The mechanism of action of artemisinin and its derivatives, the most potent of the anti-malarial drugs, is not completely understood. Here we present an unbiased chemical proteomics analysis to directly explore this mechanism in Plasmodium falciparum. We use an alkyne-tagged artemisinin analogue coupled with biotin to identify 124 artemisinin covalent binding protein targets, many of which are involved in the essential biological processes of the parasite. Such a broad targeting spectrum disrupts the biochemical landscape of the parasite and causes its death. Furthermore, using alkyne-tagged artemisinin coupled with a fluorescent dye to monitor protein binding, we show that haem, rather than free ferrous iron, is predominantly responsible for artemisinin activation. The haem derives primarily from the parasite's haem biosynthesis pathway at the early ring stage and from haemoglobin digestion at the latter stages. Our results support a unifying model to explain the action and specificity of artemisinin in parasite killing.
The mechanism of action of artemisinin, an antimalarial drug, is not well understood. Here, the authors use a labelled artemisinin analogue to show that the drug is mainly activated by haem and then binds covalently to over 120 proteins in the malaria parasite, affecting many of its cellular processes.
High fluorescence quantum yield graphene quantum dots (GQDs) have showed up as a new generation for bioimaging. In this work, luminescent GQDs were prepared by an ameliorative photo-Fenton reaction and a subsequent hydrothermal process using graphene oxide sheets as the precursor. The as-prepared GQDs were nanomaterials with size ranging from 2.3 to 6.4 nm and emitted intense green luminescence in water. The fluorescence quantum yield was as high as 24.6% (excited at 340 nm) and the fluorescence was strongest at pH 7. Moreover, the influences of low-concentration (12.5, 25 μg/mL) GQDs on the morphology, viability, membrane integrity, internal cellular reactive oxygen species level and mortality of HeLa cells were relatively weak, and the in vitro imaging demonstrated GQDs were mainly in the cytoplasm region. More strikingly, zebrafish embryos were co-cultured with GQDs for in vivo imaging, and the results of heart rate test showed the intake of small amounts of GQDs brought little harm to the cardiovascular of zebrafish. GQDs with high quantum yield and strong photoluminescence show good biocompatibility, thus they show good promising for cell imaging, biolabeling and other biomedical applications.
Growth arrest and DNA-damage inducible protein 45 beta (Gadd45b) is serving as a neuronal activity sensor. Brain ischemia induces the expression of Gadd45b, which stimulates recovery after stroke and may play a protective role in cerebral ischemia. However, little is known of the molecular mechanisms of how Gadd45b expression regulated and the down-stream targets in brain ischemia. Here, using an oxygen-glucose deprivation and reperfusion (OGD/R) model, we identified Huwe1/Mule/ARF-BP1, a HECT domain containing ubiquitin ligase, involved in the control of Gadd45b protein level. In this study, we also investigated the role of Huwe1-Gadd45b mediated pathway in BDNF methylation.
We found that the depletion of Huwe1 by lentivirus shRNA mediated interference significantly increased the expression of Gadd45b and BDNF at 24 h after OGD. Moreover, treatment with Cycloheximide (CHX) inhibited endogenous expression of Gadd45b, and promoted expression of Gadd45b after co-treated with lentivirus shRNA-Huwe1. Inhibition of Gadd45b by lentivirus shRNA decreased the expression levels of brain derived neurotrophic factor (BDNF) and phosphorylated cAMP response element-binding protein (p-CREB) pathway, while inhibition of Huwe1 increased the expression levels of BDNF and p-CREB. Moreover, shRNA-Huwe1 treatment decreased the methylation level of the fifth CpG islands (123 bp apart from BDNF IXa), while shRNA-Gadd45b treatment increased the methylation level of the forth CpG islands (105 bp apart from BDNF IXa).
These findings suggested that Huwe1 involved in the regulation of Gadd45b expression under OGD/R, providing a novel route for neurons following cerebral ischemia-reperfusion injury. It also indicated that the methylation of BDNF IXa was affected by Gadd45b as well as Huwe1 in the OGD/R model.
Electronic supplementary material
The online version of this article (doi:10.1186/s13041-015-0178-y) contains supplementary material, which is available to authorized users.
