Although previous studies have shown that γ-secretase inhibitors significantly suppress tumor growth via anti-angiogenesis, the mechanism involved in the regulation of tumor angiogenesis by γ-secretase inhibitors has not been clearly understood. The objective of this study was to investigate the regulation of vascular endothelial growth factor receptor (VEGFR) and endothelial nitric oxide synthase (eNOS) by a γ-secretase inhibitor in the H5V mouse microvascular endothelial cell line. H5V cells were cultured with different concentrations of the γ-secretase inhibitor DAPT for 48 h and with 100 μmol/l DAPT at different incubation times. Protein and mRNA expression of VEGFR-1, VEGFR-2, VEGFR-3 and eNOS was measured by Western blotting and real-time PCR, respectively. The VEGFR-2 kinase inhibitor was used to assess the role of VEGFR-2 in eNOS regulation. We found that the γ-secretase inhibitor DAPT increased protein and mRNA expression of VEGFR-2 and eNOS, but decreased VEGFR-1 expression and had no significant effect on VEGFR-3. Up-regulation of eNOS was blocked by the VEGFR-2 kinase inhibitor. In conclusion, the γ-secretase inhibitor enhances VEGFR-2 and eNOS expression, and the up-regulation of eNOS is dependent on an increase in VEGFR-2. Thus, we suggest that administration of the γ-secretase inhibitor be combined with disruption of eNOS or interruption of VEGF signaling, which may improve the anti-angiogenic efficacy in tumor treatments.
angiogenesis; vascular endothelial growth factor receptor; endothelial nitric oxide synthase
Transgene spreading is a major concern in cultivating genetically modified (GM) corn. Cross-pollination may cause the spread of transgenes from GM cornfields to conventional fields. Occasionally, seed lot contamination, volunteers, mixing during sowing, harvest, and trade can also lead to transgene escape. Obviously, new biological confinement technologies are highly desired to mitigate transgene spreading in addition to physical separation and isolation methods. In this study, we report the development of a built-in containment method to mitigate transgene spreading in corn. In this method, an RNAi cassette for suppressing the expression of the nicosulfuron detoxifying enzyme CYP81A9 and an expression cassette for the glyphosate tolerant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene G10 were constructed and transformed into corn via Agrobacterium-mediated transformation. The GM corn plants that were generated were found to be sensitive to nicosulfuron but resistant to glyphosate, which is exactly the opposite of conventional corn. Field tests demonstrated that GM corn plants with silenced CYP81A9 could be killed by applying nicosulfuron at 40 g/ha, which is the recommended dose for weed control in cornfields. This study suggests that this built-in containment method for controlling the spread of corn transgenes is effective and easy to implement.
Differential co-expression analysis (DCEA) has emerged in recent years as a novel, systematic investigation into gene expression data. While most DCEA studies or tools focus on the co-expression relationships among genes, some are developing a potentially more promising research domain, differential regulation analysis (DRA). In our previously proposed R package DCGL v1.0, we provided functions to facilitate basic differential co-expression analyses; however, the output from DCGL v1.0 could not be translated into differential regulation mechanisms in a straightforward manner.
To advance from DCEA to DRA, we upgraded the DCGL package from v1.0 to v2.0. A new module named “Differential Regulation Analysis” (DRA) was designed, which consists of three major functions: DRsort, DRplot, and DRrank. DRsort selects differentially regulated genes (DRGs) and differentially regulated links (DRLs) according to the transcription factor (TF)-to-target information. DRrank prioritizes the TFs in terms of their potential relevance to the phenotype of interest. DRplot graphically visualizes differentially co-expressed links (DCLs) and/or TF-to-target links in a network context. In addition to these new modules, we streamlined the codes from v1.0. The evaluation results proved that our differential regulation analysis is able to capture the regulators relevant to the biological subject.
With ample functions to facilitate differential regulation analysis, DCGL v2.0 was upgraded from a DCEA tool to a DRA tool, which may unveil the underlying differential regulation from the observed differential co-expression. DCGL v2.0 can be applied to a wide range of gene expression data in order to systematically identify novel regulators that have not yet been documented as critical.
DCGL v2.0 package is available at http://cran.r-project.org/web/packages/DCGL/index.html or at our project home page http://lifecenter.sgst.cn/main/en/dcgl.jsp.
