Phosphorylated AKT (p-AKT), constitutive activation of AKT, is a potentially interesting prognostic marker and therapeutic target in non-small cell lung cancer (NSCLC). However, the available results of p-AKT expression in NSCLC are heterogeneous. Therefore, a meta-analysis of published researches investigating the prognostic relevance of p-AKT expression in patients with NSCLC was performed.
Materials and Methods
A literature search via PubMed, EMBASE and CNKI (China National Knowledge Infrastructure) databases was conducted. Data from eligible studies were extracted and included into meta-analysis using a random effects model.
A total of 1049 patients from nine studies were included in the meta-analysis. Nine studies investigated the relationship between p-AKT expression and overall survival using univariate analysis, and five of these undertook multivariate analysis. The pooled hazard ratio (HR) for overall survival was 1.49 (95% confidence interval (CI): 1.01-2.20) by univariate analysis and 1.02 (95% CI: 0.54-1.95) by multivariate analysis.
Our study shows that positive expression of p-AKT is associated with poor prognosis in patients with NSCLC. However, adequately designed prospective studies need to perform.
Aminoacyl-tRNA synthetases (AARSs) are ligases (EC.6.1.1.-) that catalyze the acylation of amino acids to their cognate tRNAs in the process of translating genetic information from mRNA to protein. Their amino acid and tRNA specificity are crucial for correctly translating the genetic code. Glycine is the smallest amino acid and the glycyl-tRNA synthetase (GlyRS) belongs to Class II AARSs. The enzyme is unusual because it can assume different quaternary structures. In eukaryotes, archaebac-teria and some bacteria, it forms an α2 homodimer. In some bacteria, GlyRS is an α2β2 heterotetramer and shows a distant similarity to α2 GlyRSs. The human pathogen eubacterium Campylobacter jejuni GlyRS (CjGlyRS) is an α2β2 heterotetramer and is similar to Escherichia coli GlyRS; both are members of Class IIc AARSs. The two-step aminoacylation reaction of tetrameric GlyRSs requires the involvement of both α- and β-subunits. At present, the structure of the GlyRS α2β2 class and the details of the enzymatic mechanism of this enzyme remain unknown. Here we report the crystal structures of the catalytic α-subunit of CjGlyRS and its complexes with ATP, and ATP and glycine. These structures provide detailed information on substrate binding and show evidence for a proposed mechanism for amino acid activation and the formation of the glycyl-adenylate intermediate for Class II AARSs.
Gly-tRNA synthetase; Catalytic subunit; ATP binding; Glycine binding
AIM: To investigate the effects of photodynamic therapy with quantum dots-arginine-glycine-aspartic acid (RGD) probe as photosensitizer on the proliferation and apoptosis of pancreatic carcinoma cells.
METHODS: Construction of quantum dots-RGD probe as photosensitizer for integrin-targeted photodynamic therapy was accomplished. After cells were treated with photodynamic therapy (PDT), the proliferation of SW1990 cells were measured by methyl thiazolyl tetrazolium assay. Morphologic changes, cell cycle retardance and apoptosis were observed under fluoroscope and flow cytometry. The expression of myeloid cell leukemia-1 (Mcl-1), protein kinase B (Akt) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) mRNA were detected by reverse transcription-polymerase chain reaction. The amount of reactive oxygen species were also evaluated by fluorescence probe.
RESULTS: The photodynamic therapy with quantum dots-RGD probe as photosensitizer significantly inhibited cell proliferation (P < 0.01). Apoptotic cells and morphologic changes could be found under optical microscope. The FCM revealed PDT group had more significant cell apoptosis rate compared to control cells (F = 130.617, P < 0.01) and cell cycle G0/G1 and S retardance (P < 0.05) compared to control cells. The expression of Mcl-1 and Akt mRNA were down-regulated, while expression of TRAIL mRNA was up-regulated after cells treated with PDT. PDT group had more significant number of cells producing reactive oxygen species compared to control cells (F = 3262.559, P < 0.01).
CONCLUSION: The photodynamic therapy with quantum dots-RGD probe as photosensitizer significantly inhibits cell proliferation and increases apoptosis in SW1990 cells.
Pancreatic carcinoma; Targeted probe; Photodynamic therapy; Apoptosis; Reactive oxygen species
The Clustered Regularly Interspaced Palindromic Repeats (CRISPR) system is an adaptive immune system in prokaryotes. Interference complexes encoded by CRISPR-associated (cas) genes utilize small RNAs for homology-directed detection and subsequent degradation of invading genetic elements, and they have been classified into three main types (I–III). Type III complexes share the Cas10 subunit but are subclassifed as type IIIA (CSM) and type IIIB (CMR), depending on their specificity for DNA or RNA targets, respectively. The role of CSM in limiting the spread of conjugative plasmids in Staphylococcus epidermidis was first described in 2008. Here, we report a detailed investigation of the composition and structure of the CSM complex from the archaeon Sulfolobus solfataricus, using a combination of electron microscopy, mass spectrometry, and deep sequencing. This reveals a three-dimensional model for the CSM complex that includes a helical component strikingly reminiscent of the backbone structure of the type I (Cascade) family.