Gadd45b; Huwe1; BDNF; Oxygen-glucose deprivation and reperfusion; Methylation
Vascular complications induced by intercavernous sinus injury during dural opening in the transsphenoidal surgery may contribute to incomplete tumour resections. Preoperative neuro-imaging is of crucial importance in planning surgical approach. The aim of this study is to correlate the microanatomy of intercavernous sinuses with its contrast-enhanced magnetic resonance venography (CE-MRV).
Eighteen human adult cadavers and 24 patients were examined based on autopsy and CE-MRV. Through dissection of the cadavers and CE-MRV, the location, shape, number, diameter and type of intercavernous sinuses were measured and compared.
Different intercavernous sinuses were identified by their location and shape in all the cadavers and CE-MRV. Compared to the cadavers, CE-MRV revealed 37% of the anterior intercavernous sinus, 48% of the inferior intercavernous sinus, 30% of the posterior intercavernous sinus, 30% of the dorsum sellae sinus and 100% of the basilar sinus. The smaller intercavernous sinuses were not seen in the neuro-images. According to the presence of the anterior and inferior intercavernous sinus, four types of the intercavernous sinuses were identified in cadavers and CE-MRV, and the corresponding operative space in the transsphenoidal surgical approach was implemented.
The morphology and classification of the cavernous sinus can be identified by CE-MRV, especially for the larger vessels, which cause bleeding more easily. Therefore, CE-MRV provides a reliable measure for individualized preoperative planning during transsphenoidal surgery.
Diabetes has been shown to be significantly associated with poor outcome after stroke. However, the sex differences in stroke outcome among patients with diabetes are unknown. Therefore, we aimed to assess the sex differences in long-term prognosis among acute ischemic stroke patients with diabetes.
The ischemic stroke patients with diabetes were recruited to this study between May 2005 and September 2014 in Tianjin, China. Sex differences in mortality, dependency (modified rank scale > 2), and recurrence at 3, 12, and 36 months after stroke were analyzed.
A total of 2360 patients were recruited in this study. The age of stroke onset, National Institute of Health stroke scale (NIHSS), and modified rank scale (mRS) on admission were greater in women than in men (P < 0.05). Women were more likely to have hypertension, obesity, atrial fibrillation, and dyslipidemias. In contrast, men were more likely to have artery stenosis, current smoking, and alcohol consumption (P < 0.001). There was higher mortality in women than in men at 3 months (7.9 % vs 5.2 %), 12 months (12.2 % vs 8.2 %), and 36 months (21.9 % vs 16.1 %) after stroke; but no differences were found in dependency and recurrence. Sex differences were found in associated factors of outcomes by time-point. Trial of Org 10172 in Acute Stroke Treatment (TOAST) of large artery atherothrombosis (LAA), cardioembolism (CE), and smoking were risk factors of outcomes in women at short term and medium term; but atrial fibrillation (AF), obesity, and alcohol were risk factors of outcomes in men at medium term and long term.
These findings suggest that it is crucial to establish the individual scheme of therapy for every patient by different risk factors of stroke, strengthen the rehabilitation of stroke, and carry on the health education early for the secondary prevention of stroke in patients with diabetes mellitus (DM).
Sex differences; Acute ischemic stroke; Diabetes mellitus; Outcome; Risk factors
Male breast carcinoma (MBC) is rarely encountered in clinical practice. Due to its paucity, our knowledge of MBC only rely on small or single-institutional studies and sporadic cases. The current guidelines for MBC are extrapolated from its female counterparts Rudlowski (Breast Care (Basel) 3(3):183–189, 2008). Nowadays, MBC is actively studied and viewed as a potentially different entity on the aspects of etiology, biological behavior and prognosis. Thus, special treatment strategy guidelines should be established for MBC. Additionally, advance in the systemic chemotherapy and hormonal therapy also contribute to the local control. The indication of radiotherapy need to be clarified and over-treatment should be avoided. Here we present two cases of MBC in which radiotherapy help to sustain a satisfactory disease free survival. Our cases will provide valuable experience for identifying the role of radiotherapy in MBC.
