of ABCG2 in the progression of Laryngeal squamous cell carcinoma (LSCC) were investigated. We found that introduction of ABCG2 siRNA into Hep-2 and Hep-2T cells significantly enhanced the intracellular accumulation of mitoxantrone (MX). Down-regulation of ABCG2 by transient RNAi inhibited cell proliferation and blocked cell cycle progression by regulating the expression of cyclin D3 and p21 Cip1. ABCG2 silence also induced cell apoptosis by regulating the expression of surviving, bcl-2 and the cleavage of poly (ADP-ribose) polymerase (PARP) in Hep-2 and Hep-2T cells. ABCG2-specific inhibitor, fumitremorgin C (FTC), and mitogen-activated protein kinase (MAPK) pathway inhibitor, U0126, inhibited cell proliferation and promoted cell apoptosis by degrading endogenous ABCG2 in Hep-2T cells. Furthermore, inhibition of MAPK pathway by U0126 enhanced anti-cancer effects of MX in vivo. In conclusion, suppression of ABCG2 inhibits the procession of LSCC tumor growth by regulating cell proliferation and apoptosis. Our data also provide more evidence for the importance of the MAPK pathway as a suitable therapeutic target for LSCC.
PMCID: PMC4266705  PMID: 25520861
BCG2; laryngeal squamous cell carcinoma; fumitremorgin C; MAPK pathway; mitoxantrone
3.  Post-infectious myositis ossificans in medial, lateral pterygoid muscles: A case report and review of the literature 
Oncology Letters  2014;9(2):920-926.
Myositis ossificans (MO) is a disease where heterotropic bone forms within a muscle or other type of soft tissue. MO is classified into two groups, MO progressiva and post-traumatic MO. It rarely occurs in the masticatory muscles and thus, only 20 cases involving the masticatory muscles have been reported since 2001. The majority of the reported cases occurred due to trauma, repeated injury or surgical manipulation. However, in a small number of cases, no specific traumatic event was identified as the cause of MO. To the best of our knowledge, this is the first case of post-infectious MO to be reported in the medial and lateral pterygoid muscles.
PMCID: PMC4301508  PMID: 25621069
myositis ossificans; infection; pterygoid muscle; computed tomography
4.  Inactivated vaccine with adjuvants consisting of pattern recognition receptor agonists confers protection against avian influenza viruses in chickens 
Veterinary microbiology  2014;172(0):120-128.
Use of adjuvant containing pathogen pattern recognition receptor agonists is one of the effective strategies to enhance the efficacy of licensed vaccines. In this study, we investigated the efficacy of avian influenza vaccines containing an adjuvant (CVCVA5) which was composed of polyriboinosinic polyribocytidylic, resiquimod, imiquimod, muramyl dipeptide and levomisole. Avian influenza vaccines adjuvanted with CVCVA5 were found to induce significantly higher titers of hemagglutiniton inhibition antibodies (P ≤ 0.01) than those of commercial vaccines at 2-, 3- and 4-week post vaccination in both specific pathogen free (SPF) chickens and field application. Furthermore, virus shedding was reduced in SPF chickens immunized with H9-CVCVA5 vaccine after H9 subtype heterologous virus challenge. The ratios of both CD3+CD4+ and CD3+CD8+ lymphocytes were slowly elevated in chickens immunized with H9-CVCVA5 vaccine. Lymphocytes adoptive transfer study indicates that CD8+ T lymphocyte subpopulation might have contributed to improved protection against heterologous virus challenge. Results of this study suggest that the adjuvant CVCVA5 was capable of enhancing the potency of existing avian influenza vaccines by increasing humoral and cellular immune response.
PMCID: PMC4208718  PMID: 24894132
Avian influenza virus; Inactivated vaccine; Adjuvants; Pattern recognition receptor; Agonist; Toll-like receptor
5.  RASSF1A Promoter Hypermethylation Is a Strong Biomarker of Poor Survival in Patients with Salivary Adenoid Cystic Carcinoma in a Chinese Population 
PLoS ONE  2014;9(10):e110159.
In addition to the clinicopathological parameters, molecular biomarkers are becoming increasingly important in the prognostic evaluation of cancer patients. This study aimed to determine the molecular alterations in the RAS association domain family protein1A gene (RASSF1A) in salivary adenoid cystic carcinoma (ACC) and to evaluate the potential of such alterations as prognostic markers. One hundred and sixty-seven ACC tumor tissues and 50 samples of matched normal salivary gland tissues from the same patients were analyzed for RASSF1A promoter methylation status by bisulfite sequencing PCR (BSP) and/or methylation-specific PCR (MSP). Fifty ACC tumor tissues and matched normal salivary gland tissues were analyzed for loss of heterozygosity (LOH) by examining two microsatellite markers (D3S1478, D3S1621) at 3p21. RASSF1A gene mutations were detected by direct sequencing of all six exons in 50 tumor and normal tissue specimens. Over-all, RASSF1A promoter hypermethylation was detected in 35.3% (59/167) of ACC tissues and was associated with histologically solid tumor pattern (P = 0.002) and advanced TNM stage (P = 0.014). RASSF1A LOH was observed in 18.0% (9/50) of cases, and no somatic mutation of RASSF1A was detected in any cases. RASSF1A promoter methylation was associated with the poor over-all survival (Log-rank test, P <0.001) and disease-free survival (Log-rank test, P <0.001) and identified as an independent predicator of over-all patient survival (P = 0.009) and disease-free survival (P <0.001). It was concluded that RASSF1A methylation is involved in the development, differentiation and progression of ACC and is a strong independent biomarker of poor survival in ACC patients in a Chinese population.
