A few studies focused on unilateral or bilateral pedicle screw (PS) fixation of minimally invasive transforaminal lumbar interbody fusion (MIS-TLIF) to treat lumbar degenerative diseases have been published. There is still debate over whether one method is superior to another. A systematic review and meta-analysis of randomized controlled trials (RCT) was performed to compare the efficacy of the two methods.
Focal epilepsies often originate in the hippocampal formation of the temporal lobe (temporal lobe epilepsy) and are generally acquired after transient brain insults. Such insults induce cellular and structural reorganization processes of the hippocampus, referred to as epileptogenesis that finally convert the brain spontaneous epileptic. Here, we developed a new molecular imaging strategy in a state-of-the-art animal model to provide insights into key epileptogenic mechanisms. Our new approach combines recombinant adeno-associated virus (rAAV) gene delivery with in vivo bioluminescence imaging. rAAV particles harboring the luciferase reporter gene under control of the minimal T type Ca(2+)-channel subunit Ca V 3.2-promoter were generated and injected stereotaxically in the hippocampal region of mice. Bioluminescent signals, corresponding to Ca V 3.2 promoter activation, were imaged in vivo in the pilocarpine model of status epilepticus (SE). We detected activation of key Ca V 3.2 promoter motifs at 3 and 10 days after SE but not after the onset of chronic seizures. These data suggest Ca V 3.2 promoter activation as novel anti-epileptogenic target. In more general terms, we have established an experimental approach that allows to follow cerebral gene promoter dynamics longitudinally and to correlate this activity to behavioral parameters in the same mice.
To determine: (1) the relationship of thoracic cage parameters and preoperative pulmonary function tests (PFTs) in congenital scoliosis (CS) patients. (2) if patients with rib deformity have greater impairment of PFTs than those without rib deformity.
Natural killer T (NKT) cells share phenotypic and functional properties with both conventional natural killer cells and T cells. These cells might have an important role in the pathogenesis of ulcerative colitis (UC). The interaction of chemokine ligand 25 (CCL25) with chemokine receptor 9 (CCR9) is involved in gut-specific migration of leukocytes and induces regulatory T cells (Tregs) to migrate to the intestine in chronic ileitis.
Cathelicidins, a class of gene-encoded effector molecules of vertebrate innate immunity, provide a first line of defense against microbial invasions. Although cathelicidins from mammals, birds, reptiles and fishes have been extensively studied, little is known about cathelicidins from amphibians. Here we report the identification and characterization of two cathelicidins (cathelicidin-RC1 and cathelicidin-RC2) from the bullfrog Rana catesbeiana. The cDNA sequences (677 and 700 bp, respectively) encoding the two peptides were successfully cloned from the constructed lung cDNA library of R. catesbeiana. And the deduced mature peptides are composed of 28 and 33 residues, respectively. Structural analysis indicated that cathelicidin-RC1 mainly assumes an amphipathic alpha-helical conformation, while cathelicidin-RC2 could not form stable amphipathic structure. Antimicrobial and bacterial killing kinetic analysis indicated that the synthetic cathelicidin-RC1 possesses potent, broad-spectrum and rapid antimicrobial potency, while cathelicidin-RC2 exhibited very weak antimicrobial activity. Besides, the antimicrobial activity of cathelicidin-RC1 is salt-independent and highly stable. Scanning electron microscopy (SEM) analysis indicated that cathelicidin-RC1 kills microorganisms through the disruption of microbial membrane. Moreover, cathelicidin-RC1 exhibited low cytotoxic activity against mammalian normal or tumor cell lines, and low hemolytic activity against human erythrocytes. The potent, broad-spectrum and rapid antimicrobial activity combined with the salt-independence, high stability, low cytotoxic and hemolytic activities make cathelicidin-RC1 an ideal template for the development of novel peptide antibiotics.