Male breast carcinoma; Radiotherapy
Mitochondria, vital organelles existing
in almost all eukaryotic cells, play a crucial role in energy metabolism
and apoptosis of aerobic organisms. In this work, we report two new
flavone-based fluorescent probes, MC-Mito1 and MC-Mito2, for monitoring mitochondria in living cells. These
two probes exhibit remarkably low toxicity, good cell permeability,
and high specificity; these probes complement the existing library
of mitochondrial imaging agents. The new dyes give nearly no background
fluorescence, and their application does not require tedious postwashing
after cell staining. The appreciable tolerance of MC-Mito2 encourages a broader range of biological applications for understanding
the cell degeneration and apoptosis mechanism.
imaging agents; fluorescence; mitochondria; flavone; biocompatibility; wash-free
Sheep is an important livestock in the world for meat, dairy and wool production. The third version of sheep reference genome has been recently assembled, but sheep DNA methylome has not been profiled yet. In this study, we report the comprehensive sheep methylome with 94.38% cytosine coverage at single base resolution by sequencing DNA samples from Longissimus dorsi of dizygotic Sunit sheep, which were bred in different habitats. We also compared methylomes between the twin sheep. DNA methylation status at genome-scale differentially methylated regions (DMRs), functional genomic regions and 248 DMR-containing genes were identified between the twin sheep. Gene ontology (GO) and KEGG annotations of these genes were performed to discover computationally predicted function. Lipid metabolism, sexual maturity and tumor-associated categories were observed to significantly enrich DMR-containing genes. These findings could be used to illustrate the relationship between phenotypic variations and gene methylation patterns.
The central mechanisms by which interleukin-1 beta (IL-1β) and angiotension II receptor 1 (AT1-R) contribute to sympathoexcitation in heart failure (HF) are unclear. In this study, we determined whether an interaction between IL-1β and AT1-R in the paraventricular nucleus (PVN) contributes to progression of HF. Rats were implanted with bilateral PVN cannulae and subjected to coronary artery ligation or sham surgery (Sham). Subsequently, animals were treated for 4 weeks through PVN infusion with either vehicle, losartan (LOS, 200 μg/day), IL-1β (IL, 1 μg/day), or IL-1β along with LOS (LOS+IL). HF rats had higher levels of corticotropin-releasing hormone (CRH), norepinephrine (NE), and glutamate (Glu); lower levels of gamma-aminobutyric acid (GABA); and more positive fra-like activity in PVN when compared with Sham rats. HF rats also had higher levels of NE, epinephrine (EPI), and IL-1β in plasma. PVN infusion of LOS attenuated the decreases in GABA and the increases in CRH, NE, and Glu in the PVN of HF rats. IL-1β could further increase the expression of CRH, NE, Glu, EPI, and IL-1β and decrease GABA expression. Treatment with IL-1β along with LOS could eliminate the effects of IL-1β. These findings suggest that an interaction between AT1-R and IL-1β in the PVN contributes to progression in HF.
Increased nicotinamide phosphoribosyltransferase (NAMPT) transcription is mechanistically linked to ventilator-induced inflammatory lung injury (VILI), with VILI severity attenuated by reduced NAMPT bioavailability. The molecular mechanisms of NAMPT promoter regulation in response to excessive mechanical stress remain poorly understood. The objective of this study was to define the contribution of specific transcription factors, acute respiratory distress syndrome (ARDS)-associated single nucleotide polymorphisms (SNPs), and promoter demethylation to NAMPT transcriptional regulation in response to mechanical stress. In vivo NAMPT protein expression levels were examined in mice exposed to high tidal volume mechanical ventilation. In vitro NAMPT expression levels were examined in human pulmonary artery endothelial cells exposed to 5 or 18% cyclic stretch (CS), with NAMPT promoter activity assessed using NAMPT promoter luciferase reporter constructs with a series of nested deletions. In vitro NAMPT transcriptional regulation was further characterized by measuring luciferase activity, DNA demethylation, and chromatin immunoprecipitation. VILI-challenged mice exhibited significantly increased NAMPT expression in bronchoalveolar lavage leukocytes and in lung endothelium. A mechanical stress–inducible region (MSIR) was identified in the NAMPT promoter from −2,428 to −2,128 bp. This MSIR regulates NAMPT promoter activity, mRNA expression, and signal transducer and activator of transcription 5 (STAT5) binding, which is significantly increased by 18% CS. In addition, NAMPT promoter activity was increased by pharmacologic promoter demethylation and inhibited by STAT5 silencing. ARDS-associated NAMPT promoter SNPs rs59744560 (−948G/T) and rs7789066 (−2,422A/G) each significantly elevated NAMPT promoter activity in response to 18% CS in a STAT5-dependent manner. Our results show that NAMPT is a key novel ARDS therapeutic target and candidate gene with genetic/epigenetic transcriptional regulation in response to excessive mechanical stress.