PMCID: PMC4193867  PMID: 25302792
6.  Neuroprotective effects of Asiaticoside 
Neural Regeneration Research  2014;9(13):1275-1282.
In the central nervous system, Asiaticoside has been shown to attenuate in vitro neuronal damage caused by exposure to β-amyloid. In vivo studies demonstrated that Asiaticoside could attenuate neurobehavioral, neurochemical and histological changes in transient focal middle cerebral artery occlusion animals. In addition, Asiaticoside showed anxiolytic effects in acute and chronic stress animals. However, its potential neuroprotective properties in glutamate-induced excitotoxicity have not been fully studied. We investigated the neuroprotective effects of Asiaticoside in primary cultured mouse cortical neurons exposed to glutamate-induced excitotoxicity invoked by N-methyl-D-aspartate. Pretreatment with Asiaticoside decreased neuronal cell loss in a concentration-dependent manner and restored changes in expression of apoptotic-related proteins Bcl-2 and Bax. Asiaticoside pretreatment also attenuated the upregulation of NR2B expression, a subunit of N-methyl-D-aspartate receptors, but did not affect expression of NR2A subunits. Additionally, in cultured neurons, Asiaticoside significantly inhibited Ca2+ influx induced by N-methyl-D-aspartate. These experimental findings provide preliminary evidence that during excitotoxicity induced by N-methyl-D-aspartate exposure in cultured cortical neurons, the neuroprotective effects of Asiaticoside are mediated through inhibition of calcium influx. Aside from its anti-oxidant activity, down-regulation of NR2B-containing N-methyl-D-aspartate receptors may be one of the underlying mechanisms in Asiaticoside neuroprotection.
PMCID: PMC4160853  PMID: 25221579
nerve regeneration; brain injury; Asiaticoside; apoptosis; N-methyl-D-aspartate; glutamate; neurotransmitter; neurotoxicity; calcium imaging; Bcl-2; Bax; NSFC grant; neural regeneration
7.  Low Dose CT Reconstruction via Edge-preserving Total Variation Regularization 
Physics in medicine and biology  2011;56(18):5949-5967.
High radiation dose in CT scans increases a lifetime risk of cancer and has become a major clinical concern. Recently, iterative reconstruction algorithms with Total Variation (TV) regularization have been developed to reconstruct CT images from highly undersampled data acquired at low mAs levels in order to reduce the imaging dose. Nonetheless, the low contrast structures tend to be smoothed out by the TV regularization, posing a great challenge for the TV method. To solve this problem, in this work we develop an iterative CT reconstruction algorithm with edge-preserving TV regularization to reconstruct CT images from highly undersampled data obtained at low mAs levels. The CT image is reconstructed by minimizing an energy consisting of an edge-preserving TV norm and a data fidelity term posed by the x-ray projections. The edge-preserving TV term is proposed to preferentially perform smoothing only on non-edge part of the image in order to better preserve the edges, which is realized by introducing a penalty weight to the original total variation norm. During the reconstruction process, the pixels at edges would be gradually identified and given small penalty weight. Our iterative algorithm is implemented on GPU to improve its speed. We test our reconstruction algorithm on a digital NCAT phantom, a physical chest phantom, and a Catphan phantom. Reconstruction results from a conventional FBP algorithm and a TV regularization method without edge preserving penalty are also presented for comparison purpose. The experimental results illustrate that both TV-based algorithm and our edge-preserving TV algorithm outperform the conventional FBP algorithm in suppressing the streaking artifacts and image noise under the low dose context. Our edge-preserving algorithm is superior to the TV-based algorithm in that it can preserve more information of low contrast structures and therefore maintain acceptable spatial resolution.
PMCID: PMC4026331  PMID: 21860076
8.  Aerobic Exercise Inhibits Sympathetic Nerve Sprouting and Restores β-Adrenergic Receptor Balance in Rats with Myocardial Infarction 
PLoS ONE  2014;9(5):e97810.
Cardiac sympathetic nerve sprouting and the dysregulation of β-adrenergic receptor (β-AR) play a critical role in the deterioration of cardiac function after myocardial infarction (MI). Growing evidence indicates that exercise provides protection against MI. The aims of this study were to investigate whether aerobic exercise following MI could inhibit sympathetic nerve sprouting and restore the balance of β3-AR/β1-AR.
Male Sprague-Dawley rats were divided into three groups: sham-operated control group (SC), MI group (MI), and MI with aerobic exercise group (ME). The rats in ME group were assigned to 8 weeks of exercise protocol (16 m/min, 50 min/d, 5 d/wk). The expression of nerve growth factor (NGF), the sympathetic nerve marker-tyrosine hydroxylase (TH), the nerve sprouting marker-growth associated protein 43 (GAP43), and β1- and β2-AR expression in the peri-infarct area of the left ventricle (LV) were measured by Western blot and immunohistochemistry, while β3-AR expression was determined by Western blot and immunofluorescence. Endothelial nitric oxide synthase (NOS2), phospho-NOS2 (p-NOS2), and neuronal nitric oxide synthase (NOS1) were measured by Western blot.
MI increased LV end-diastolic pressure (LVEDP), and decreased LV systolic pressure (LVSP). Compared with the MI group, aerobic exercise significantly decreased LVEDP and increased LVSP. The protein expression of TH, GAP43 and NGF was significantly increased after MI, which was normalized by exercise. Compared with the SC group, the ratios of β2-AR/β1-AR and β3-AR/β1-AR were elevated in the MI group, and the protein expression of β3-AR and NOS1 increased after MI. Compared with the MI group, the ratios of β2-AR/β1-AR and β3-AR/β1-AR were normalized in the ME group, while the protein expression of β3-AR and NOS1 significantly increased, and NOS2 was activated by exercise.