Prostate cancer is relatively common cancer occurring in males. Radical prostatectomy (RP) is the most effective treatment for a localized tumor but erectile dysfunction (ED) is common complication, even when bilateral nerve-sparing RP (BNSRP) is performed. Clinical trials have shown varied effectiveness of phosphodiesterase type-5 inhibitors (PDE5-Is) for treatment of post-BNSRP ED, but there remains controversy over the application of this treatment and no formal systematic review and meta-analysis for the use of PDE5-Is for this condition has been conducted. This review was to systematically assess the efficacy and safety of oral PDE5-Is for post-BNSRP ED. A database search was conducted to identify randomized controlled trials (RCTs). The comparative efficacy of treatments was analyzed by fixed or random effect modeling. Erectile function was measured using the International Index of Erectile Function (IIEF), Sexual Encounter Profile (SEP) question-2, 3 and the Global Assessment Question (GAQ). The rate and incidence of adverse events (AEs) were determined. The quality of included studies was appraised using the Cochrane Collaboration bias appraisal tool. Eight RCTs were included in the analyses. PDE5-Is were effective for treating post-BNSRP ED compared to placebo when erectile function was determined using the IIEF score [mean difference (MD) 5.63, 95% confidence interval (CI) (4.26-6.99)], SEP-2 [relative risk (RR) 1.63, 95% CI (1.18-2.25) ], SEP-3 [RR 2.00, 95% CI (1.27-3.15) ] and GAQ [RR 3.35, 95% CI (2.68-4.67) ]. The subgroup analysis could find a trend that longer treatment duration, higher dosage, on-demand dosing, sildenafil and mild ED are associated with more responsiveness to PDE5-Is. PDE5-Is were overall well tolerated with headache being the most commonly reported AE. Our data provides compelling evidence for the use of PDE5-Is as a primary treatment for post-BNSRP ED. However, further studies are required to optomize usage parameters (such as dosage and duration of treatment).
No recent studies have analyzed the rates of or reasons for unanticipated revision surgery within 30 days of primary surgery in spinal deformity patients. Our aim was to examine the incidence, characteristics, reasons, and risk factors for unplanned revision surgery in spinal deformity patients treated at one institution. All patients with a diagnosis of spinal deformity presenting for primary instrumented spinal fusion at a single institution from 1998 to 2012 were reviewed. All unplanned reoperations performed within 30 days after primary surgery were analyzed in terms of demographics, surgical data, and complications. Statistical analyses were performed to obtain correlations and risk factors for anticipated revision. Of 2758 patients [aged 16.07 years (range, 2-71), 69.8% female] who underwent spinal fusion surgery, 59 (2.1%) required reoperation within 30 days after primary surgery. The length of follow up for each patient was more than 30 days. Of those that required reoperation, 87.0% had posterior surgery only, 5.7% had anterior surgery, and 7.3% underwent an anteroposterior approach. The reasons for reoperation included implant failure (n?=?20), wound infection (n?=?12), neurologic deficit (n?=?9), pulmonary complications (n?=?17), and coronal plane imbalance (n?=?1). The risk factors for reoperation were age, diagnosis, and surgical procedure with osteotomy.
It is difficult to construct a control group for trials of adjuvant therapy (Rx) of prostate cancer after radical prostatectomy (RP) due to ethical issues and patient acceptance. We utilized 8 curve-fitting models to estimate the time to 60%, 65%, … 95% chance of progression free survival (PFS) based on the data derived from Kattan post-RP nomogram. The 8 models were systematically applied to a training set of 153 post-RP cases without adjuvant Rx to develop 8 subsets of cases (reference case sets) whose observed PFS times were most accurately predicted by each model. To prepare a virtual control group for a single-arm adjuvant Rx trial, we first select the optimal model for the trial cases based on the minimum weighted Euclidean distance between the trial case set and the reference case set in terms of clinical features, and then compare the virtual PFS times calculated by the optimum model with the observed PFSs of the trial cases by the logrank test. The method was validated using an independent dataset of 155 post-RP patients without adjuvant Rx. We then applied the method to patients on a Phase II trial of adjuvant chemo-hormonal Rx post RP, which indicated that the adjuvant Rx is highly effective in prolonging PFS after RP in patients at high risk for prostate cancer recurrence. The method can accurately generate control groups for single-arm, post-RP adjuvant Rx trials for prostate cancer, facilitating development of new therapeutic strategies.