acute respiratory distress syndrome; cyclic stretch; nicotinamide phosphoribosyltransferase; B cell colony-enhancing factor; signal transducer and activator of transcription 5
Activity-dependent stimuli induced a calcineurin-mediated dephosphorylation of the transcriptional factor MEF2A at serine408 and promoted a switch from SUMOylation to acetylation at lysine403 which led to MEF2A transcriptional activation. We previously identified SENP2 is the de-SUMOylation enzyme for MEF2A and promotes MEF2A-dependent transcription. We report here a requirement for APCCdh1-SENP2-MEF2A axis in the regulation of MEF2A transcriptional activation. APCCdh1 interacts with and targets SENP2 for ubiquitination and destruction in the cytoplasm by recognizing a conserved canonical D-box motif in SENP2. Moreover, Cdh1 regulates the transcriptional activity of MEF2A in a SENP2 dependent manner. Activity-dependent stimuli prevented APCCdh1-induced SENP2 ubiquitination, promoted SENP2 nuclear accumulations, and caused MEF2A de-SUMOylation and MEF2A acetylation, leading to MEF2A transcriptional activation. Thus, our findings defined a post-transcriptional mechanism underlying activity-dependent stimuli-induced MEF2A transcriptional activation.
activity-dependent stimuli; APC-Cdh1; MEF2A; SENP2
Quantitation of lipid storage, desaturation, and oxidation in live C. elegans has been a long-standing obstacle. By hyperspectral stimulated Raman scattering imaging and multivariate analysis in fingerprint vibration region, we present a platform that allows quantitative mapping of fat distribution, degree of fat unsaturation, lipid oxidation, and cholesterol storage in vivo in whole worm. Our results reveal for the first time that lysosome related organelles in intestinal cells are sites for storage of cholesterol in C. elegans.
Cell metabolism; C. elegans; Lipids; Stimulated Raman scattering imaging; Vibrational spectroscopy
The one-step synthesis of oleic acid-coated TiO2 nanoparticles with visible light-driven photocatalytic activity was reported by this manuscript, using oleic acid-ethanol as crucial starting materials. The photocatalytic degradation of nitrogen monoxide (deNOx) in the gas phase was investigated in a continuous reactor using a series of TiO2 semiconductors, prepared from oleic acid- or acetic acid-ethanol solution. The surface modification on TiO2 by organic fatty acid, oleic acid, could reinvest TiO2 photocatalyst with the excellent visible light response. The deNOx ability is almost as high as 30 % destruction in the visible light region (λ > 510 nm) which is similar to the nitrogen-doped TiO2. Meanwhile, acetic acid, a monobasic acid, has a weaker ability on visible light modification of TiO2.
Electronic supplementary material
The online version of this article (doi:10.1186/s11671-015-1133-7) contains supplementary material, which is available to authorized users.
TiO2; Visible light; Photocatalysis; Surface modification; Oleic acid
An approach based on immunomagnetic separation (IMS) and SYBR Green I real-time PCR (real-time PCR) with species-specific primers and melting curve analysis was proposed as a rapid and effective method for detecting Alicyclobacillus spp. in fruit juices. Specific primers targeting the 16S rDNA sequences of Alicyclobacillus spp. were designed and then confirmed by the amplification of DNA extracted from standard strains and isolates. Spiked samples containing known amounts of target bacteria were used to obtain standard curves; the correlation coefficient was greater than 0.986 and the real-time PCR amplification efficiencies were 98.9%- 101.8%. The detection limit of the testing system was 2.8×101 CFU/mL. The coefficient of variation for intra-assay and inter-assay variability were all within the acceptable limit of 5%. Besides, the performance of the IMS-real-time PCR assay was further investigated by detecting naturally contaminated kiwi fruit juice; the sensitivity, specificity and accuracy were 91.7%, 95.9% and 95.3%, respectively. The established IMS-real-time PCR procedure provides a new method for identification and quantitative detection of Alicyclobacillus spp. in fruit juice.