•The CSM complex from Sulfolobus solfataricus has been purified and characterized•EM reveals a helical backbone with striking similarities to the Cascade complex•Mass spectrometry defines the subunit stoichiometry and organization of the complex•CSM subunits are modified by methylation, acetylation, and phosphorylation
Photoacoustic tomography (PAT) is an emerging molecular imaging modality. Here, we demonstrate use of semiconductor copper sulfide nanoparticles (CuS NP) for PAT with an Nd:YAG laser at a wavelength of 1064 nm. CuS NP allowed visualization of mouse brain after intracranial injection, rat lymph nodes 12 mm below the skin after interstitial injection, and CuS NP-containing agarose gel embedded in chicken breast muscle at the depth of ~ 5 cm. This imaging approach has great potential for molecular imaging of breast cancer.
photoacoustic tomography; CuS nanoparticles; 1064-nm laser; optical imaging
To determine the susceptibility genes of lung cancer, we investigated the frequency distributions of the xeroderma pigmentosum complementary group D (XPD) and cytidine deaminase (CDA) genes in patients. A case-control study was conducted involving lung cancer patients and healthy controls. The genotypic distributions of XPD exon 10 G→A (Asp312Asn) and 23 T→G (Lys751Gln), and CDA 79 A→C (Lys27Gln) and 208 G→A (Ala70Thr), were determined using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The results demonstrated that the XPD Asp312Asn genotype distribution was G/G (82.52%) and A/G (17.48%) in the lung cancer patients, and G/G (82.52%), A/G (16.50%) and A/A (10.98%) in the controls. The genotypes of Lys751Gln were T/T (83.49%) and T/G (16.50%) in the lung cancer patients, and T/T (84.47%) and T/G (15.53%) in the controls. Mutations in the XPD single nucleotide polymorphism loci did not demonstrate a significant difference between the two groups (P>0.05). The risk of lung cancer in individuals with mutations at positions 312 and 751 increased 6.13-fold (P=0.047). The CDA Lys27Gln genotype distribution was A/A (78.65%), A/C (20.39%) and C/C (0.98%) in the lung cancer patients, and A/A (79.61%), A/C (19.42%) and C/C (0.98%) in the controls (P=0.985). The CDA Ala70Thr genotype distribution was G/G (98.06%) and A/G (1.94%) in the controls, while all the genotypes were wild-type in the lung cancer patients. The difference between the lung cancer patients and the controls was not statistically significant (P=0.155). There was also no significant difference in the frequency distribution of XPD or CDA between the different pathological types (P>0.05). Our findings demonstrate that the mutation of XPD codons 312 and 751 increases the risk of lung cancer. By contrast, polymorphisms of CDA appear to have little association with lung cancer.
genetic polymorphisms; xeroderma pigmentosum complementary group D; cytidine deaminase; lung cancer
Microcin C (McC) is heptapeptide-adenylate antibiotic produced by Escherichia coli strains carrying the mccABCDEF gene cluster encoding enzymes, in addition to the heptapeptide structural gene mccA, necessary for McC biosynthesis and self-immunity of the producing cell. The heptapeptide facilitates McC transport into susceptible cells, where it is processed releasing a non-hydrolyzable aminoacyl adenylate that inhibits an essential aminoacyl-tRNA synthetase. The self-immunity gene mccF encodes a specialized serine-peptidase that cleaves an amide bond connecting the peptidyl or aminoacyl moieties of, respectively, intact and processed McC with the nucleotidyl moiety. Most mccF orthologs from organisms other than E. coli are not linked to the McC biosynthesis gene cluster. Here, we show that a protein product of one such gene, MccF from Bacillus anthracis (BaMccF), is able to cleave intact and processed McC and we present a series of structures of this protein. Structural analysis of apo-BaMccF and its AMP-complex reveal specific features of MccF-like peptidases that allow them to interact with substrates containing nucleotidyl moieties. Sequence analyses and phylogenetic reconstructions suggest that several distinct subfamilies form the MccF clade of the large S66 family of bacterial serine peptidases. We show that various representatives of the MccF clade can specifically detoxify non-hydrolyzable aminoacyl adenylates differing in their aminoacyl moieties. We hypothesize that bacterial mccF genes serve as a source of bacterial antibiotic resistance.