Lysozymes are key proteins that play important roles in innate immune defense in many animal phyla by breaking down the bacterial cell-walls. In this study, we report the molecular cloning, sequence analysis and phylogeny of the first caudate amphibian g-lysozyme: a full-length spleen cDNA library from axolotl (Ambystoma mexicanum). A goose-type (g-lysozyme) EST was identified and the full-length cDNA was obtained using RACE-PCR. The axolotl g-lysozyme sequence represents an open reading frame for a putative signal peptide and the mature protein composed of 184 amino acids. The calculated molecular mass and the theoretical isoelectric point (pl) of this mature protein are 21523.0 Da and 4.37, respectively. Expression of g-lysozyme mRNA is predominantly found in skin, with lower levels in spleen, liver, muscle, and lung. Phylogenetic analysis revealed that caudate amphibian g-lysozyme had distinct evolution pattern for being juxtaposed with not only anura amphibian, but also with the fish, bird and mammal. Although the first complete cDNA sequence for caudate amphibian g-lysozyme is reported in the present study, clones encoding axolotls other functional immune molecules in the full-length cDNA library will have to be further sequenced to gain insight into the fundamental aspects of antibacterial mechanisms in caudate.
Antifreeze proteins (AFPs) refer to a class of polypeptides that are produced by certain vertebrates, plants, fungi, and bacteria and which permit their survival in subzero environments. In this study, we report the molecular cloning, sequence analysis and three-dimensional structure of the axolotl antifreeze-like protein (AFLP) by homology modeling of the first caudate amphibian AFLP. We constructed a full-length spleen cDNA library of axolotl (Ambystoma mexicanum). An EST having highest similarity (?42%) with freeze-responsive liver protein Li16 from Rana sylvatica was identified, and the full-length cDNA was subsequently obtained by RACE-PCR. The axolotl antifreeze-like protein sequence represents an open reading frame for a putative signal peptide and the mature protein composed of 93 amino acids. The calculated molecular mass and the theoretical isoelectric point (pl) of this mature protein were 10128.6 Da and 8.97, respectively. The molecular characterization of this gene and its deduced protein were further performed by detailed bioinformatics analysis. The three-dimensional structure of current AFLP was predicted by homology modeling, and the conserved residues required for functionality were identified. The homology model constructed could be of use for effective drug design. This is the first report of an antifreeze-like protein identified from a caudate amphibian.
In case-control profiling studies, increasing the sample size does not always improve statistical power because the variance may also be increased if samples are highly heterogeneous. For instance, tumor samples used for gene expression assay are often heterogeneous in terms of tissue composition or mechanism of progression, or both; however, such variation is rarely taken into account in expression profiles analysis. We use a prostate cancer prognosis study as an example to demonstrate that solely recruiting more patient samples may not increase power for biomarker detection at all. In response to the heterogeneity due to mixed tissue, we developed a sample selection strategy termed Stepwise Enrichment by which samples are systematically culled based on tumor content and analyzed with t-test to determine an optimal threshold for tissue percentage. The selected tissue-percentage threshold identified the most significant data by balancing the sample size and the sample homogeneity; therefore, the power is substantially increased for identifying the prognostic biomarkers in prostate tumor epithelium cells as well as in prostate stroma cells. This strategy can be generally applied to profiling studies where the level of sample heterogeneity can be measured or estimated.