Functional recovery is usually poor following peripheral nerve injury when reinnervation is delayed. Early innervation by sensory nerve has been indicated to prevent atrophy of the denervated muscle. It is hypothesized that early protection with sensory axons is adequate to improve functional recovery of skeletal muscle following prolonged denervation of mixed nerve injury. In this study, four groups of rats received surgical denervation of the tibial nerve. The proximal and distal stumps of the tibial nerve were ligated in all animals except for those in the immediate repair group. The experimental groups underwent denervation with nerve protection of peroneal nerve (mixed protection) or sural nerve (sensory protection). The experimental and unprotected groups had a stage II surgery in which the trimmed proximal and distal tibial nerve stumps were sutured together. After 3 months of recovery, electrophysiological, histological and morphometric parameters were assessed. It was detected that the significant muscle atrophy and a good preserved structure of the muscle were observed in the unprotected and protective experimental groups, respectively. Significantly fewer numbers of regenerated myelinated axons were observed in the sensory-protected group. Enhanced recovery in the mixed protection group was indicated by the results of the muscle contraction force tests, regenerated myelinated fiber, and the results of the histological analysis. Our results suggest that early axons protection by mixed nerve may complement sensory axons which are required for promoting functional recovery of the denervated muscle natively innervated by mixed nerve.
Many tumors are stiffer than their surrounding tissue. This increase in stiffness has been attributed, in part, to a Rho-dependent elevation of myosin II light chain phosphorylation. To characterize this mechanism further, we studied myosin light chain kinase (MLCK), the main enzyme that phosphorylates myosin II light chains. We anticipated that increases in MLCK expression and activity would contribute to the increased stiffness of cancer cells. However, we find that MLCK mRNA and protein levels are substantially less in cancer cells and tissues than in normal cells. Consistent with this observation, cancer cells contract 3D collagen matrices much more slowly than normal cells. Interestingly, inhibiting MLCK or Rho kinase did not affect the 3D gel contractions while blebbistatin partially and cytochalasin D maximally inhibited contractions. Live cell imaging of cells in collagen gels showed that cytochalasin D inhibited filopodia-like projections that formed between cells while a MLCK inhibitor had no effect on these projections. These data suggest that myosin II phosphorylation is dispensable in regulating the mechanical properties of tumors.
The development of suitable methods to deliver peptides specifically to the endoplasmic reticulum (ER) can provide some potential therapeutic applications of such peptides. Ankylosing spondylitis (AS) is strongly associated with the expression of human leukocytic antigen-B27 (HLA-B27). HLA-B27 heavy chain (HC) has a propensity to fold slowly resulting in the accumulation of misfolded HLA-B27 HC in the ER, triggering the unfolded protein response, and forming a homodimer, (B27-HC)2. Natural killer cells and T-helper 17 cells are then activated, contributing to the major pathogenic potentials of AS. The HLA-B27 HC is thus an important target, and delivery of an HLA-B27-binding peptide to the ER capable of promoting HLA-B27 HC folding is a potential mechanism for AS therapy. Here, we demonstrate that a His6-ubiquitin-tagged Tat-derived peptide (THU) can deliver an HLA-B27-binding peptide to the ER promoting HLA-B27 HC folding. The THU-HLA-B27-binding peptide fusion protein crossed the cell membrane to the cytosol through the Tat-derived peptide. The HLA-B27-binding peptide was specifically cleaved from THU by cytosolic ubiquitin C-terminal hydrolases and subsequently transported into the ER by the transporter associated with antigen processing. This approach has potential application in the development of peptide therapy for AS.
Telehealthcare has been used to provide healthcare service, and information technology infrastructure appears to be essential while providing telehealthcare service. Insufficiencies have been identified, such as lack of integration, need of accommodation of diverse biometric sensors, and accessing diverse networks as different houses have varying facilities, which challenge the promotion of telehealthcare. This study designs an information technology framework to strengthen telehealthcare delivery.
Materials and Methods:
The proposed framework consists of a system architecture design and a network transmission design. The aim of the framework is to integrate data from existing information systems, to adopt medical informatics standards, to integrate diverse biometric sensors, and to provide different data transmission networks to support a patient's house network despite the facilities. The proposed framework has been evaluated with a case study of two telehealthcare programs, with and without the adoption of the framework.
The proposed framework facilitates the functionality of the program and enables steady patient enrollments. The overall patient participations are increased, and the patient outcomes appear positive. The attitudes toward the service and self-improvement also are positive.