MccF; serine peptidase; nucleophilic elbow; catalytic triad (Ser-His-Glu); substrate binding loop
A simple and rapid multiplex substrate profiling method has been developed to reveal the substrate specificity of any endo- or exo-peptidase using LC-MS/MS sequencing. A physicochemically diverse library of peptides was generated by incorporating all combinations of neighbor and near-neighbor amino acid pairs into decapeptide sequences that are flanked by unique dipeptides at each terminus. Addition of a panel of evolutionarily diverse peptidases to a mixture of these tetradecapeptides generated prime and non-prime site information and substrate specificity matched or expanded upon previous substrate motifs. This method biochemically confirmed the activity of the klassevirus 3C gene responsible for polypeptide processing and allowed Granzyme B substrates to be ranked by enzymatic turnover efficiency using label-free quantitation of precursor ion abundance. Furthermore, the proteolytic secretions from a parasitic flatworm larvae and a pancreatic cancer cell line were deconvoluted in a subtractive strategy using class-specific peptidase inhibitors.
Lack of immunogenicity of cancer cells has been considered a major reason for their failure in induction of a tumor specific T cell response. In this paper, we present evidence that decitabine (DAC), a DNA methylation inhibitor that is currently used for the treatment of myelodysplastic syndrome (MDS), acute myeloid leukemia (AML) and other malignant neoplasms, is capable of eliciting an anti-tumor cytotoxic T lymphocyte (CTL) response in mouse EL4 tumor model. C57BL/6 mice with established EL4 tumors were treated with DAC (1.0 mg/kg body weight) once daily for 5 days. We found that DAC treatment resulted in infiltration of IFN-γ producing T lymphocytes into tumors and caused tumor rejection. Depletion of CD8+, but not CD4+ T cells resumed tumor growth. DAC-induced CTL response appeared to be elicited by the induction of CD80 expression on tumor cells. Epigenetic evidence suggests that DAC induces CD80 expression in EL4 cells via demethylation of CpG dinucleotide sites in the promoter of CD80 gene. In addition, we also showed that a transient, low-dose DAC treatment can induce CD80 gene expression in a variety of human cancer cells. This study provides the first evidence that epigenetic modulation can induce the expression of a major T cell co-stimulatory molecule on cancer cells, which can overcome immune tolerance, and induce an efficient anti-tumor CTL response. The results have important implications in designing DAC-based cancer immunotherapy.
AIM: To evaluate the use of medical adhesive spray in endoscopic submucosal dissection (ESD).
METHODS: Patients who underwent ESD between January 2009 and June 2012 (n = 173) were enrolled in the prospective randomized study. Two patients undergoing surgery due to severe intraoperative hemorrhage and failed hemostasis were excluded, and the remaining 171 patients were randomly divided into two groups: group A (medical adhesive group, n = 89) and group B (control group, n = 82). In group A, a medical adhesive spray was evenly applied after routine electrocoagulation and hemostasis using hemostatic clip after ESD. Patients in group B only treated with routine wound management. Intraoperative and postoperative data were collected and compared.
RESULTS: In all 171 patients, ESD was successfully completed. There was no significant difference in the average treatment time between groups A and B (59.4 min vs 55.0 min, respectively). The average length of hospital stay was significantly different between group A and B (8.89 d vs 9.90 d, respectively). The incidence of intraoperative perforation was 10.1% in group A and 9.8% in group B, and was not significantly different between the two groups. In all cases, perforations were successfully managed endoscopically and with conservative treatment. The incidence of postoperative delayed bleeding in group A was significantly lower than that in group B (0.00% vs 4.88%, respectively).
CONCLUSION: ESD is an effective minimally invasive treatment for gastrointestinal precancerous lesions or early-stage gastrointestinal cancer. Medical adhesive spray is effective in preventing delayed bleeding after ESD, and can thus reduce the average length of hospital stay.
Endoscopic submucosal dissection; Medical adhesive; Early-stage gastrointestinal cancer; Postoperative delayed bleeding; Intraoperative hemorrhage
To determine whether the use of idarubicin+cytarabine (IA) is more effective than the use of daunorubicin+cytarabine (DA) as induction chemotherapy for patients with newly diagnosed acute myeloid leukaemia.
A computer-based search was performed. Randomised trials comparing IA with DA as induction therapy for newly diagnosed AML were included in this meta-analysis. The primary outcome of interest for our analysis was survival (disease-free survival, event-free survival and overall survival); the secondary endpoint was complete remission.
Ten trials with 4,060 patients were eligible for this meta-analysis. Our pooled results suggest that IA is associated with a significant advantage in CR (RR = 1·23; 95% CI = 1·07–1·41, p = 0.004), EFS (HR = 0·64; 95% CI = 0·45–0·91, p = 0.013), and OS (HR = 0·88; 95% CI = 0·81–0·95, p = 0.02) but not in DFS (HR = 0·90; 95% CI = 0·80–1·00, p = 0.06). In the subgroup analysis, age had a significant interaction with OS and CR benefits.