To protect themselves against the invasion of microorganisms, amphibians, especially the Rana frogs, are possibly equipped with complex combinations of antimicrobial peptides (AMPs). The two major AMP families, ranid skin secretion AMPs and cathelicidins might together constitute the host innate immune system of amphibians. Cathelicidins are a group of cationic peptides found in leukocytes and epithelial cells, and they play a central role in the early innate immune defense found in virtually all species of mammals. However, they have rarely been reported from amphibians. Here, we report the identification and discovery of polymorphism cathelicidins in Limnonectes fragilis. The expression profile indicated high cathelicidin transcript levels in frog spleen, liver and kidney, but lower levels in lung, skin and stomach. According to the amphibians unique proteolytic pattern, R125 and L121 of the prepropeptides are predicted to be the processing positions for protease to generate the mature peptides, Lf-CATH1 and -2, respectively. Both consist of 30 amino acid residues, of which two were cysteines positionally conserved among a few known amphibian cathelicidins. Homology modeling analysis revealed that Lf-CATH1 and -2 adopt a tertiary structure with a mostly ? helix that is representative of small cationic cathelicidin family peptides. Recombinant Lf-CATH1 (rLf-CATH1) was produced in Escherichia coli. Synthetic Lf-CATH1 and -2 displayed potent antimicrobial activities in vitro against a broad spectrum of microorganisms, including standard and clinically isolated drug-resistant strains, while showing neglectable hemolysis and cytotoxicities.
Enzymatic synthesis of some industrially important compounds depends heavily on cofactor NADPH as the reducing agent. This is especially true in the synthesis of chiral compounds that are often used as pharmaceutical intermediates to generate the correct stereochemistry in bioactive products. The high cost and technical difficulty of cofactor regeneration often pose a challenge for such biocatalytic reactions. In this study, to increase NADPH bioavailability, the native NAD(+)-dependent glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gapA gene in Escherichia coli was replaced with a NADP(+)-dependent gapB from Bacillus subtilis. To overcome the limitation of NADP(+) availability, E. coli NAD kinase, nadK was also coexpressed with gapB. The recombinant strains were then tested in three reporting systems: biosynthesis of lycopene, oxidation of cyclohexanone with cyclohexanone monooxygenase (CHMO), and an anaerobic system utilizing 2-haloacrylate reductase (CAA43). In all the reporting systems, replacing NAD(+)-dependent GapA activity with NADP(+)-dependent GapB activity increased the synthesis of NADPH-dependent compounds. The increase was more pronounced when NAD kinase was also overexpressed in the case of the one-step reaction catalyzed by CAA43 which approximately doubled the product yield. These results validate this novel approach to improve NADPH bioavailability in E. coli and suggest that the strategy can be applied in E. coli or other bacterium-based production of NADPH-dependent compounds.
Cofactors provide redox carriers for biosynthetic reactions, catabolic reactions and act as important agents in transfer of energy for the cell. Recent advances in manipulating cofactors include culture conditions or additive alterations, genetic modification of host pathways for increased availability of desired cofactor, changes in enzyme cofactor specificity, and introduction of novel redox partners to form effective circuits for biochemical processes and biocatalysts. Genetic strategies to employ ferredoxin, NADH and NADPH most effectively in natural or novel pathways have improved yield and efficiency of large-scale processes for fuels and chemicals and have been demonstrated with a variety of microbial organisms.
NADPH-dependent reactions play important roles in production of industrially valuable compounds. In this study, we used phosphofructokinase (PFK)-deficient strains to direct fructose-6-phosphate to be oxidized through the pentose phosphate pathway (PPP) to increase NADPH generation. pfkA or pfkB single deletion and double-deletion strains were tested for their ability to produce lycopene. Since lycopene biosynthesis requires many NADPH, levels of lycopene were compared in a set of isogenic strains, with the pfkA single deletion strain showing the highest lycopene yield. Using another NADPH-requiring process, a one-step reduction reaction of 2-chloroacrylate to 2-chloropropionic acid by 2-haloacrylate reductase, the pfkA pfkB double-deletion strain showed the highest yield of 2-chloropropionic acid product. The combined effect of glucose-6-phosphate dehydrogenase overexpression or lactate dehydrogenase deletion with PFK deficiency on NADPH bioavailability was also studied. The results indicated that the flux distribution of fructose-6-phosphate between glycolysis and the pentose phosphate pathway determines the amount of NAPDH available for reductive biosynthesis.