Aerobic exercise inhibits cardiac sympathetic nerve sprouting, restores β3-AR/β1-AR balance and increases β3-AR expression through the activation of NOS2 and NOS1 after myocardial infarction.
PMCID: PMC4026473  PMID: 24842290
9.  Astrocyte Elevated Gene-1 Mediates Glycolysis and Tumorigenesis in Colorectal Carcinoma Cells via AMPK Signaling 
Mediators of Inflammation  2014;2014:287381.
To investigate the role of AEG-1 in glycolysis and tumorigenesis, we construct myc-AEG-1 expression vector and demonstrate a novel mechanism that AEG-1 may increase the activity of AMPK by Thr172 phosphorylation. The higher expression levels of AEG-1 in colorectal carcinoma cells were found but showed significant difference in different cell lines. To study the role of AEG-1 in colorectal cells, myc-AEG-1 vector was constructed and transfected into NCM460 colonic epithelial cells. We observed consistent increasing of glucose consumption and lactate production, typical features of anaerobic glycolysis, suggesting that AEG-1 may promote anaerobic glycolysis. Moreover, we noted that AMPK phosphorylation at Thr172 as well as pPFK2 (Ser466) was increased in NCM460 cells overexpressing AEG-1. Compound C may block AMPK and PFK2 phosphorylation in both control and AEG-1-overexpressed cells and decrease the glucose consumption and lactate production. The present findings indicated that reduced AEG-1 protein levels by RNAi may decrease the glucose consumption and lactate production in HCT116 colorectal carcinoma cells. The present identified AEG-1/AMPK/PFK2 glycolysis cascade may be essential to cell proliferation and tumor growth. The present results may provide us with a mechanistic insight into novel targets controlled by AEG-1, and the components in the AEG-1/AMPK/PFK2 glycolysis process may be targeted for the clinical treatment of cancer.
PMCID: PMC4009221  PMID: 24829520
10.  Refractive Error and Risk of Early or Late Age-Related Macular Degeneration: A Systematic Review and Meta-Analysis 
PLoS ONE  2014;9(3):e90897.
To summarize relevant evidence investigating the associations between refractive error and age-related macular degeneration (AMD).
Systematic review and meta-analysis.
We searched Medline, Web of Science, and Cochrane databases as well as the reference lists of retrieved articles to identify studies that met the inclusion criteria. Extracted data were combined using a random-effects meta-analysis. Studies that were pertinent to our topic but did not meet the criteria for quantitative analysis were reported in a systematic review instead.
Main outcome measures
Pooled odds ratios (ORs) and 95% confidence intervals (CIs) for the associations between refractive error (hyperopia, myopia, per-diopter increase in spherical equivalent [SE] toward hyperopia, per-millimeter increase in axial length [AL]) and AMD (early and late, prevalent and incident).
Fourteen studies comprising over 5800 patients were eligible. Significant associations were found between hyperopia, myopia, per-diopter increase in SE, per-millimeter increase in AL, and prevalent early AMD. The pooled ORs and 95% CIs were 1.13 (1.06–1.20), 0.75 (0.56–0.94), 1.10 (1.07–1.14), and 0.79 (0.73–0.85), respectively. The per-diopter increase in SE was also significantly associated with early AMD incidence (OR, 1.06; 95% CI, 1.02–1.10). However, no significant association was found between hyperopia or myopia and early AMD incidence. Furthermore, neither prevalent nor incident late AMD was associated with refractive error. Considerable heterogeneity was found among studies investigating the association between myopia and prevalent early AMD (P = 0.001, I2 = 72.2%). Geographic location might play a role; the heterogeneity became non-significant after stratifying these studies into Asian and non-Asian subgroups.
Refractive error is associated with early AMD but not with late AMD. More large-scale longitudinal studies are needed to further investigate such associations.
PMCID: PMC3946285  PMID: 24603619
11.  Exercise Training Attenuates Hypertension and Cardiac Hypertrophy by Modulating Neurotransmitters and Cytokines in Hypothalamic Paraventricular Nucleus 
PLoS ONE  2014;9(1):e85481.
Regular exercise as an effective non-pharmacological antihypertensive therapy is beneficial for prevention and control of hypertension, but the central mechanisms are unclear. In this study, we hypothesized that chronic exercise training (ExT) delays the progression of hypertension and attenuates cardiac hypertrophy by up-regulating anti-inflammatory cytokines, reducing pro-inflammatory cytokines (PICs) and restoring the neurotransmitters balance in the hypothalamic paraventricular nucleus (PVN) in young spontaneously hypertensive rats (SHR). In addition, we also investigated the involvement of nuclear factor-κB (NF-κB) p65 and NAD(P)H oxidase in exercise-induced effects.
Methods and results
Moderate-intensity ExT was administrated to young normotensive Wistar-Kyoto (WKY) and SHR rats for 16 weeks. SHR rats had a significant increase in mean arterial pressure and cardiac hypertrophy. SHR rats also had higher levels of glutamate, norepinephrine (NE), phosphorylated IKKβ, NF-κB p65 activity, NAD(P)H oxidase subunit gp91phox, PICs and the monocyte chemokine protein-1 (MCP-1), and lower levels of gamma-aminobutyric acid (GABA) and interleukin-10 (IL-10) in the PVN. These SHR rats also exhibited higher renal sympathetic nerve activity (RSNA), and higher plasma levels of PICs, and lower plasma IL-10. However, ExT ameliorates all these changes in SHR rats.