Our analysis indicated that IA could improve the duration of overall survival compared to DA as induction therapy for young patients with newly diagnosed AML. Further study is needed to determine whether IA can produce clinical benefits in selected genetic or molecular subgroups of young AML patients.
To study the mechanisms in gambogic acid (GA) -induced JeKo-1 human Mantle Cell Lymphoma cell apoptosis in vitro.
The proliferation of GA-treated JeKo-1 cells was measured by CCK-8 assay and Ki-67 immunocytochemical detection. Apoptosis, cell cycle and mitochondrial membrane potential were measured by flow cytometric analysis. Caspase-3, -8 and -9 were detected by colorimetric assay. Bcl-2 and Bax were analyzed by Western blotting.
GA inhibited cell growth in a time- and dose- dependent manner. GA induces apoptosis in JeKo-1 cells but not in normal bone marrow cells, which was involved in reducing the membrane potential of mitochondria, activating caspases-3, -8 and -9 and decreasing the ratio of Bcl-2 and Bax without cell cycle arresting.
GA induced apoptosis in human MCL JeKo-1 cells by regulating Bcl-2/Bax and activating caspase-3, -8 and -9 via mitochondrial pathway without affecting cell cycle.
Gambogic acid; JeKo-1 cells; cell cycle arrest, apoptosis; membrane potential of mitochondria; caspase-3; caspase-8; caspase-9; Bax; Bcl-2
In this paper, we combine the acoustic vector-sensor array parameter estimation problem with the parallel profiles with linear dependencies (PARALIND) model, which was originally applied to biology and chemistry. Exploiting the PARALIND decomposition approach, we propose a blind coherent two-dimensional direction of arrival (2D-DOA) estimation algorithm for arbitrarily spaced acoustic vector-sensor arrays subject to unknown locations. The proposed algorithm works well to achieve automatically paired azimuth and elevation angles for coherent and incoherent angle estimation of acoustic vector-sensor arrays, as well as the paired correlated matrix of the sources. Our algorithm, in contrast with conventional coherent angle estimation algorithms such as the forward backward spatial smoothing (FBSS) estimation of signal parameters via rotational invariance technique (ESPRIT) algorithm, not only has much better angle estimation performance, even for closely-spaced sources, but is also available for arbitrary arrays. Simulation results verify the effectiveness of our algorithm.
arbitrary array; acoustic vector-sensor; coherent angle estimation; PARALIND decomposition
This meta-analysis was performed to evaluate the efficiency and the safety of absorbable implants. Five major electronic databases (PubMed, Embase, Cochrane Library, SinoMed and Wanfang Data) were systematically searched for randomized controlled trials (RCTs) from their establishment to November 2012. Studies on absorbable implants and metal implants for ankle fractures were selected. The meta-analysis was performed using RevMan 5.1. Ten studies with 762 patients were included and analyzed. Compared with metal implants, absorbable implants used for the internal fixation of ankle fractures produce similar radiographic and functional outcomes (P= 0.52). Normally, removal of the internal fixation is unnecessary (P<0.0001) and the incidence of palpable implants is lower (P=0.02) for absorbable implants. No statistically significant difference was observed between the two groups with regard to foreign body reactions (P=0.07), infection (P= 0.69), osteoarthritis (P= 0.39), pain (P= 0.06), refracture (P=0.67), skin necrosis (P=0.99), deep vein thrombosis (P=0.21) and nerve injury (P=0.94). Absorbable implants used in ankle fractures rarely require reoperation and result in similar functional outcomes and complications compared with metal implants. These characteristics make them efficient and reasonably safe for the treatment of ankle fractures.
absorbable implant; metal implant; ankle fracture; meta-analysis
Whether PET scan maximum standard uptake value (SUVmax) could differentiate luminal A from luminal B and help predict the survival of metastatic breast cancer (MBC) patients with luminal subtype is still unknown and need to be investigated.
305 MBC patients with luminal subtypes were screened with PET/CT. Eligible patients were prospectively followed up.
In total, 134 patients were eligible for this study. SUVmax was significantly related to the number of metastatic sites and presence of visceral metastasis on univariate analysis. SUVmax could not effectively differentiate patients with luminal A from luminal B subtype. Although luminal subtype at diagnosis could predict the relapse-free interval, it could not predict progression-free survival (PFS) or overall survival (OS) after developing relapse. In contrast, SUVmax was predictive of both PFS and OS and this effect was maintained in multivariate COX regression model.