The findings of this study add up to the construction of a telehealthcare system. Implementing the proposed framework further assists the functionality of the service and enhances the availability of the service and patient acceptances.
telehealthcare; information technology framework; integrated information system
Human serum albumin (HSA) is widely utilized for medical purposes and biochemical research. Transgenic rice has proved to be an attractive bioreactor for mass production of recombinant HSA (rHSA). However, transgene spread is a major environmental and food safety concern for transgenic rice expressing proteins of medical value. This study aimed to develop a selectively terminable transgenic rice line expressing HSA in rice seeds, and a simple process for recovery and purification of rHSA for economical manufacture. An HSA expression cassette was inserted into a T-DNA vector encoding an RNA interference (RNAi) cassette suppressing the CYP81A6 gene. This gene detoxifies the herbicide bentazon and is linked to the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) cassette which confers glyphosate tolerance. ANX Sepharose Fast Flow (ANX FF) anion exchange chromatography coupled with Butyl Sepharose High Performance (Butyl HP) hydrophobic interaction chromatography was used to purify rHSA. A transgenic rice line, HSA-84, was obtained with stable expression of rHSA of up to 0.72% of the total dry weight of the dehusked rice seeds. This line also demonstrated high sensitivity to bentazon, and thus could be killed selectively by a spray of bentazon. A two-step chromatography purification scheme was established to purify the rHSA from rice seeds to a purity of 99% with a recovery of 62.4%. Results from mass spectrometry and N-terminus sequencing suggested that the purified rHSA was identical to natural plasma-derived HSA. This study provides an alternative strategy for large-scale production of HSA with a built-in transgene safety control mechanism.
Recombinant human serum albumin (rHSA); Selectively terminable transgenic rice; Purification
gp120 is a substrate for protein engineering both for HIV immunogen design and as a bait for isolating anti-HIV antibodies from patient samples. In this work we describe the display of a stripped core gp120 on the yeast cell surface. Validation against a panel of neutralizing antibodies confirms that yeast-displayed gp120 presents the CD4 binding site in the correct conformation. We map the epitope of the broadly neutralizing anti-gp120 antibody VRC01 using both a random mutagenesis library and a defined mutant panel, and find the resultant epitope maps are consistent with one another and with the crystallographically identified contact residues. Mapping the VRC01-competitive antibodies b12 and b13 reveals energetic differences in their epitopes that are not obvious from existing crystal structures. These data suggest mutation sets that abrogate binding to broadly neutralizing antibodies with greater specificity than the canonical mutation D368R, useful in rapidly assessing the nature of a vaccine response.
New onset diabetes after transplantation (NODAT) occurs less frequently in living donor liver transplant (LDLT) recipients than in deceased donor liver transplant (DDLT) recipients. The aim of this study was to compare the incidence and predictive factors for NODAT in LDLT versus DDLT recipients. The Organ Procurement and Transplant Network/United Network for Organ Sharing database was reviewed from 2004 to 2010, and 902 LDLT and 19,582 DDLT nondiabetic recipients were included. The overall incidence of NODAT was 12.2% at 1 year after liver transplantation. At 1, 3, and 5 years after transplant, the incidence of NODAT in LDLT recipients was 7.4, 2.1, and 2.6%, respectively, compared to 12.5, 3.4, and 1.9%, respectively, in DDLT recipients. LDLT recipients have a lower risk of NODAT compared to DDLT recipients (hazard ratio = 0.63 (0.52–0.75), P < 0.001). Predictors for NODAT in LDLT recipients were hepatitis C (HCV) and treated acute cellular rejection (ACR). Risk factors in DDLT recipients were recipient male gender, recipient age, body mass index, donor age, donor diabetes, HCV, and treated ACR. LDLT recipients have a lower incidence and fewer risk factors for NODAT compared to DDLT recipients. Early identification of risk factors will assist timely clinical interventions to prevent NODAT complications.
Skeletal muscles in the limb and body trunk are composed of heterogeneous myofibers expressing different isoforms of myosin heavy chain (Myh), including type I (slow, Myh7), IIA (intermediate, Myh2), IIX (fast, Myh1) and IIB (very fast, Myh4). While the contraction force and speed of a muscle are known to be determined by the relative abundance of myofibers expressing each Myh isoform, it is unclear how specific combinations of myofiber types are formed and regulated at the cellular and molecular level. We report here that myostatin (Mstn) positively regulates slow but negatively regulates fast Myh isoforms. Mstn was expressed at higher levels in the fast muscle myoblasts and myofibers than in the slow muscle counterparts. Interestingly, Mstn knockout led to a shift of Myh towards faster isoforms, suggesting an inhibitory role of Mstn in fast Myh expression. Consistently, when induced to differentiate, Mstn null myoblasts formed myotubes preferentially expressing fast Myh. Conversely, treatment of myoblasts with a recombinant Mstn protein upregulated Myh7 but downregulated Myh4 gene expression in newly formed myotubes. Importantly, both Mstn antibody and soluble activin type 2B receptor inhibited slow Myh7 and promoted fast Myh4 expression, indicating that myostatin acts through canonical activin receptor to regulate the expression of Myh genes. These results demonstrate a role of myostatin in the specification of myofiber types during myogenic differentiation.