Thick-lipped lenok, Brachymystax lenok is one of the ancient fish species in China and northeast Asia countries. Due to the overfishing, the population of lenok has been declined significantly. Cathelicidins are innate immune effectors that possess both bactericidal activities and immunomodulatory functions. This report identifies and characterizes the salmonoid cathelicidin (CATH_BRALE) from this ancient fish. It consists of open reading frame (ORF) of 886 bp encoding the putative peptide of 199 amino acids. Sequence alignment with other representative salmonid cathelicidins displayed two distinctive features of current lenok cathelicidin: high level of arginine, resulting in high positive charge and glycine residues, which is significantly different from most acknowledged types of cathelicidins; and the six-aminoacid tandem repeated sequence of RPGGGS detected in a variable number of copies among fish cathelicidins, suggesting the existence of a genetically unstable region similar to that found in some mammalian cathelicidins. Expression of CATH_BRALE is predominantly found in gill, with lower levels in the gastrointestinal tract and spleen. The homology modeled structure of CATH_BRALE exhibits structural features of antiparallel b-sheets flanked by a-helices that are representative of small cationic cathelicidin family peptides. CATH_BRALE possesses much stronger antimicrobial activity against gram-negative bacteria than that of the human ortholog, LL-37. The growth of two typical fish bacterial pathogens, gram-negative bacterium of Aeromonas salmonicida and Aeromonas hydrophila was substantially inhibited by synthetic CATH_BRALE, with both MICs as low as 9.38 mM.
To compare the clinical effectiveness of posterior lumbar interbody fusion (PLIF) and posterolateral fusion (PLF) for lumbar spondylolisthesis and to collect scientific evidence for determining which fusion method is better.
Increasing evidence suggests that innate immunity plays an important role in alcohol-induced liver injury and most studies have focused on positive regulation of innate immunity. The main objective of this study was to investigate the negative regulator of innate immunity, IL-1/Toll-like receptor (TLR) signaling pathways and interleukin receptor-associated kinase-M (IRAK-M) in alcoholic liver injury. We established an alcohol-induced liver injury model using wild type and IRAK-M deficient B6 mice and investigated the possible mechanisms. We found that in the absence of IRAK-M, liver damage by alcohol was worse with higher alanine transaminase (ALT), more immune cell infiltration and increased numbers of IFN? producing cells. We also found enhanced phagocytic activity in CD68(+) cells. Moreover, our results revealed altered gut bacteria after alcohol consumption and this was more striking in the absence of IRAK-M. Our study provides evidence that IRAK-M plays an important role in alcohol-induced liver injury and IRAK-M negatively regulates the innate and possibly the adaptive immune response in the liver reacting to acute insult by alcohol. In the absence of IRAK-M, the hosts developed worse liver injury, enhanced gut permeability and altered gut microbiota.
With the use of ChIP on microarray assays in primary leukemia samples, we report that acute myeloid leukemia (AML) blasts exhibit significant alterations in histone H3 acetylation (H3Ac) levels at > 1000 genomic loci compared with CD34(+) progenitor cells. Importantly, core promoter regions tended to have lower H3Ac levels in AML compared with progenitor cells, which suggested that a large number of genes are epigenetically silenced in AML. Intriguingly, we identified peroxiredoxin 2 (PRDX2) as a novel potential tumor suppressor gene in AML. H3Ac was decreased at the PRDX2 gene promoter in AML, which correlated with low mRNA and protein expression. We also observed DNA hypermethylation at the PRDX2 promoter in AML. Low protein expression of the antioxidant PRDX2 gene was clinically associated with poor prognosis in patients with AML. Functionally, PRDX2 acted as inhibitor of myeloid cell growth by reducing levels of reactive oxygen species (ROS) generated in response to cytokines. Forced PRDX2 expression inhibited c-Myc-induced leukemogenesis in vivo on BM transplantation in mice. Taken together, epigenome-wide analyses of H3Ac in AML led to the identification of PRDX2 as an epigenetically silenced growth suppressor, suggesting a possible role of ROS in the malignant phenotype in AML.