SUVmax of MBC did not correlate with molecular subtypes of primary tumor. While molecular subtype may be a valuable prognostic factor at primary diagnosis of breast cancer, the SUVmax, rather than molecular subtype, does have a potential to predict independently in multivariate analysis for the PFS and OS in patients with metastatic disease of luminal subtype.
Metastatic breast cancer; Luminal subtype; PET/CT; SUVmax; Prognosis
Chronic hepatitis B (CHB) is a serious public health problem, and Traditional Chinese Medicine (TCM) plays an important role in the control and treatment for CHB. In the treatment of TCM, zheng discrimination is the most important step. In this paper, an approach based on CFS-GA (Correlation based Feature Selection and Genetic Algorithm) and C5.0 boost decision tree is used for zheng classification and progression in the TCM treatment of CHB. The CFS-GA performs better than the typical method of CFS. By CFS-GA, the acquired attribute subset is classified by C5.0 boost decision tree for TCM zheng classification of CHB, and C5.0 decision tree outperforms two typical decision trees of NBTree and REPTree on CFS-GA, CFS, and nonselection in comparison. Based on the critical indicators from C5.0 decision tree, important lab indicators in zheng progression are obtained by the method of stepwise discriminant analysis for expressing TCM zhengs in CHB, and alterations of the important indicators are also analyzed in zheng progression. In conclusion, all the three decision trees perform better on CFS-GA than on CFS and nonselection, and C5.0 decision tree outperforms the two typical decision trees both on attribute selection and nonselection.
AIM: To evaluate the safety and efficacy of CO2 insufflation compared with air insufflation in the endoscopic submucosal excavation (ESE) of gastrointestinal stromal tumors.
METHODS: Sixty patients were randomized to undergo endoscopic submucosal excavation, with the CO2 group (n = 30) and the air group (n = 30) undergoing CO2 insufflation and air insufflation in the ESE, respectively. The end-tidal CO2 level (pETCO2) was observed at 4 time points: at the beginning of ESE, at total removal of the tumors, at completed wound management, and 10 min after ESE. Additionally, the patients’ experience of pain at 1, 3, 6 and 24 h after the examination was registered using a visual analog scale (VAS).
RESULTS: Both the CO2 group and air group were similar in mean age, sex, body mass index (all P > 0.05). There were no significant differences in PetCO2 values before and after the procedure (P > 0.05). However, the pain scores after the ESE at different time points in the CO2 group decreased significantly compared with the air group (1 h: 21.2 ± 3.4 vs 61.5 ± 1.7; 3 h: 8.5 ± 0.7 vs 42.9 ± 1.3; 6 h: 4.4 ± 1.6 vs 27.6 ± 1.2; 24 h: 2.3 ± 0.4 vs 21.4 ± 0.7, P < 0.05). Meanwhile, the percentage of VAS scores of 0 in the CO2 group after 1, 3, 6 and 24 h was significantly higher than that in the air group (60.7 ± 1.4 vs 18.9 ± 1.5, 81.5 ± 2.3 vs 20.6 ± 1.2, 89.2 ± 0.7 vs 36.8 ± 0.9, 91.3 ± 0.8 vs 63.8 ± 1.3, respectively, P < 0.05). Moreover, the condition of the CO2 group was better than that of the air group with respect to anal exsufflation.
CONCLUSION: Insufflation of CO2 in the ESE of gastrointestinal stromal tumors will not cause CO2 retention and it may significantly reduce the level of pain, thus it is safe and effective.
Carbon dioxide insufflation; Endoscopic submucosal excavation; Gastrointestinal tract; Stromal tumor; Treatment
Abnormalities in the Notch signaling system are considered to play a role in the tumorigenesis of bronchiogenic carcinoma. The present study aimed to investigate the expression of Notch3 in adult lung cancer patients and its role in the pathogenesis of primary bronchiogenic carcinoma. The expression of the Notch3 protein in lung squamous cell carcinoma, adenocarcinoma, small cell carcinoma and corresponding non-tumor tissues was detected by immunohistochemistry. To investigate the expression of Notch3 in adenocarcinoma tissues, Notch3 mRNA and protein expression were measured with reverse transcription polymerase chain reaction (RT-PCR) and western blot analysis, respectively. It was demonstrated that Notch3 had a stronger positive degree of expression in lung squamous cell carcinoma and adenocarcinoma compared with the corresponding non-tumor tissue (P<0.01). The expression of Notch3 in small cell carcinoma tissue was lower compared with that of the corresponding non-tumor tissue (P<0.01). The expression of Notch3 in the lung adenocarcinoma group was the highest of the three lung carcinoma groups (P<0.01). RT-PCR revealed that the expression of Notch3 mRNA in the lung adenocarcinoma group was higher than that of the normal lung group, but there was no statistically significant difference (P=0.05). The expression of Notch1 protein in the lung adenocarcinoma group was significantly higher compared with the normal lung group (P<0.01), as shown by western blot analysis. Notch3 may be involved in the pathogenesis of bronchogenic carcinoma, in particular in the promotion of the lung cancer oncogene, and a difference in its expression may exist in the various pathological types.