Myostatin (Mstn); satellite cells; myofiber; myosin heavy chain; TGFβ
The Gram-negative bacteria type VI secretion system (T6SS) has been found to play an important role in interbacterial competition, biofilm formation and many other virulence-related processes. The bacteria harboring T6SS inject the effectors into their recipient’s cytoplasm or periplasm to kill them and meanwhile, to avoid inhibiting itself, the cognate immunity proteins were produced to acts as the effector inhibitor. Tae4 (type VI amidase effector 4) and Tai4 (type VI amidase immunity 4) are newly identified T6SS effector-immunity (EI) pairs. We have recently solved the structures of StTae4-Tai4 and EcTae4-Tai4 complexes from the human pathogens Salmonella typhimurium and Enterobacter cloacae, respectively. It is very interesting and important to discover whether there is cross-neutralization between St- and EcTai4 and whether their effector inhibition mechanism is conserved. Here, we determined the crystal structure of StTae4 in complex with EcTai4. The solution conformation study revealed it is a compact heterotetramer that consists of an EcTai4 homodimer binding two StTae4 molecules in solution, different from that in crystal. A remarkable shift can be observed in both the flexible winding loop of StTae4 and protruding loop of EcTai4 and disulfide bonds are formed to stabilize their overall conformations. The in vitro and in vivo interactions studies showed EcTai4 can efficiently rescue the cells from the toxicity of its cognate effectors StTae4, but can not neutralize the toxic activities of the effectors from other families. These findings provide clear structural evidence to support the previous observation of cross-immunity within T6SS families and provide a basis for understanding their important roles in polymicrobial environments.
In the present study, a set of self-designed measurement protocols for the precision of coaxial needle placement (PCNP) was proposed and applied in a computed tomography (CT)-guided transthoracic needle biopsy (TNB) audit of an interventional radiologist to determine if the PCNP was commensurate with the experience of the operator. A total of 102 patients (98 with lung lesions and four with mediastinum lesions) consented to be subjected to CT-guided TNB performed by staff interventional radiologists. The patients were divided into two groups based on appointment date. Group A consisted of the first 51 patients and group B comprised of the latter 51 patients. A set of self-designed measurement protocols for PCNP was proposed, and the PCNP was classified into four grades, from grade 1 (most accurate) to grade 4 (least accurate). PCNPs were independently measured by three staff radiologists who were blind to the grouping. The anatomical features of the lesions were also analyzed between the two groups. A significant difference in the PCNP gained after the first needle placement was identified between the two groups (P=0.003, two-tailed). The number of patients in group B with grade I PCNP (51.0%) was significantly higher than that in group A (21.6%) (P<0.05). The number of patients in group B with grade III PCNP (11.8%) was significantly lower than that in group A (29.4%, P<0.05). The PCNP was observed to be commensurate with the experience of the operator and should be considered as a routine audit index in CT-guided TNB.
biopsy; computed tomography; thorax; audit
Monoclonal antibodies against GD2 ganglioside, such as ch14.18, the human–mouse chimeric antibody, have been shown to be effective for the treatment of neuroblastoma. However, treatment is associated with generalized, relatively opiate-resistant pain. We investigated if a point mutation in ch14.18 antibody (hu14.18K332A) to limit complement-dependent cytotoxicity (CDC) would ameliorate the pain behavior, while preserving antibody-dependent cellular cytotoxicity (ADCC). In vitro, CDC and ADCC were measured using europium-TDA assay. In vivo, allodynia was evaluated by measuring thresholds to von Frey filaments applied to the hindpaws after injection of either ch14.18 or hu14.18K332 into wild type rats or rats with deficient complement factor 6. Other rats were pretreated with complement factor C5a receptor antagonist and tested following ch14.18 injection. The mutation reduces the antibody’s ability to activate complement, while maintaining its ADCC capabilities. Injection of hu14.18K322 (1 or 3 mg/kg) produced faster resolving allodynia than that engendered by ch14.18 (1 mg/kg). Injection of ch14.18 (1 mg/kg) into rats with C6 complement deficiency further reduced antibody-induced allodynia, while pre-treatment with complement factor C5a receptor antagonist completely abolished ch14.18-induced allodynia. These findings showed that mutant hu14.18 K322 elicited less allodynia than ch14.18 and that ch14.18-elicited allodynia is due to activation of the complement cascade: in part, to formation of membrane attack complex, but more importantly to release of complement factor C5a. Development of immunotherapeutic agents with decreased complement-dependent lysis while maintaining cellular cytotoxicity may offer treatment options with reduced adverse side effects, thereby allowing dose escalation of therapeutic antibodies.