Purpose: A meta-analysis was undertaken to examine the correlation between ankylosing spondylitis (AS) progression and serum levels of pro-inflammatory cytokines, Interleukin-6 (IL-6) and Interleukin-17 (IL-17) in AS patients. Methods: PubMed, EBSCO, Cochrane Library database, Ovid, Springer link, WANFANG, China national knowledge infrastructure (CNKI) and VIP databases(last updated search in October, 2014) were exhaustively searched for published case-control studies using keywords related to IL-6, IL-17 and AS. The search results were screened using stringent inclusion and exclusion criteria, and the data from selected high-quality studies was analyzed with Comprehensive Meta-analysis 2.0 software. Results: Thirteen case-control studies were selected for this meta-analysis and contained a pooled total of 514 AS patients and 358 healthy controls. Our main result revealed strikingly higher serum levels of IL-6 and IL-17 in AS patients, compared to healthy controls (IL-6: SMD = 2.51, 95% CI = 1.33~3.70, P = 0.01; IL-17: SMD = 3.05, 95% CI = 2.09~4.02, P < 0.001). Ethnicity-based subgroup analysis showed a statistically correlation of high IL-6 and IL-17 serum levels with AS both in Asian (IL-6: SMD = 3.15, 95% CI = 0.75~5.55, P < 0.001; IL-17: SMD = 3.30, 95% CI = 1.93~4.66, P < 0.001) and Caucasian populations (IL-6: SMD = 1.34, 95% CI = 0.33~2.35, P = 0.009; IL-17: SMD = 2.52, 95% CI = 1.06~3.98, P = 0.001). Conclusion: Meta-analysis of pooled data from thirteen high-quality studies revealed a strong correlation between elevated IL-6 and IL-17 serum levels and the development of AS. Therefore, IL-6 and IL-17 could be used as markers for diagnosis and assessment of treatment outcomes in AS patients.
Ankylosing spondylitis; interleukin-6; interleukin-17; rheumatism; spondyloarthropathies; meta-analysis
Objective of this study is to analyze preoperative concerns of patients suffering from spinal degenerative disease in a Chinese population. A total of 94 patients with spinal degenerative disease were included, and they were divided into four groups: male and female group, older (≥60 year-old) and younger group (<60 year-old). Questionnaire was designed through patients counseling, preliminary formulation, pilot test and final revision. Each patient was required to select three items of greatest concern. “Attention rate” (AR) was defined as ratio of selected times of one item upon case number within the group. AR of three most concerned items between male and female group, as well as older and younger group were compared and analyzed. All participants selected “recurrence of symptoms following operation” (41/94), “clinical outcome” (35/94) and “postoperative rehabilitation and daily activity” (30/94) as their three top items of concern. Both male and female groups selected “recurrence of symptoms following operation” (22/47, 19/47), “clinical outcome” (21/47, 14/47), “postoperative rehabilitation and daily activity” (15/47, 15/47) and “limb paralysis” (13/47, 14/47) as their most concerned items, revealing no statistical difference (P>0.05). Older group chose “clinical outcome” (17/46) as their most concerned item, followed by “limb paralysis” (14/46), “postoperative rehabilitation and daily activity” (14/46) and “recurrence of symptoms following operation” (12/46). Younger group chose “recurrence of symptoms following operation” (29/48), “clinical outcome” (18/48) and “postoperative rehabilitation and daily activity” (16/48) as their three top concerned items. AR of “recurrence of symptoms following operation” between older and younger group demonstrated statistical difference (P<0.001), while AR of remaining items of greatest concern between both groups were not statistically different (P>0.05). For Chinese patients, high level concerns are associated with surgical outcome and most of them reveal no gender-associated or age-associated difference. Assessing preoperative concerns empowers better preoperative counseling between surgeons and patients and more informed decision for patients.
Concern; preoperation; spine; degenerative disease
Asiaticoside is one of the main functional components of the natural plant Centella asiatica urban. Studies have reported it has several functions such as anti-depression and nerve cell protection. Asiaticoside can reduce the cerebral infarct size in acute focal cerebral ischemia in a mouse model and asiatic acid glycosides can significantly improve neurobehavioral scores. Currently, there is a lack of understanding of asiaticoside in regard to its neural protective mechanism in cerebral ischemia. This study aimed to solve this problem by using an ischemia-hypoxia cell model in vitro.