bronchiogenic carcinoma; Notch3; lung cancer
Streptomyces species are widely distributed in natural habitats, such as soils, lakes, plants and some extreme environments. Replication loci of several Streptomyces theta-type plasmids have been reported, but are not characterized in details. Conjugation loci of some Streptomyces rolling-circle-type plasmids are identified and mechanism of conjugal transferring are described.
We report the detection of a widely distributed Streptomyces strain Y27 and its indigenous plasmid pWTY27 from fourteen plants and four soil samples cross China by both culturing and nonculturing methods. The complete nucleotide sequence of pWTY27 consisted of 14,288 bp. A basic locus for plasmid replication comprised repAB genes and an adjacent iteron sequence, to a long inverted-repeat (ca. 105 bp) of which the RepA protein bound specifically in vitro, suggesting that RepA may recognize a second structure (e.g. a long stem-loop) of the iteron DNA. A plasmid containing the locus propagated in linear mode when the telomeres of a linear plasmid were attached, indicating a bi-directional replication mode for pWTY27. As for rolling-circle plasmids, a single traA gene and a clt sequence (covering 16 bp within traA and its adjacent 159 bp) on pWTY27 were required for plasmid transfer. TraA recognized and bound specifically to the two regions of the clt sequence, one containing all the four DC1 of 7 bp (TGACACC) and one DC2 (CCCGCCC) and most of IC1, and another covering two DC2 and part of IC1, suggesting formation of a high-ordered DNA-protein complex.
This work (i) isolates a widespread Streptomyces strain Y27 and sequences its indigenous theta-type plasmid pWTY27; (ii) identifies the replication and conjugation loci of pWTY27 and; (iii) characterizes the binding sequences of the RepA and TraA proteins.
Streptomyces; Plasmid; Replication; Conjugation
To investigate the release profile of met-enkephalin, β-endorphin, and dynorphin-A in ruminants' CNS, goats were stimulated by electroacupuncture of 0, 2, 40, 60, 80, or 100 Hz for 30 min. The pain threshold was measured using potassium iontophoresis. The peptide levels were determined with SABC immunohistochemisty. The results showed that 60 Hz increased pain threshold by 91%; its increasing rate was higher (P < 0.01) than any other frequency did. 2 Hz and 100 Hz increased met-enkephalin immunoactivities (P < 0.05) in nucleus accumbens, septal area, caudate nucleus, amygdala, paraventricular nucleus of hypothalamus, periaqueductal gray, dorsal raphe nucleus, and locus ceruleus. The two frequencies elicited β-endorphin release (P < 0.05) in nucleus accumbens, septal area, supraoptic nucleus, ventromedial nucleus of hypothalamus, periaqueductal gray, dorsal raphe nucleus, locus ceruleus, solitary nucleus and amygdala. 60 Hz increased (P < 0.05) met-enkephalin or β-endorphin immunoactivities in the nuclei and areas mentioned above, and habenular nucleus, substantia nigra, parabrachial nucleus, and nucleus raphe magnus. High frequencies increased dynorphin-A release (P < 0.05) in spinal cord dorsal horn and most analgesia-related nuclei. It suggested that 60 Hz induced the simultaneous release of the three peptides in extensive analgesia-related nuclei and areas of the CNS, which may be contributive to optimal analgesic effects and species variation.
Nanomaterials that interact with light provide a unique opportunity for biophotonic nanomedicine. Multifunctional nanoparticles (NPs) that have strong and tunable surface plasmon resonance absorption in the near-infrared region combined with visibility with multiple imaging modalities (magnetic resonance imaging, nuclear imaging, and photoacoustic imaging) have great potential in image-guided therapies. These novel nanostructures, once introduced, are expected to home to solid tumors via the enhanced permeability and retention effect (a passive targeting mechanism) or via targeting ligands bound to their surfaces (an active targeting mechanism). The primary mode of action for photothermal conducting NPs is to convert photoenergy into heat, causing temperature in the treatment volume to be elevated above the thermal damage threshold, which results in irreversible cell killing. It is now recognized that this process, termed photothermal ablation therapy or PTA, although very effective, is unlikely to kill all tumor cells when used alone. In addition to PTA, photothermal conducting NPs can also efficiently trigger drug release and activate RNA interference. Such a multimodal approach, which permits simultaneous PTA therapy, chemotherapy, and therapeutic RNA interference, should provide an opportunity for complete eradication of residual disease.