Triple-negative breast cancer (TNBC) is an aggressive breast cancer subtype with a high rate of proliferation and metastasis, as well as poor prognosis for advanced-stage disease. Although TNBC was previously classified together with basal-like and BRCA1/2-related breast cancers, genomic profiling now shows that there is incomplete overlap, with important distinctions associated with each subtype. The biology of TNBC is still poorly understood; therefore, to define the relative contributions of major cellular pathways in TNBC, we have studied its molecular signature based on analysis of gene expression. Comparisons were then made with normal breast tissue. Our results suggest the existence of molecular networks in TNBC, characterized by explicit alterations in the cell cycle, DNA repair, nucleotide synthesis, metabolic pathways, NF-?B signaling, inflammatory response, and angiogenesis. Moreover, we also characterized TNBC as a cancer of mixed phenotypes, suggesting that TNBC extends beyond the basal-like molecular signature and may constitute an independent subtype of breast cancer. The data provide a new insight into the biology of TNBC.
Non-small cell lung cancer (NSCLC) is the most common type of lung cancer, with a poor response to chemotherapy and low survival rate. This unfavorable treatment response is likely to derive from both late diagnosis and from complex, incompletely understood biology, and heterogeneity among NSCLC subtypes. To define the relative contributions of major cellular pathways to the biogenesis of NSCLC and highlight major differences between NSCLC subtypes, we studied the molecular signatures of lung adenocarcinoma (ADC) and squamous cell carcinoma (SCC), based on analysis of gene expression and comparison of tumor samples with normal lung tissue. Our results suggest the existence of specific molecular networks and subtype-specific differences between lung ADC and SCC subtypes, mostly found in cell cycle, DNA repair, and metabolic pathways. However, we also observed similarities across major gene interaction networks and pathways in ADC and SCC. These data provide a new insight into the biology of ADC and SCC and can be used to explore novel therapeutic interventions in lung cancer chemoprevention and treatment.
Genus Odorrana, among all amphibians studied, is generally reported to have the most abundant and diversified anti-microbial peptides even from a single individual frog. In our previous work, 46 cDNA sequences encoding precursors of 22 different anti-microbial peptides (AMPs) were characterized from the skin of frog, Odorrana tiannanensis. In this work, we reported the purification of three AMPs from skin secretions of O. tiannanensis. Their amino acid sequences matched well with the sequences deduced from cDNAs and they were designated as Odorranain-C7HSa, Brevinin-1-OT2 and Odorranain-G-OT, respectively. Furthermore, we selected to analyze the four most structurally diversified sequences among the 22 AMPs that are significantly different from all reported AMPs. By structural characterization, three of them were designated as pleurain-E-OT, odorranain-G-OT, odorranain-A-OT, belonging to AMP families already identified. The forth one with a unique 14-mer sequence of AILTTLANWARKFLa and C-terminal amidation represents the prototypes of a new class of amphibian AMP, and thereby named tiannanensin. Such broad diversity in sequences and structures are consistent with other species in Genus Odorrana. Multi-functions of the synthesized four special AMPs were screened, including anti-microbial, antioxidant, cytotoxic and hemolytic activities. The results suggest that these AMPs may employ sophisticated mechanisms of action in host defense in addition to anti-microbial, although their precise contribution to host defense still seems unclear.