These findings suggest that there are the imbalances between excitatory and inhibitory neurotransmitters and between pro- and anti-inflammatory cytokines in the PVN of SHR rats, which at least partly contributing to sympathoexcitation, hypertension and cardiac hypertrophy; chronic exercise training attenuates hypertension and cardiac hypertrophy by restoring the balances between excitatory and inhibitory neurotransmitters and between pro- and anti-inflammatory cytokines in the PVN; NF-κB and oxidative stress in the PVN may be involved in these exercise-induced effects.
PMCID: PMC3901693  PMID: 24482680
12.  Autophagy in Retinal Ganglion Cells in a Rhesus Monkey Chronic Hypertensive Glaucoma Model  
PLoS ONE  2013;8(10):e77100.
Primary open angle glaucoma (POAG) is a neurodegenerative disease characterized by physiological intraocular hypertension that causes damage to the retinal ganglion cells (RGCs). In the past, RGC damage in POAG was suggested to have been attributed to RGC apoptosis. However, in the present study, we applied a model closer to human POAG through the use of a chronic hypertensive glaucoma model in rhesus monkeys to investigate whether another mode of progressive cell death, autophagy, was activated in the glaucomatous retinas. First, in the glaucomatous retinas, the levels of LC3B-II, LC3B-II/LC3B-I and Beclin 1 increased as demonstrated by Western blot analyses, whereas early or initial autophagic vacuoles (AVi) and late or degraded autophagic vacuoles (AVd) accumulated in the ganglion cell layer (GCL) and in the inner plexiform layer (IPL) as determined by transmission electron microscopy (TEM) analysis. Second, lysosome activity and autophagosome-lysosomal fusion increased in the RGCs of the glaucomatous retinas, as demonstrated by Western blotting against lysosome associated membrane protein-1 (LAMP1) and double labeling against LC3B and LAMP1. Third, apoptosis was activated in the glaucomatous eyes with increased levels of caspase-3 and cleaved caspase-3 and an increased number of TUNEL-positive RGCs. Our results suggested that autophagy was activated in RGCs in the chronic hypertensive glaucoma model of rhesus monkeys and that autophagy may have potential as a new target for intervention in glaucoma treatment.
PMCID: PMC3797129  PMID: 24143204
13.  Ocular Hypertension after Pediatric Cataract Surgery: Baseline Characteristics and First-Year Report 
PLoS ONE  2013;8(7):e69867.
Monitoring intraocular pressure (IOP) is essential for pediatric cataract treatment but always difficult due to lack of cooperation in young children. We present the baseline characteristics and the first-year results of a long-term prospective cohort study, which are aimed to determine the relationship of the incidence of ocular hypertension (OH) in children after cataract surgery during the first-year period and the risk of developing late-onset glaucoma. Children were included with the following criteria: they were≤10 years old and scheduled to undergo cataract surgery with/without intraocular lens implantation; they were compliant with our follow-up protocol, which included monitoring IOP using a Tono-Pen under sedation or anesthesia. Incidence of OH, peak OH value, OH onset time and OH duration within a 12-month period following surgery were measured. In brief, 206 patients (379 eyes) were included and OH developed in 66 of 379 (17.4%) eyes. The mean follow-up period was 14.0±3.2 months (median, 12 months; range, 10–16 months). Moreover, 33 of 196 (16.8%) aphakic eyes and 33 of 183 (18.0%) IOL eyes were diagnosed with OH. The peak OH onset times were at 1-week (34/66, 51.5%) and 1-month (14/66, 21.2%) appointments postsurgery. The peak IOP value in the OH eyes was 29.9±7.5 mmHg (median, 29 mmHg; range, 21–48 mmHg). The duration of OH was 30.9±31.2 days (median, 30 days; range, 3–150 days). OH recurred in 13 eyes with a history of OH diagnosed within 1 month postsurgery (13/54, 24.1%), which needed temporary or long term use of antiglaucoma medications. In conclusion, the incidence of OH in children after cataract surgery was 17.4% during the first-year period. Children who have suffered elevated IOP in the first year after cataract surgery should be followed closely to determine if there is an increased risk of developing late-onset glaucoma.
PMCID: PMC3726742  PMID: 23922832
14.  Prognostic significance of p21, p27 and survivin protein expression in patients with oral squamous cell carcinoma 
Oncology Letters  2013;6(2):381-386.
Oral squamous cell carcinoma (OSCC) accounts for >80% of head and neck malignancies. p21, p27 and survivin proteins are abnormally expressed in OSCC and have been previously reported to correlate with cell proliferation and apoptosis. However, the prognostic significance of p21, p27 and survivin remains controversial. The aim of the present study was to investigate the association of clinical parameters and prognosis with the levels of p21, p27 and survivin expression in patients with OSCC. The levels of the three biomarkers were evaluated by immunohistochemical staining in specimens from 110 patients with OSCC and each section was scored according to the percentage of positive tumor cells and staining intensity. Log-rank test and Cox proportional hazards regression were performed to assess the correlation between biomarkers and clinical events. The association between the immunoexpression of p21, p27 and survivin and clinical pathological variables were analyzed by the χ2 test and a non-parametric analysis. The expression of p21 in patients with OSCC was found to correlate with the expression of p27 and survivin. The results of the current study revealed that the five-year survival rate was significantly lower in patients with high p21 expression. In addition, the expression of p27 also showed a negative correlation with the five-year survival rate of OSCC, but to a lesser extent. By contrast, the expression of survivin was not a prognostic factor for OSCC. A Kaplan-Meier analysis and Cox proportional hazards model showed that lymph node metastasis and p21 expression were independent prognostic factors of OSCC.