Anti-GD2; Pain; C5a complement factor; Membrane attack complex; Neuroblastoma; Neuroblastoma therapy; Complement-dependent cytotoxicity; Antibody-dependent cellular cytotoxicity
To explore the correlation between the retinal nerve fiber layer (RNFL) thickness by using optical coherence tomography (OCT) and by histological measurements in normal adult rats and optic nerve transected rats.
The RNFL thickness of 36 rats was scanned in a circle 3.46mm far from the optic disc by OCT. The two experimental groups were the normal group (n=20 rats) and the optic nerve transected group (n=16 rats). The latter group included 4 groups (n=4/group) surviving for 1 day, 3, 5 and 7 days. Then the RNFL thickness of the same retina area was also measured by NF-200 immunohistochemical staining method. Linear regression was used to analyze the correlation between the data obtained from these two methods.
The RNFL thickness of normal right eyes around optic disc by OCT was 72.35±5.71µm and that of the left eyes was 72.65±5.88µm (P=0.074). The RNFL thickness of the corresponding histological section by immunohistochemistry was 37.54±4.05µm (right eyes) and 37.38±4.23µm (left eyes) (P=0.059). There was a good correlation between the RNFL thickness measured by OCT and that measured by histology (R2=0.8131). After optic nerve transection, the trend of the RNFL thickness was thinner with the prolonged survival time. The correlation of the thickness detected by the above two methods was approximately (R2=0.8265). Value of the RNFL thickness in rats around optic disc measured by OCT was obviously higher than that measured by common histological measurement in normal adult rats and optic nerve transected rats.
The RNFL thickness measured by OCT has a strong correlation with that measured by histological method. Through OCT scanning, we found that the thickness of RNFL gradually becomes thinner in a time-dependent manner.
retinal nerve fiber layer thickness; optical coherence tomography; optic nerve transection; immunohistochemistry staining; relevance analysis; rat
S. erythraea is a Gram-positive filamentous bacterium used for the industrial-scale production of erythromycin A which is of high clinical importance. In this work, we sequenced the whole genome of a high-producing strain (E3) obtained by random mutagenesis and screening from the wild-type strain NRRL23338, and examined time-series expression profiles of both E3 and NRRL23338. Based on the genomic data and transcriptpmic data of these two strains, we carried out comparative analysis of high-producing strain and wild-type strain at both the genomic level and the transcriptomic level.
We observed a large number of genetic variants including 60 insertions, 46 deletions and 584 single nucleotide variations (SNV) in E3 in comparison with NRRL23338, and the analysis of time series transcriptomic data indicated that the genes involved in erythromycin biosynthesis and feeder pathways were significantly up-regulated during the 60 hours time-course. According to our data, BldD, a previously identified ery cluster regulator, did not show any positive correlations with the expression of ery cluster, suggesting the existence of alternative regulation mechanisms of erythromycin synthesis in S. erythraea. Several potential regulators were then proposed by integration analysis of genomic and transcriptomic data.
This is a demonstration of the functional comparative genomics between an industrial S. erythraea strain and the wild-type strain. These findings help to understand the global regulation mechanisms of erythromycin biosynthesis in S. erythraea, providing useful clues for genetic and metabolic engineering in the future.
S. erythraea; Erythromycin biosynthesis; Functional comparative genetics; Regulation mechanism
Transverse myelitis is a rare inflammatory myelopathy characterized by loss of motor and sensory function below the affected level of the spinal cord, and causes neurogenic bowel and bladder. Occasionally, it also causes neuropathic pain with spasticity. Traditional therapies for neuropathic pain are multiple, including multimodal analgesic regimens, antiepileptic or antidepressant medications, opioids, sympathetic blocks, and spinal cord stimulation. Persistent neuropathic pain can cause emotional distress by affecting sleep, work, recreation, and emotional well-being. Here we report the case of a patient suffering from intractable neuropathic pain following acute transverse myelitis that was not relieved by combinations of nonsteroidal anti-inflammatory, anti-epileptic, antidepressant, and opioid medications, or by acupuncture. Implantation of an intrathecal morphine pump controlled the pain successfully without side effects, and enabled the patient to embark on intensive rehabilitation. The patient’s muscle strength has improved significantly and the patient may soon be able to use a walker with minimal assistance.
intrathecal morphine pump; neuropathic pain; rehabilitation; transverse myelitis
The crystal structure of POTRA4–5 has been determined to 1.50 Å resolution with an R factor of 14.7% and an R
free of 18.9%.