An in vitro ischemia hypoxia cell model was successfully established by primary cultured newborn rat cortical neurons. After being treated by asiaticoside for 24 h, cell survival rate, lactate dehydrogenase release quantity, and B-cell lymphoma gene-2 (BCL-2), Bax, and caspase-3 protein expressions was detected.
After 10 nmol/L or 100 nmol/L of asiaticoside were given to the cells, cell survival rate increased significantly and presented concentration dependence. Asiaticoside can reduce lactate dehydrogenase release. Lactate dehydrogenase release in model cells is gradually reduced with the increase of asiaticoside concentration. The lactate dehydrogenase release in asiaticoside 10 nmol/L group, asiaticoside 100 nmol/L group and ischemia hypoxia group were 26.75±1.05, 22.36±2.87 and 52.35±5.46%, respectively (p<0.05). It was also found that asiaticoside could modulate the expression of apoptotic factors, including bcl-2, Bax, and caspase-3.
Asiaticoside helps to protect in vitro ischemia hypoxia neurons. This nerve cell protection may be mediated by the BCL-2 protein.
Abstracting and Indexing as Topic; Dilazep; Hypoxia-Ischemia, Brain
A hydroxybenzohydrazide-based Schiff base ligand was conveniently synthesized. Upon addition of Zn2+ cation, the ligand exhibited a high tendency to form a binuclear structure with a 2:2 ligand-to-zinc ratio, which was accompanied with a large fluorescence turn-on (λem = 507 nm, ϕfl ≈ 0.28). The reactivity of zinc complex was examined by using different phosphate anions, which reveals a higher response to acid pyrophosphate anion. Detailed spectroscopic studies revealed that the pyrophosphate response is based on the ligand displacement mechanism.
Activation of cancer stem cell signaling is central to acquired resistance to therapy in esophageal cancer (EC). ABT-263, a potent Bcl-2 family inhibitor, is active against many tumor types. However, effect of ABT-263 on EC cells and their resistant counterparts are unknown. Here we report that ABT-263 inhibited cell proliferation and induced apoptosis in human EC cells and their chemo-resistant counterparts. The combination of ABT-263 with 5-FU had synergistic lethal effects and amplified apoptosis that does not depend fully on its inhibition of BCL-2 family proteins in EC cells. To further explore the novel mechanisms of ABT-263, proteomic array (RPPAs) were performed and gene set enriched analysis demonstrated that ABT-263 suppresses the expression of many oncogenes including genes that govern stemness pathways. Immunoblotting and immunofluorescence further confirmed reduction in protein expression and transcription in Wnt/β-catenin and YAP/SOX9 axes. Furthermore, ABT263 strongly suppresses cancer stem cell properties in EC cells and the combination of ABT-263 and 5-FU significantly reduced tumor growth in vivo and suppresses the expression of stemness genes. Thus, our findings demonstrated a novel mechanism of ABT-263 antitumor effect in EC and indicating that combination of ABT-263 with cytotoxic drugs is worthy of pursuit in patients with EC.
esophageal cancer; stemness pathways; cancer stem cells; ABT-263; 5-fluorouracil
The aim of the present study was to investigate the expression, localization and role of centromere-associated protein E (Cenp-E) in hepatoma cells. The Cenp-E mRNA expression levels in the HepG-2 human hepatocellular carcinoma and LO2 normal hepatic cell lines following treatment with nocodazole were detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Furthermore, the localization and expression of Cenp-E protein in the two cell types was visualized using indirect immunofluorescence. RT-qPCR was also performed to detect the Cenp-E mRNA expression levels in LO2 cells before and after RNA interference. Additional evaluation of the function of interfered cells was performed using indirect immunofluorescence. The results of RT-qPCR demonstrated that the protein expression levels of Cenp-E in the two cell lines prior to treatment with nocodazole were not significantly different (P>0.05). However, the upregulation of Cenp-E expression levels in the LO2 cells was significantly higher compared with that in the HepG-2 cells during cell division (P<0.05). Indirect immunofluorescence analysis indicated that the Cenp-E protein was predominantly located in the nucleus, and that Cenp-E protein expression in nuclei with abnormal mitosis was markedly lower compared with that in nuclei exhibiting normal mitosis. Indirect immunofluorescence also determined that the ratio of dyskaryosis was significantly higher in cells that had undergone Cenp-E interference compared with normal cells. Thus, the present study indicated that the low expression of Cenp-E mRNA may be an important reason for numerical chromosomal abnormalities in human hepatoma cells.