PMCID: PMC3789108  PMID: 24137333
p21; p27; survivin; oral squamous cell carcinoma; biomarker; prognosis
15.  Expression of ERBB3 binding protein 1 (EBP1) in salivary adenoid cystic carcinoma and its clinicopathological relevance 
BMC Cancer  2012;12:499.
ERBB3 binding protein 1 (EBP1) gene transfer into human salivary adenoid cystic carcinoma cells has been shown to significantly inhibit cell proliferation and reduce tumor metastasis in mouse models. In the current study, to evaluate if EBP1 is a novel biomarker capable of identifying patients at higher risk of disease progression and recurrence, we examined the EBP1 expression profile in adenoid cystic carcinoma (ACC) patients and analyzed its clinicopathological relevance. To understand the underlying anti-metastatic mechanism, we investigated if EBP1 regulates invasion-related molecules.
We performed immunohistochemical analysis on 132 primary adenoid cystic carcinoma and adjacent non-cancerous tissues using commercial EBP1, MMP9, E-cadherin and ICAM-1 antibodies. Results were correlated to clinicopathological parameters, long-term survival and invasion-related molecules by statistical analysis. Cell motility and invasiveness of vector or wild-type EBP1-transfected ACC-M cell lines were evaluated using wound healing and Boyden chamber assays. MMP9, E-cadherin and ICAM-1 proteins in these cell lines were detected using western blot assay.
The expression of EBP1 was significantly higher in non-cancerous adjacent tissues compared with corresponding cancer tissues. The intensity and percentage of cells that reacted with EBP1 antibodies were significantly higher in cases with tubular pattern than those with solid pattern (P<0.0001). We also found adenoid cystic carcinoma with local lymphatic metastasis had significantly lower EBP1 expression than ACC with no local lymphatic node metastasis (P<0.0001). Similar findings were observed in ACC with lung metastasis compared with cases with no lung metastasis (P<0.0001), in particular, in cases with perineural invasion compared with cases with no perineural invasion (P<0.0001). Furthermore, a decrease in EBP1 expression was positively associated with a reduction in overall survival of ACC patients. Of note, EBP1 inhibits migration and invasiveness of ACC cells by upregulating E-cadherin but downregulating MMP9. In clinical adenoid cystic carcinoma patients, higher EBP1 expression was positively correlated with E-cadherin levels (P<0.001) but negatively correlated with MMP9 expression (P=0.0002).
EBP1 expression is reduced in adenoid cystic carcinoma, indicating unfavorable prognosis of ACC patients. Its regulation of MMP9 and E-cadherin protein levels suggests a critical therapeutic potential.
PMCID: PMC3499390  PMID: 23110497
16.  Microwave-assisted one-pot synthesis of water-soluble rare-earth doped fluoride luminescent nanoparticles with tunable colors 
Polyethyleneimine (PEI) functionalized multicolor luminescent LaF3 nanoparticles were synthesized via a novel microwave-assisted method, which can achieve fast and uniform heating under eco-friendly and energy efficient conditions. The as-prepared nanoparticles possess a pure hexagonal structure with an average size of about 12 nm. When doped with different ions (Tb3+ and Eu3+), the morphology and structure of the nanoparticles were not changed, whereas the optical properties varied with doped ions and their molar ratio, and as a result emission of four different colors (green, yellow, orange and red) were achieved by simply switching the types of doping ions (Eu3+ versus Tb3 +) and the molar ratio of the two doping ions.
PMCID: PMC3413302  PMID: 22879690
Microwave-assisted synthesis; Multicolor luminescence; Nanomaterials
17.  Birth Weight, Postnatal Weight Change, and Risk for High Blood Pressure Among Chinese Children 
Pediatrics  2011;127(5):e1272-e1279.
It is hypothesized that a physiological predisposition toward hypertension results from a combination of intrauterine growth restriction or overgrowth and excessive postnatal weight gain. Previous studies were conducted largely in Western countries however the hypothesis may also be relevant in developing countries where metabolic disorders are increasing.
We investigated the association of birth weight and postnatal weight gain with hypertension among Chinese children.
A population based study was conducted among 15 600 children aged 3 to 6 years from Tianjin, China. Weight was expressed as z scores. Postnatal weight gain was defined as changes in z scores from birth to 3 to younger than 4 years, 4 to younger than 5 years, and 5 to 6 years. Hypertension was defined as greater than the 90th percentile of either systolic or diastolic blood pressure. Logistic regression-derived odds ratios and 95% confidence intervals were generated to estimate the association between birth weight and postnatal weight gain with hypertension risk in childhood.
Birth weight was positively associated with childhood hypertension in boys and girls (odds ratios [95% confidence interval] comparing extreme quartiles [high versus low] were 5.67 [3.83–8.39] and 2.58 [3.83–8.39], respectively). Postnatal weight gain was positively associated with hypertension and the association did not significantly vary by birth size for gestational age.
Greater birth weight or postnatal weight gain was associated with increased childhood hypertension risk, suggesting that intrauterine growth and postnatal weight gain may have implications on health during childhood.