In Escherichia coli, the BAM complex is employed to mediate correct folding of the outer membrane (OM) proteins into β-barrels and their insertion into the OM. BamA, which is an essential component of the complex, consists of a C-terminal transmembrane region and five N-terminal polypeptide transport-associated (POTRA) domains. Although deletion studies have shown that each of the POTRA domains plays an important role in the process of BAM complex formation, only POTRA5 is essential for cell viability. Here, the crystal structure of POTRA4–5 has been determined to 1.50 Å resolution with an R factor of 14.7% and an R
free of 18.9%.
BAM complex; POTRA domains; BamA; Escherichia coli
The diagnosis of coccidioidomycosis relies heavily on serologic test results in addition to clinical history, physical examination, and radiographic findings. Use of the enzyme immunoassay (EIA) has increased because it is rapidly performed and does not require referral to a reference laboratory, as do complement fixation and immunodiffusion tests. However, interpretation of immunoglobulin M (IgM) reactivity by EIA in the absence of immunoglobulin G (IgG) reactivity has been problematic. We conducted a retrospective medical record review of all patients with such IgM reactivity at our institution to identify situations where the finding was more likely to be clinically specific for coccidioidal infection. From 1 January 2004 through 31 December 2008, a total of 1,117 patients had positive EIA coccidioidal serology or EIA IgM-only reactivity; of these, 102 patients (9%) had EIA IgM-only reactivity. Among the 102 patients with EIA IgM-only reactivity, 60 were tested to evaluate symptomatic illness, 13 for follow-up of previously abnormal serology, and 29 for screening purposes. Of the 102 patients, 80 (78%) had positive serologic findings by other methods or had positive culture or histology. Fifty-four (90%) of the 60 patients whose serology was performed to evaluate symptomatic illness had coccidioidal infection, whereas 13 (45%) of 29 patients whose serology was performed for screening purposes had coccidioidal infection. Of the 102 patients with isolated IgM reactivity by EIA, 12 later seroconverted to IgG and IgM reactivity. The use of EIA for screening in 29 asymptomatic persons was associated with unconfirmable results in 13 (45%). Although the majority of patients in our study with isolated IgM reactivity by EIA had probable or confirmed coccidioidomycosis, this result must be interpreted with caution for asymptomatic patients.
In order to understand the composition and dynamics of planktonic viruses and their relationship with environmental parameters in natural freshwater, flow cytometry was optimized with filtration/fixation/staining/dilution and then applied to the analysis of samples collected from 9 stations (covering urban, rural, and estuarial areas) along the Haihe River, China, over a one-year period of study. The total viral abundance exhibited an apparent peak in the spring. Spatially, the highest viral abundance was recorded in estuarial areas. The correlation analysis indicated that the bacteria in the Haihe River significantly influenced viral abundance. The relationship between abiotic variables and viral abundance remained the same as with bacterial abundance, indicating that environmental parameters could possibly influence viral abundance in virtue of their bacterial host cells. The influence of environmental factors on viral abundance differed in the three sampling areas, suggesting different drivers of viral abundance in different stretches of the river associated with their utilization and surroundings.
Transcription factors of the FoxO (forkhead box O) family regulate a wide range of cellular physiological processes, including metabolic adaptation and myogenic differentiation. The transcriptional activity of most FoxO members is inhibitory to myogenic differentiation and overexpression of FoxO1 inhibits the development of oxidative type I fibres in vivo. In this study, we found that FoxO6, the last discovered FoxO family member, is expressed ubiquitously in various tissues but with higher expression levels in oxidative tissues, such as brain and oxidative muscles. Both the expression level and promoter activity of FoxO6 were found to be enhanced by PGC-1α (peroxisome-proliferator-activated receptor γ co-activator 1α), thus explained its enriched expression in oxidative tissues. We further demonstrated that FoxO6 represses the expression of PGC-1α via direct binding to an upstream A/T-rich element (AAGATATCAAAACA,−2228–2215) in the PGC-1α promoter. Oxidative low-intensity exercise induced PGC-1α but reduced FoxO6 expression levels in hind leg muscles, and the binding of FoxO6 to PGC-1α promoter was also prevented by exercise. As FoxO6 promoter can be co-activated by PGC-1α and its promoter in turn can be repressed by FoxO6, it suggests that FoxO6 and PGC-1α form a regulatory loop for setting oxidative metabolism level in the skeletal muscle, which can be entrained by exercise.