In this Account, we provide an up-to-date review of the synthesis and characterization, functionalization, and in vitro and in vivo evaluation of NIR light-activatable multifunctional nanostructures used for imaging and therapy, with an emphasis on hollow gold nanospheres, magnetic core–shell gold nanoshells, and semiconductor copper monosulfide NPs. We discuss three types novel drug delivery systems in which hollow gold nanospheres are used to mediate controlled drug release.
Theranostics; hollow gold nanospheres; copper sulfide nanoparticles; near-infrared; drug delivery
AIM: To investigate the preventive effect of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) on bile duct ligation (BDL)-induced liver fibrosis in rats.
METHODS: Liver fibrosis in rats was induced by BDL and AcSDKP was infused subcutaneously for 2 wk via a osmotic minipump (Alzet 2ML4) immediately after BDL operation. After scarifying, serum and liver specimens were collected. Hematoxylin and eosin staining, Sirius red staining, enzyme linked immunosorbent assay, Western blot or real-time polymerase chain reaction were used to determinate liver functions, histological alterations, collagen deposition, mRNA expression of markers for fibroblasts, transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-7 (BMP-7).
RESULTS: When compared to model rats, chronic exogenous AcSDKP infusion suppressed profibrogenic TGF-β1 signaling, α-smooth muscle actin positivity (α-SMA), fibroblast specific protein-1 (FSP-1) staining and collagen gene expression. Col I, Col III, matrix metalloproteinase-2, tissue inhibitors of metalloproteinase-1 and tissue inhibitors of metalloproteinase-2 mRNA expressions were all significantly downregulated by AcSDKP infusion (2.02 ± 1.10 vs 14.16 ± 6.50, 2.02 ± 0.45 vs 10.00 ± 3.35, 2.91 ± 0.30 vs 7.83 ± 1.10, 4.64 ± 1.25 vs 18.52 ± 7.61, 0.46 ± 0.16 vs 0.34 ± 0.12, respectively, P < 0.05). Chronic exogenous AcSDKP infusion attenuated BDL-induced liver injury, inflammation and fibrosis. BDL caused a remarkable increase in alanine transaminase, aspartate transaminase, total bilirubin, and prothrombin time, all of which were reduced by AcSDKP infusion. Mast cells, collagen accumulation, α-SMA, TGF-β1, FSP-1 and BMP-7 increased. The histological appearance of liver specimens was also improved.
CONCLUSION: Infusion of exogenous AcSDKP attenuated BDL-induced fibrosis in the rat liver. Preservation of AcSDKP may be a useful therapeutic approach in the management of liver fibrosis.
N-acetyl-seryl-aspartyl-lysyl-proline; Liver fibrosis; Transformating growth factor-β1; α-smooth mucle actin; Bone morphological protein-7; Fibroblast specific protein-1; Epithelial-mesenchymal transition
Imaging of apoptosis can allow noninvasive assessment of disease states and response to therapeutic intervention for a variety of diseases. The purpose of this study was to develop and evaluate a multimodal nanoplatform for the detection of apoptosis.
To modulate the pharmacokinetics of annexin A5, a 36-kDa protein that binds specifically with phosphatidylserine, annexin A5 was conjugated to polyethylene glycol-coated, core-crosslinked polymeric micelles (CCPM) dually labeled with near-infrared fluorescence fluorophores and a radioisotope (indium 111). To evaluate the specificity of the binding of annexin A5-CCPM to apoptotic cells, both fluorescence microscopy and cell binding studies were performed in vitro. Pharmacokinetics, biodistribution, dual nuclear and optical imaging, and immunohistochemical studies were carried out in 2 xenografted tumor models to evaluate the potential applications of annexin A5-CCPM.
In cell-based studies, annexin A5-CCPM exhibited strongly specific binding to apoptotic tumor cells. This binding could be efficiently blocked by annexin A5. In mice, annexin A5-CCPM displayed a mean elimination half-life of 12.5 h. The mean initial concentration in blood was predicted to be 22.4% of the injected dose/mL, and annexin A5-CCPM was mainly distributed in the central blood compartment. In mice bearing EL4 lymphoma treated with cyclophosphamide and etoposide and in mice bearing MDA-MB-468 breast tumors treated with poly(L-glutamic acid)-paclitaxel and cetuximab (IMC-C225) anti-EGFR antibody, the tumor apoptosis was clearly visualized by both single photon emission computed tomography and fluorescence molecular tomography. In contrast, there was little accumulation of this nanoradiotracer in the tumors of untreated mice. The biodistribution data were consistent with the imaging data, with tumor-to-muscle and tumor-to-blood ratios of 38.8 and 4.1, respectively, in treated mice, and 14.8 and 2.2, respectively, in untreated mice bearing EL4 lymphoma. Moreover, further studies demonstrated that the conventional Tc-99m-labeled HYNIC-annexin A5 and the plain CCPM control exhibited significantly lower uptake in the tumors of the treated mice than annexin A5-CCPM. Immunohistochemistry staining study showed that radioactivity count correlated with fluorescence signal from the nanoparticles, and both signals co-localized with the region of tumor apoptosis.