PMCID: PMC3387869  PMID: 21502227
birth weight; hypertension; fetal origins
18.  Simultaneous Identification and Quantification of Canrenone and 11-α-Hydroxy-Canrenone by LC-MS and HPLC-UVD 
A procedure for simultaneous identification and quantification of canrenone and its biotransformed product 11-α-hydroxy-canrenone by high-performance liquid chromatography with ultraviolet detector (HPLC-UVD) and mass spectrometry (LC-MS) methods was proposed. The optimal determination variables on the HPLC-UVD or LC-MS coupled with a ZORBAX Eclipse XDB-C18 column (150 mm × 4.6 mm, 5 μm) were set as follows: detection wavelength of 280 nm, mobile phase of water and methanol gradient elution, temperature for the chromatographic column of 30°C, flow rate of mobile phase of 0.8 mL/min, sample injection volume of 5 μL, and elution time of 40 min. The MS conditions were set as follows: the flow rate of sheath gas, aux gas, and sweep gas were kept at 35 arb, 5 arb, and 0 arb, respectively. The temperature of capillary was held at 300°C, and capillary voltage was set at 30.00 V. Tube lens were performed at 100.00 V. The proposed method was validated by linearity (r2 ≥ 0.9910), average recovery (94.93%, RSD1.21%), precision (RSD ≤ 1.31%), limit of detection, and limit of quantification (LOD 0.1~0.12 mg/L, LOQ 0.5~0.67 mg/L), which proved to be affordable for simultaneously determining canrenone and its bio-transformed product 11-α-hydroxy-canrenone.
PMCID: PMC3238810  PMID: 22203787
19.  Hepatoma cell line HepG2.2.15 demonstrates distinct biological features compared with parental HepG2 
AIM: To investigate the biological features of hepatitis B virus (HBV)-transfected HepG2.2.15 cells.
METHODS: The cell ultrastructure, cell cycle and apoptosis, and the abilities of proliferation and invasion of HBV-transfected HepG2.2.15 and the parent HepG2 cells were examined by electron microscopy, flow cytometry, 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and trans-well assay. Oncogenicity of the two cell lines was compared via subcutaneous injection and orthotopic injection or implantation in nude mice, and the pathological analysis of tumor formation was performed. Two cytoskeletal proteins were detected by Western blotting.
RESULTS: Compared with HepG2 cells, HepG2.2.15 cells showed organelle degeneration and filopodia disappearance under electron microscope. HepG2.2.15 cells proliferated and migrated slowly in vitro, and hardly formed tumor and lung metastasis in nude mice. Flow cytometry showed that the majority of HepG2.2.15 cells were arrested in G1 phase, and apoptosis was minor in both cell lines. Furthermore, the levels of cytoskeletal proteins F-actin and Ezrin were decreased in HepG2.2.15 cells.
CONCLUSION: HepG2.2.15 cells demonstrated a lower proliferation and invasion ability than the HepG2 cells due to HBV transfection.
PMCID: PMC3063907  PMID: 21448419
HepG2.2.15; HepG2; Hepatitis B virus; Biological feature; Tumor
20.  Anti-proliferation effects of Sirolimus sustained delivery film in rabbit glaucoma filtration surgery 
Molecular Vision  2011;17:2495-2506.
To investigate the efficacy, safety, and mechanisms of Sirolimus sustained delivery film on prevention of scar formation in a rabbit model of glaucoma filtration surgery.
Sixty-four New Zealand white rabbits who underwent trabeculectomy in the right eye were randomly allocated to one of the four treatment regimens: Sirolimus sustained delivery film treatment group (Group A), or drug-free film treatment group (Group B), or 30 ng/ml Sirolimus-soaked sponge treatment group (Group C), or no adjunctive treatment group (Group D), and each group consists of 16 rabbits. Intraocular pressure (IOP), morphologic changes of bleb, anterior chamber flare, and corneal endothelial cell count and complications were evaluated over a 28-day period follow-up time. Aqueous humor samples were gathered from Group A, and the concentration of Sirolimus was measured regularly post-operation. Rabbits were sacrificed on the 7th, 14th, and 28th day post-operation separately, and the fibroblast hypertrophy, infiltration of inflammatory, and proliferation of new collagen fiber formation in each group were evaluated with HE and Masson staining. Proliferative cell nuclear antigen (PCNA) and fibroblast apoptosis were evaluated by immunohistochemistry and terminal deoxynucleotidyl transferasemediated dUTP nick end labeling (TUNEL) assay at the 28th day post-operation.
Both Sirolimus sustained delivery film (Group A) and Sirolimus alone (Group C) were well tolerated in this model, and significantly prolonged bleb survival compared with no drug treatment group (Group B and D; p<0.001). Group A had the longest bleb survival time in comparison with other groups (p<0.001). There were significant differences in IOP readings between Group A and other groups at the last follow-up (p<0.05). The concentration of Group A maintained stable for over 2 weeks, drops from (10.56 ±0.05) ng/ml at day 3 to (7.74 ±0.05) ng/ml at day 14. The number of corneal endothelial cells of Group A was not statistically significant between pre and post-operation. Histologic examination demonstrated that eyes treated with Sirolimus, especially the Sirolimus sustained delivery film, showed an obvious reduction in subconjunctival fibroblast scar tissue formation compared with no drug treatment groups, and had minimal evidence of inflammatory cell infiltration and new collagen deposition in the subconjunctiva. Immunohistochemistry assay showed that PCNA-expression was lower in the Group A (16.25±3.24%) compared to other groups (p<0.01). TUNEL assay showed a significant increase in the number of apoptotic fibroblasts around the surgical area in Group A and Group C (9.75±1.71% and 8.50±1.92%) compared to the Group B and D (p<0.01).