FoxO6; muscle; oxidative metabolism; PGC-1α; promoter; transcription; CHIP, chromatin immunoprecipitation; DM, differentiation medium; DMEM, Dulbecco’s modified Eagle’s medium; EMSA, electrophoretic mobility-shift assay; FCS, fetal calf serum; FoxO, forkhead box O; Gapdh, glyceraldehydes-3-phosphate dehydrogenase; GC, gastrocnemius; GM, growth medium; HEK-293T, HEK-293 cells expressing the large T-antigen of SV40; HRP, horseradish peroxidase; MBP, maltose-binding protein; MRFs, myogenic regulatory factors; PBST, PBS containing 0.5% Tween 20; PGC-1α, peroxisome-proliferator-activated receptor γ co-activator 1α; PI3k, phosphoinositide 3-kinase; qRT-PCR, quantitative reverse transcription PCR; T2D, type 2 diabetes
The rise of multidrug-resistant (MDR) pathogens causes an increasing challenge to public health. Antimicrobial peptides are considered a possible solution to this problem. HBV core protein (HBc) contains an arginine-rich domain (ARD) at its C-terminus, which consists of 16 arginine residues separated into four clusters (ARD I to IV). In this study, we demonstrated that the peptide containing the full-length ARD I–IV (HBc147-183) has a broad-spectrum antimicrobial activity at micro-molar concentrations, including some MDR and colistin (polymyxin E)-resistant Acinetobacter baumannii. Furthermore, confocal fluorescence microscopy and SYTOX Green uptake assay indicated that this peptide killed Gram-negative and Gram-positive bacteria by membrane permeabilization or DNA binding. In addition, peptide ARD II–IV (HBc153-176) and ARD I–III (HBc147-167) were found to be necessary and sufficient for the activity against P. aeruginosa and K. peumoniae. The antimicrobial activity of HBc ARD peptides can be attenuated by the addition of LPS. HBc ARD peptide was shown to be capable of direct binding to the Lipid A of lipopolysaccharide (LPS) in several in vitro binding assays. Peptide ARD I–IV (HBc147-183) had no detectable cytotoxicity in various tissue culture systems and a mouse animal model. In the mouse model by intraperitoneal (i.p.) inoculation with Staphylococcus aureus, timely treatment by i.p. injection with ARD peptide resulted in 100-fold reduction of bacteria load in blood, liver and spleen, as well as 100% protection of inoculated animals from death. If peptide was injected when bacterial load in the blood reached its peak, the protection rate dropped to 40%. Similar results were observed in K. peumoniae using an IVIS imaging system. The finding of anti-microbial HBc ARD is discussed in the context of commensal gut microbiota, development of intrahepatic anti-viral immunity and establishment of chronic infection with HBV. Our current results suggested that HBc ARD could be a new promising antimicrobial peptide.
Antibiotics-resistant pathogens have been a major problem to our public health. Recently, in our studies of human hepatitis B virus (HBV), we accidentally discovered potent and broad spectrum antimicrobial peptides from HBV core protein (HBc) arginine-rich domain (ARD). The peptides are mainly composed of SPRRR repeats and are effective against both Gram-positive and Gram-negative bacteria, as well as fungi. We found different bactericidal mechanisms of the ARD peptides, which involved LPS binding, DNA binding and membrane permeabilization in various tested bacteria, such as P. aeruginosa, K. pneumoniae, E. coli and S. aureus. We also found that this ARD peptide was effective for colistin-resistant A. baumannii. The peptides exhibited no hemolysis activity to human red blood cells and no cytotoxicity to human hepatoma cells and kidney cells. Furthermore, the ARD peptide was shown to be safe and protective in the animal model. Recently, intestinal flora was found to influence the development of immunity. We discussed here the potential involvement of the antimicrobial activity of HBc ARD in the establishment of HBV chronic infection in the newborns. We proposed here that the HBc ARD peptides could serve as an alternative to the conventional antibiotics in clinical medicine.
Flu vaccination is effective for preventing infection, but coverage levels in the USA remain low—especially among racial/ethnic minorities. This study examines factors associated with flu vaccination in a predominantly Hispanic community in Manhattan, New York.
Households were recruited during the 2006–2007 and 2007–2008 flu seasons. Primary household respondents were interviewed to determine knowledge of flu transmission/treatment and vaccination status and demographic information for all household members.
Vaccination coverage was 47.3% among children <5, 39.3% among 5–17-year-olds, 15.3% among 18–49-year-olds, 31.0% among 50–64-year-olds and 37.1% among adults ≥65 in year 1; and 53.1% among children <5, 43.6% among 5–17-year-olds, 19.5% among 18–49-year-olds, 34.1% among 50–64-year-olds and 34.3% among adults ≥65 in year 2. For children, younger age, having a chronic respiratory condition (eg, asthma), and greater primary respondent knowledge of flu were positively associated with vaccination. Among adults, female gender, older age, higher education, greater primary respondent knowledge of flu, having been born in the USA and having a chronic respiratory condition were positively associated with vaccination. The most common reasons cited for not being vaccinated were the beliefs that flu vaccination was unnecessary or ineffective.