centromere-associated protein E; chromosome numerical abnormality; hepatoma; spindle checkpoint protein
Objective: To conduct a meta-analysis of all association studies on two of the collagen 1 alpha 1 (COL1A1) gene polymorphisms, the -1997G/T (rs1107946) and the -1663indelT (rs2412298) polymorphisms and osteoporosis/BMD and fracture. Methods: PubMed/Medline and Web of Knowledge were searched for relevant association studies published in English. Pooled OR and its corresponding 95% CI or pooled MD and its corresponding 95% CI was calculated with the Cochrane Review Manager (Revman, version 5.2) using a random-effect or a fixed effect model. Results: No significant association between the -1997G/T polymorphism and Lumbar Spine (LS) and Femoral Neck (FN) BMD except for the Caucasian subpopulation wherein subjects with the T allele of the -1997G/T polymorphism was associated with significantly higher LS BMD. Our analysis did reveal that women, especially postmenopausal or perimenopausal women with the GG genotype, had significantly higher Total Hip (TH) BMD than those with the GT. Additionally, our meta-analysis did not show significant association between the -1997G/T polymorphism and risk of fracture, between the -1663indelT polymorphism and LS BMD in postmenopausal or perimenopausal women, or between the -1663indelT polymorphism and the risk of fracture. Conclusions: Our results suggested the possibility of the COL1A1 -1997G/T and the -1663indelT polymorphisms individually playing very little role in osteoporosis and fracture, although more studies are needed especially for the analysis of association between these two polymorphisms and fracture. Haplotype studies may become one important future direction of study to further elucidate whether and how various COL1A1 polymorphisms affect bone health, osteoporosis and fracture.
COL1A1; -1997G/T; -1663indelT; polymorphism; osteoporosis; bone mineral density; fracture
Trastuzumab resistance in HER-2 positive breast cancer cells is closely related to overexpression of both epidermal growth factor receptor (EGFR) and human epidermal receptor (HER-2). SHP-1 has been demonstrated to downregulate tyrosine kinase activity including EGFR via its phosphatase function, but its effect on HER-2 activity is still unknown. Here, we examined the hypothesis that SHP-1 enhances the anticancer efficacy of trastuzumab in EGFR/HER-2 positive breast cancer cells through combining dual inhibition of EGFR and HER-2.
Trastuzumab-resistant breast cancer SKBr-3 cells were generated by long-term in vitro culture of SKBr-3cells in the presence of trastuzumab. The SHP-1 was ectopically expressed by stable transfection. The activity and expression of EGFR, HER-2, and downstream signaling pathways were tested by Western blot. Cell viability was examined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and apoptosis was examined by flow cytometry. The binding between SHP-1 and EGFR/HER-2 was evaluated by immunoprecipitation assay and bimolecular fluorescence complementation. The effects of SHP-1 on tumorigenicity and trastuzumab sensitivity were confirmed via in vivo xenograft model.
Trastuzumab-resistant SKBr-3 cells showed aberrant co-expression of EGFR and HER-2. Introduction of wild-type SHP-1 inhibited cell proliferation, clone formation, and promoted the apoptosis induced by trastuzumab. Meanwhile, SHP-1 overexpression reduced phosphorylation levels of EGFR and HER-2 both in parental and trastuzumab-resistant SKBr-3 cells. In vivo study showed an increased antitumor effect of trastuzumab in SHP-1 overexpressed xenografts. At last, we discovered that SHP-1 can make complexes with both EGFR and HER-2, and both phospho-EGFR and phosphor-HER-2 levels in wild-type SHP-1 immunoprecipitates were less than those in phosphatase-inactive SHP-1 (C453S) immunoprecipitates, indicating that EGFR and HER-2 are potential substrates of SHP-1.