Sirolimus drug sustained delivery film can inhibit inflammatory cell activity, impede fibroblast proliferation activity, and induce fibroblast apoptosis in the filtration surgery sites in rabbit. The results indicate a safe and effective treatment strategy in anti-scaring treatment in glaucoma surgery.
PMCID: PMC3185021  PMID: 21976960
21.  Cantilever measurements of surface stress, surface reconstruction, film stress and magnetoelastic stress of monolayers 
Sensors (Basel, Switzerland)  2008;8(7):4466-4486.
We review the application of cantilever-based stress measurements in surface science and magnetism. The application of thin (thickness appr. 0.1 mm) single crystalline substrates as cantilevers has been used successfully to measure adsorbate-induced surface stress changes, lattice misfit induced film stress, and magneto-elastic stress of ferromagnetic monolayers. Surface stress changes as small as 0.01 N/m can be readily measured, and this translates into a sensitivity for adsorbate-coverage well below 0.01 of one layer. Stress as large as several GPa, beyond the elasticity limits of high strength materials, is measured, and it is ascribed to the lattice misfit between film and substrate. Our results point at the intimate relation between surface stress and surface reconstruction, stress-induced structural changes in epitaxially strained films, and strain-induced modifications of the magneto-elastic coupling in ferromagnetic monolayers.
PMCID: PMC3697186
cantilever stress measurement; surface stress; surface reconstruction; film stress; magnetoelastic stress; torque magnetometry
22.  Temporal and Spatial Control of HGC1 Expression Results in Hgc1 Localization to the Apical Cells of Hyphae in Candida albicans▿  
Eukaryotic Cell  2006;6(2):253-261.
The human fungal pathogen Candida albicans can undergo a morphological transition from a unicellular yeast growth form to a multicellular hyphal growth form. During hyphal growth, cell division is asymmetric. Only the apical cell divides, whereas subapical cells remain in G1, and cell surface growth is highly restricted to the tip of the apical cell. Hgc1, a hypha-specific, G1 cyclin-like protein, is essential for hyphal development. Here, we report, using indirect immunofluorescence, that Hgc1 is preferentially localized to the dividing apical cells of hyphae. Hgc1 protein is rapidly degraded in a cell cycle-independent manner, and the protein turnover likely occurs in both the apical and the subapical cells of hyphae. In addition to rapid protein turnover, the HGC1 transcript is also dynamically regulated during cell cycle progression in hyphal growth. It is induced upon germ tube formation in early G1; the transcript level is reduced during the G1/S transition and peaks again around the G2/M phase in the subsequent cell cycles. Transcription from the HGC1 promoter is essential for its apical cell localization, as Hgc1 no longer exhibits preferential apical localization when expressed under the MAL2 promoter. Using fluorescence in situ hybridization, the HGC1 transcript is detected only in the apical cells of hyphae, suggesting that HGC1 is transcribed in the apical cell. Therefore, the preferential localization of Hgc1 to the apical cells of hyphae results from the dynamic temporal and spatial control of HGC1 expression.
PMCID: PMC1797949  PMID: 17172437
23.  Effect of Fructus Psoraleae on motility of gallbladder isolated smooth muscle strips from guinea pigs 
AIM: To observe the effect of Fructus Psoraleae on motility of isolated gallbladder muscle strips of guinea pigs and its mechanism.
METHODS: Guinea pigs were hit to lose consciousness and the whole gallbladder was removed quickly. Two or three smooth muscle strips (8 mm × 3 mm) were cut along a longitudinal direction. The mucosa was gently removed. Every longitudinal muscle strip was suspended in a tissue chamber which was continuously perfused with 5 mL Krebs solution (37°C), pH 7.4, and aerated with 950 mL/L O2 and 50 mL /L CO2. The isometric response was recorded with an ink-writing recorder. After 2 h equilibration under 1 g-load, 50 μL Fructus Psoraleae (10, 20, 70, 200, 700, 1000 g/L) was added cumulatively into the tissue chamber in turn every 2 min to observe their effects on gallbladder muscle strips (cumulating final concentration of Fructus Psoraleae was 0.1, 0.3, 1.0, 3.0, 10.0, 20.0 g/L). The antagonists, including 4-DAMP, benzhydramine, hexamethonium, phentolamine, verapamil and idomethine were given 2 min before Fructus Psoraleae respectively to investigate the mechanisms involved.
RESULTS: Fructus Psoraleae dose-dependently increased the resting tension (r = 0.992, P < 0.001), decreased the mean contractile amplitude (r = 0.970, P < 0.001) and meanwhile increased the contractile frequency of the gallbladder muscle strip in vitro (r = 0.965, P < 0.001). The exciting action of Fructus Psoraleae on the resting tension could be partially blocked by 4-DAMP (the resting tension decreased from 1.37 ± 0.41 to 0.70 ± 0.35, P < 0.001), benzhydramine (from 1.37 ± 0.41 to 0.45 ± 0.38, P < 0.001), hexamethonium (from 1.37 ± 0.41 to 0.94 ± 0.23, P < 0.05), phentolamine ( from 1.37 ± 0.41 to 0.89 ± 0.22, P < 0.01) and verapamil (from 1.37 ± 0.41 to 0.94 ± 0.26, P < 0.05). But the above antagonists had no significant effect on the action of Fructus Psoraleae–induced mean contractile amplitude (P > 0.05). Moreover, the increase of the contractile frequency due to Fructus Psoraleae was inhibited by 4-DAMP (decreased from 8.3 ± 1.2 to 6.8 ± 0.5, P < 0.01) and hexamethonium (from 8.3 ± 1.2 to 7.0 ± 0.9, P < 0.05). Idomethine had no significant effect on the Fructus Psoraleae-induced responses (P > 0.05).