Modulating the tau level may represent a therapeutic target for Alzheimers disease (AD), as accumulating evidence shows that Abeta-induced neurodegeneration is mediated by tau. It is therefore important to understand the expression and degradation of tau in neurons. Recently we showed that overexpressed mutant tau and tau aggregates are degraded via the autophagic pathway in an N2a cell model. Here we investigated whether autophagy is involved in the degradation of endogenous tau in cultured primary neurons. We activated this pathway in primary neurons with trehalose, an enhancer of autophagy. This resulted in the reduction of endogenous tau protein. Tau phosphorylation at several sites elevated in AD pathology had little influence on its degradation by autophagy. Furthermore, by using a neuronal cell model of tauopathy, we showed that activation of autophagy suppresses tau aggregation and eliminates cytotoxicity. Notably, apart from activating autophagy, trehalose also inhibits tau aggregation directly. Thus, trehalose may be a good candidate for developing therapeutic strategies for AD and other tauopathies.
More than one million prostate biopsies are performed in the United States every year. A failure to find cancer is not definitive in a significant percentage of patients due to the presence of equivocal structures or continuing clinical suspicion. We have identified gene expression changes in stroma that can detect tumor nearby. We compared gene expression profiles of 13 biopsies containing stroma near tumor and 15 biopsies from volunteers without prostate cancer. About 3,800 significant expression changes were found and thereafter filtered using independent expression profiles to eliminate possible age-related genes and genes expressed at detectable levels in tumor cells. A stroma-specific classifier for nearby tumor was constructed on the basis of 114 candidate genes and tested on 364 independent samples including 243 tumor-bearing samples and 121 nontumor samples (normal biopsies, normal autopsies, remote stroma, as well as stroma within a few millimeters of tumor). The classifier predicted the tumor status of patients using tumor-free samples with an average accuracy of 97% (sensitivity = 98% and specificity = 88%) whereas classifiers trained with sets of 100 randomly generated genes had no diagnostic value. These results indicate that the prostate cancer microenvironment exhibits reproducible changes useful for categorizing the presence of tumor in patients when a prostate sample is derived from near the tumor but does not contain any recognizable tumor.
Cathelicidins comprise a family of antimicrobial peptides sharing a highly conserved cathelin domain, which play a central role in the early innate host defense against infection. In the present study, we report three novel avian cathelicidin orthologs cloned from a constructed spleen cDNA library of Coturnix coturnix, using a nested-PCR-based cloning strategy. Three coding sequences containing ORFs of 447, 465 and 456 bp encode three mature antimicrobial peptides (named Cc-CATH1, 2 and 3) of 26, 32 and 29 amino acid residues, respectively. Phylogenetic analysis indicated that precursors of Cc-CATHs are significantly conserved with known avian cathelicidins. Synthetic Cc-CATH2 and 3 displayed broad and potent antimicrobial activity against most of the 41 strains of bacteria and fungi tested, especially the clinically isolated drug-resistant strains, with minimum inhibitory concentration values in the range 0.3-2.5 ?m for most strains with or without the presence of 100 mm NaCl. Cc-CATH2 and 3 showed considerable reduction of cytotoxic activity compared to other avian cathelicidins, with average IC(50) values of 20.18 and 17.16 ?m, respectively. They also exerted a negligible hemolytic activity against human erythrocytes, lysing only 3.6% of erythrocytes at a dose up to 100 ?g·mL(-1) . As expected, the recombinant Cc-CATH2 (rCc-CATH2) also showed potent bactericidal activity. All these features of Cc-CATHs encourage further studies aiming to estimate their therapeutic potential as drug leads, as well as coping with current widespread antibiotic resistance, especially the new prevalent and dangerous superbug that is resistant to almost all antibiotics.