Taken together, we have demonstrated that the SHP-1 is a negative regulatory factor of the tyrosine kinase activity of HER-2 and EGFR through inhibiting phosphorylation. Dual targeting of EGFR and HER-2, by combining trastuzumab with SHP-1 overexpression, may improve response in HER-2 overexpressing breast cancer cells that also express high levels of EGFR.
breast cancer; trastuzumab; EGFR; HER-2; SHP-1; drug resistance
Aldehyde dehydrogenase-2 (ALDH2) has a protective effect on ischemic heart disease. Here, we examined the protective effects of ALDH2 on cardiac fibrosis through modulation of the Wnt/β-catenin signaling pathway in a rat model of myocardial infarction (MI).
Wistar rats were divided into the sham (control), MI (model), and ALDH2 activator (Alda-1) groups. After 10 days of treatment, the left ventricular (LV) remodeling parameters of each animal were evaluated by echocardiography. Myocardial fibrosis was evaluated by Masson’s trichrome staining and Sirius Red staining. Expression levels of collagen types I and III and α-smooth muscle actin (α-SMA) were examined. Finally, the expression and activity of ALDH2 and the levels of several Wnt-related proteins and genes, such as phosphoglycogen synthase kinase (GSK)-3β, GSK-3β, β-catenin, Wnt-1, WNT1-inducible signaling-pathway protein 1, and tumor necrosis factor (TNF)-α, were also analyzed.
After MI, the heart weight/body weight ratio, LV dimension at end diastole, and LV dimension at end systole were decreased, while the LV ejection fraction and LV fractional shortening were increased in the Alda-1 group. Myocardial fibrosis was also reduced in the Alda-1 group, accompanied by decreased expression collagen types I and III and α-SMA. β-Catenin, phosphorylated GSK-3β, and Wnt-1 levels were significantly increased in the model group. Interestingly, this alteration was partly reversed by Alda-1 treatment. Immunohistochemical staining showed that numerous WNT1-inducible signaling-pathway protein 1 (WISP-1)- and TNF-α-positive cells were found in the model group. However, few WISP-1- and TNF-α-positive cells were detected in the Alda-1 group.
The reduction of cardiac fibrosis and the down-regulation of β-catenin, phosphorylated GSK-3β, Wnt-1, and WISP-1 may be mediated by increased ALDH2 activity, leading to reduction of MI-related cardiac fibrosis.
myocardial infarction; aldehyde dehydrogenase; cardiac fibrosis; Wnt signaling
In addition to the well characterized function of chemokines in mediating the homing and accumulation of leukocytes to tissues, some chemokines also exhibit potent antimicrobial activity. Little is known of the potential role of chemokines in bovine mammary gland health and disease. The chemokine CCL28 has previously been shown to play a key role in the homing and accumulation of IgA antibody secreting cells to the lactating murine mammary gland. CCL28 has also been shown to act as an antimicrobial peptide with activity demonstrated against a wide range of pathogens including bacteria, fungi and protozoans. Here we describe the cloning and function of bovine CCL28 and document the concentration of this chemokine in bovine milk. Bovine CCL28 was shown to mediate cellular chemotaxis via the CCR10 chemokine receptor and exhibited antimicrobial activity against a variety of bovine mastitis causing organisms. The concentration of bovine CCL28 in milk was found to be highly correlated with the lactation cycle. Highest concentrations of CCL28 were observed soon after parturition, with levels decreasing over time. These results suggest a potential role for CCL28 in the prevention/resolution of bovine mastitis.
RNA interference (RNAi) has been successfully used as a promising method to inhibit the replication of different viruses, including human immunodeficiency virus (HIV). Gene mutation is a hurdle for the anti-HIV by RNAi. Although prone to mutation, some genes are conserved and limited in functionally important regions. The gag gene is conserved in different subtypes and plays an important role in the assembly of HIV viral particle. Here, we identified subtypes and conserved sequences within forty-four gag genes from the epidemic strains among men who have sex with men. Three subtypes of gag gene, including CRF01_AE (47.7 %), CRF07_BC (40.9 %) and B (11.4 %) were analyzed by online blast. We designed five small hairpin RNAs (shRNAs) based on the conserved sequences. The gag–EGFP fusion transcript reporter system was used to select the most efficient shRNA. Among the five candidate shRNAs, gag-shRNA-3 represented a broad-spectrum inhibition against all chosen targets. This broad-spectrum shRNA diminished the titer of subtypes B and C of HIV-1 for a hundred orders of magnitude. The gag-shRNA-3 described here is a potential therapeutic agent in the HIV-1 gene therapy.
HIV-1; MSM; gag; shRNA