Annexin A5-CCPM allowed visualization of tumor apoptosis by both nuclear and optical techniques. The complementary information acquired with multiple imaging techniques should be advantageous in assessing and validating early response to therapy.
Apoptosis; Annexin A5; Polymeric Micelles; Nuclear Imaging; Fluorescence Optical Imaging
Image-guided thermal ablation of tumors is becoming a more widely accepted minimally invasive alternative to surgery for patients who are not good surgical candidates, such as patients with advanced head and neck cancer. In this study, multifunctional superparamagnetic iron oxide coated with gold nanoshell (SPIO@Au NS) that have both optical and magnetic properties was conjugated with the targeting agent, C225 monoclonal antibody, against epidermal growth factor receptor (EGFR). C225-SPIO@Au NS have an average a diameter of 82±4.4 nm, contain 142±15 antibodies per nanoshell, have an absorption peak in the near infrared (~800 nm), and have transverse relaxivity (r2) of 193 and 353 mM−1s−1 versus Feridex of 171 and 300 mM−1s−1, using 1.5T and 7T MR scanners, respectively. Specific targeting of the synthesized C225-SPIO@Au NS was tested in vitro using A431 cells and oral cancer cells, FaDu, OSC-19, and HN5, all of which overexpress EGFR. Selective binding was achieved using C225-SPIO@Au NS but not with the non-targeting PEG-SPIO@Au NS and blocking group (excess of C225 + C225-SPIO@Au NS). In vivo biodistribution on mice bearing A431 tumors also showed selective targeting of C225-SPIO@Au NS compared with the non-targeting and blocking groups. The selective photothermal ablation of the nanoshells shows that without laser treatment there were no cell death and among the groups that were treated with laser at a power of 36 W/cm2 for 3 minutes, only the cells treated with C225-SPIO@Au NS had cell killing (p < 0.001). In summary, successful synthesis and characterization of targeted C225-SPIO@Au NS demonstrating both superparamagnetic and optical properties has been achieved. We have shown both in vitro and in vivo that these nanoshells are MR-active and can be selectively heated up for simultaneous imaging and photothermal ablation therapy.
Theranostics; Gold nanoshells; Magnetic resonance imaging; Laser ablation; Ultrasmall paramagnetic iron oxide (SPIO)
Quality of life (QoL) is increasingly recognized as an important outcome measure in clinical trials. The Canadian Hemophilia Outcomes-Kids Life Assessment Tool (CHO-KLAT) shows promise for use in China.
To adapt the CHO-KLAT version 2.0 for use in clinical trials in China.
Forward and back translations of the CHO-KLAT2.0 were completed in 2008. Between October 2009 and June 2010, a series of 3 focus groups were held with 20 boys and 31 parents in rural and urban China to elicit additional concepts, important to their QoL, for the Chinese CHO-KLAT2.0. All of the items identified by boys and parents were reviewed by a group of experts, resulting in a Chinese version of the CHO-KLAT2.0. This version underwent a detailed cognitive debriefing process between October 2010 and June 2011. Thirteen patient-parent pairs participated in this cognitive debriefing process until a stable and clearly understood Chinese version of the CHO-KLAT2.0 was obtained.
The initial back translation of the Chinese CHO-KLAT2.0 was slightly discrepant from the original English version on 12 items. These were all successfully adjudicated. The focus groups identified 9 new items that formed an add-on Socio-Economic Context (SEC) module for China. Linguistic improvements were made after the 2nd, 5th, 7th and 13th cognitive debriefings pairs and affected a total of 18 items. The result was a 35 item CHO-KLAT2.0 and a SEC module in Simplified Chinese, both of which have good content validity.
This detailed process proved to be extremely valuable in ensuring the items were accurately interpreted by Chinese boys with hemophilia ages ≤18 years. The need for the additional SEC module highlighted the different context that currently exists in China with regard to hemophilia care as compared to many Western countries, and will be important in tracking progress within both rural and urban China over time. Changes based on the cognitive debriefings suggest that expert verbatim translation alone is not sufficient. The Chinese version of the CHO-KLAT2.0 is well understood by boys with hemophilia in China. Next steps will be to test its construct validity and reliability in boys with hemophilia in China.
Quality of life; Hemophilia; China; Children; Cross-cultural studies