CONCLUSION: Fructus Psoraleae enhances the motility of isolated gallbladder muscle strips from guinea pigs, in a dose-dependent manner. The effect of Fructus Psoraleae is partly related to M3, N receptor, α receptor, H1 receptor, Ca2+ channel, but not related to prostaglandin.
PMCID: PMC4088023  PMID: 16937536
Fructus Psoraleae; Gallbladder smooth muscle strips; M3, N, α, H1 receptors; Ca2+ channel; Prostaglandin
24.  Effects of rhubarb on isolated gastric muscle strips of guinea pigs 
AIM: To study the effects of rhubarb (dried root of Rheum officinale Baill.) on contractile activity of isolated gastric muscle strips of guinea pigs and its possible mechanism.
METHODS: A total of 48 guinea pigs were killed to remove the whole stomach. Then, the stomach was opened and the mucosal layer was removed. Parallel to the circular fibers, muscle strips were cut from the body. Each isolated gastric muscle strip was suspended in a tissue chamber containing 5 mL Krebs solution, constantly warmed by water jacket at 37 °C and bubbled continuously with a mixed gas of 950 mL/L O2 and 50 mL/L CO2. After being incubated for 1 h with 1 g tension, rhubarb of varied concentrations (1%, 2%, 7%, 20% and 70%) was added cumulatively into the tissue chamber at intervals of 2 min. Atropine (10-6 mol/L) or isoptin (5×10-8 mol/L) or hexamethonium (10-5 mol/L) was given 2 min before the administration of rhubarb. The isometrical response was measured with an ink-writing recorder.
RESULTS: Rhubarb dose dependently increased the resting tension of gastric body circular muscle (CM) (r = 0.726, P<0.05). Atropine (r = 0.829, P<0.05), isoptin (r = 0.764, P<0.05) and hexamethonium (r = 0.797, P<0.05) did not affect its action in a dose-related manner. Atropine apparently reduced the increasing action of 1%, 3%, 10%, 30% and 100% rhubarb on the resting tension of gastric body CM. Isoptin inhibited the effect of 10%, 30% and 100% rhubarb on the resting tension of gastric body CM. Hexamethonium reduced the increasing action of 1%, 10%, 30% and 100% rhubarb on the resting tension of gastric body CM. Rhubarb increased the contractile frequency of CM of body. While atropine, isoptin and hexamethonium did not inhibit the contractile frequency of gastric body CM in comparison with rhubarb at the same concentration, rhubarb at the highest concentration (100%) decreased the mean contractile amplitude of gastric body CM. Atropine, isoptin and hexamethonium did not affect the mean contractile amplitude of gastric body CM compared to rhubarb at the same concentration.
CONCLUSION: Rhubarb has exciting actions on isolated gastric smooth muscle strips of guinea pig. The exciting action of rhubarb is partly mediated via cholinergic M receptor, cholinergic N receptor and L-type calcium channel.
PMCID: PMC4305764  PMID: 15849832
Rhubarb; Stomach; Smooth muscle; Muscle contraction; Cholinergic receptor; Calcium channel; Guinea pigs
25.  Effect of rhubarb on contractile response of gallbladder smooth muscle strips isolated from guinea pigs 
AIM: To investigate the effect of rhubarb on contractile response of isolated gallbladder muscle strips from guinea pigs and its mechanism.
METHODS: Guinea pigs were killed to remove the whole gallbladder. Two or three smooth muscle strips (8 mm×3 mm) were cut along the longitudinal direction. The mucosa on each strip was carefully removed. Each longitudinal muscle strip was suspended in a tissue chamber containing 5 mL Krebs solution (37 °C), bubbled continuously with 950 mL/L O2 and 50 mL/L CO2. The resting tension (g), mean contractile amplitude (mm), and contractile frequency (waves/min) were simultaneously recorded on recorders. After 2-h equilibration, rhubarb (10, 20, 70, 200, 700, 1000 g/L) was added cumulatively to the tissue chamber in turns every 2 min to observe their effects on gallbladder. Antagonists were given 3 min before administration of rhubarb to investigate the possible mechanism.
RESULTS: Rhubarb increased the resting tension (from 0 to 0.40±0.02, P<0.001), and decreased the mean contractile amplitude (from 5.22±0.71 to 2.73±0.41, P<0.001). It also increased the contractile frequency of the gallbladder muscle strips in guinea pigs (from 4.09±0.46 to 6.08±0.35, P<0.001). The stimulation of rhubarb on the resting tension decreased from 3.98±0.22 to 1.58±0.12 by atropine (P<0.001), from3.98±0.22 to 2.09±0.19 by verapamil (P<0.001) and from 3.98±0.22 to 2.67±0.43 by phentolamine (P<0.005). But the effect was not inhibited by hexamethonium (P>0.05). In addition, the action of mean amplitude and frequency was not inhibited by the above antagonists.
CONCLUSION: Rhubarb can stimulate the motility of isolated gallbladder muscle strips from guinea pigs. The stimulation of rhubarb might be relevant with M receptor, Ca2+ channel and α receptor partly.
PMCID: PMC4250598  PMID: 15682482
Rhubarb; Gallbladder smooth muscle strips; M receptor; Ca2+ channel; α receptor

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