Possible methods for increasing vaccination levels in urban Hispanic communities include improving health literacy, making low-cost vaccination available and encouraging providers to use other office visits as opportunities to mention vaccination to patients.
This study is registered at http://ClinicalTrials.gov (NCT00448981).
Ixabepilone is an epothilone B analogue with activity in a variety of solid malignancies, including prostate cancer. The main dose-limiting toxicity of ixabepilone is myelosuppression when administered by using an every 3-week schedule. Here we evaluate the activity of a weekly ixabepilone in men with metastatic castrate-resistant prostate cancer to minimize hematologic toxicity.
BMS-247550 (ixabepilone) is an epothilone B analogue with activity in taxane-resistant cancer cell lines. Here we report the activity and toxicity of ixabepilone, administered by using a weekly schedule, in men with metastatic castrate-resistant prostate cancer (CRPC).
Patients with metastatic CRPC received ixabepilone at 20 mg/m2 intravenous weekly × 3, in 4-week cycles. This noncomparative study stratified patients to either a chemotherapy naive (CN), prior taxane (Tax) only, or 2 prior cytotoxic (TCx) chemotherapy arm. The primary endpoint was prostate-specific antigen response by using PCWG (Prostate Cancer Working Group) 1 criteria. Secondary endpoints included radiographic response when using RECIST (Response Evaluation Criteria In Solid Tumors).
In total, 124 patients were enrolled, of whom, 109 were eligible (35 CN, 42 Tax, and 32 TCx) for the primary response determination in this study. Prostate-specific antigen responses were seen in 12 (34.3%) of 35, 12 (28.6%) of 42, and 7 (21.9%) of 32 patients with the partial objective response in 5 (22.7%) of 22, 2 (8.0%) of 25, and 0 (0.0%) of 24 patients for the CN, Tax, and TCx arms, respectively. Significant (grade 3/4) neutropenia was seen in 6 (15.4%), 7 (14.6%), and 9 (25.0%); and grade 3/4 sensory neuropathy was seen in 8 (20.5%), 12 (25.0%), and 12 (33.3%) for CN, Tax, and TCx, respectively. Grade 3/4 thrombocytopenia was infrequent and seen in only one patient on the CN and the TCx arm.
Ixabepilone was found to have an acceptable toxicity profile when administered by using a weekly schedule with less myelosuppression compared with prior studies when using the every 3-week schedule. Single-agent activity was observed and met prespecified activity levels for the Tax treated arm.
BMS-247550; Chemotherapy; Epothilone; Microtubule-inhibitor
Primula species are important early spring garden plants with a centre of diversity and speciation in the East Himalaya-Hengduan Mountains in Western China. Studies on population genetics, speciation and phylogeny of Primula have been impeded by a lack of genomic resources. In the present study, we sequenced the transcriptomes of two closely related primrose species, Primula poissonii and Primula wilsonii, using short reads on the Illumina Genome Analyzer platform.
We obtained 55,284 and 55,011 contigs with N50 values of 938 and 1,085 for P. poissonii and P. wilsonii, respectively, and 6,654 pairs of putative orthologs were identified between the two species. Estimations of non-synonymous/synonymous substitution rate ratios for these orthologs indicated that 877 of the pairs may be under positive selection (Ka/Ks > 0.5), and functional enrichment analysis revealed that significant proportions of the orthologs were in the categories DNA repair, stress resistance, which may provide some hints as to how the two closely related Primula species adapted differentially to extreme environments, such as habitats characterized by aridity, high altitude and high levels of ionizing radiation. It was possible for the first time to estimate the divergence time between the radiated species pair, P. poissonii and P. wilsonii; this was found to be approximately 0.90 ± 0.57 Mya, which falls between the Donau and Gunz glaciation in the Middle Pleistocene. Primers based on 54 pairs of orthologous SSR-containing sequences between the two Primula species were designed and verified. About half of these pairs successfully amplified for both species. Of the 959 single copy nuclear genes shared by four model plants (known as APVO genes), 111 single copy nuclear genes were verified as being present in both Primula species and exon-anchored and intron-spanned primers were designed for use.
We characterized the transcriptomes for the two Primula species, and produced an unprecedented amount of genomic resources for these important garden plants. Evolutionary analysis of these two Primula species not only revealed a more precise divergence time, but also provided some novel insights into how differential adaptations occurred in extreme habitats. Furthermore, we developed two sets of genetic markers, single copy nuclear genes and nuclear microsatellites (EST-SSR). Both these sets of markers will facilitate studies on the genetic improvement, population genetics and phylogenetics of this rapidly adapting taxon.
Adaptive radiation; East Himalaya-Hengduan Mountains; EST-SSR; Single copy nuclear gene