Cathelicidins are a family of antimicrobial peptides acting as multifunctional effector molecules in innate immunity. Cathelicidin-BF has been purified from the snake venoms of Bungarus fasciatus and it is the first identified cathelicidin antimicrobial peptide in reptiles. In this study, cathelicidin-BF was found exerting strong antibacterial activities against Propionibacterium acnes. Its minimal inhibitory concentration against two strains of P. acnes was 4.7 µg/ml. Cathelicidin-BF also effectively killed other microorganisms including Staphylococcus epidermidis, which was possible pathogen for acne vulgaris. Cathelicidin-BF significantly inhibited pro-inflammatory factors secretion in human monocytic cells and P. acnes-induced O2.- production of human HaCaT keratinocyte cells. Observed by scanning electron microscopy, the surfaces of the treated pathogens underwent obvious morphological changes compared with the untreated controls, suggesting that this antimicrobial peptide exerts its action by disrupting membranes of microorganisms. The efficacy of cathelicidin-BF gel topical administering was evaluated in experimental mice skin colonization model. In vivo anti-inflammatory effects of cathelicidin-BF were confirmed by relieving P. acnes-induced mice ear swelling and granulomatous inflammation. The anti-inflammatory effects combined with potent antimicrobial activities and O2.- production inhibition activities of cathelicidin-BF indicate its potential as a novel therapeutic option for acne vulgaris.
Direct-acting fibrin(ogen)olytic agents such as plasmin have been proved to contain effective and safety thrombolytic potential. Unfortunately, plasmin is ineffective when administered by the intravenous route because it was neutralized by plasma antiplasmin. Direct-acting fibrin(ogen)olytic agents with resistance against antiplasmin will brighten the prospect of anti-thrombosis. As reported in Compendium of Materia Medica, the insect of Eupolyphaga sinensis Walker has been used as traditional anti-thrombosis medicine without bleeding risk for several hundreds years. Currently, we have identified a fibrin(ogen)olytic protein (Eupolytin1) containing both fibrin(ogen)olytic and plasminogen-activating (PA) activities from the beetle, E. sinensis.
There have been several reports on hemivertebra resection via a posterior-only procedure. However, the number of reported cases is small, and various types of instrumentation have been used. In our study, we retrospectively investigated 56 consecutive cases of congenital scoliosis that were treated by posterior hemivertebra resection with transpedicular instrumentation. Radiographs were reviewed to determine the type and location of the hemivertebra, the coronal curve magnitude and the sagittal alignment pre-operatively, post-operatively and at the latest follow-up. Radiographs were also used to assess implant failure and inter-body fusion. Surgical reports and patient charts were reviewed to record any peri-operative complications. Fifty-eight posterior hemivertebrae resections from 56 patients aged 1.5-17 years with fully segmented non-incarcerated hemivertebra were evaluated. The average age at surgery was 9.9 years (1.5-17 years). The average follow-up was 32.9 months (24-58 months). The mean fusion level was 5.0 segments (2-11 segments). There was a mean improvement of 72.9% in the segmental scoliosis, from 42.4° before surgery to 12.3° at the time of the latest follow-up, and there was a mean improvement of 70% in segmental kyphosis from 42.0° to 14.5° over the same time period. The thoracic kyphosis (T5-T12) averaged 10.8° before surgery and 23.9° at the latest follow-up. The lumbar lordosis (L1-S1) averaged -52.8° before surgery and -51.6° at the latest follow-up. Two cases with neurological claudications had complete recovery immediately after the surgery. There was one case of delayed wound healing, two fractures of the pedicle at the instrumented level, two rod breakages and one proximal junction kyphosis that required revision. There were no neurological complications. Radiolucent gaps were found in the residual space after resection on the lateral view in five cases, without any sign of implant failure or correction loss. Our results show that one-stage posterior hemivertebra resection with transpedicular instrumentation can achieve excellent correction, 360° decompression and short fusion without neurological complications. Pedicle cutting still remains a challenge in younger children when using bisegmental instrumentation. In addition, the radiolucent gaps in the residual space require further investigation.
In order to avoid low absorption, incorporation, and undesirable side effects of inorganic oxovanadium compounds, the antidiabetic activities of organic oxovanadium (IV) compounds in alloxan-induced diabetic mice were investigated. Vanadyl carboxymethyl carrageenan (VOCCA) and vanadyl carboxymethyl chitosan (VOCCH) were synthesized and administrated through intragastric administration in different doses for 20 days in alloxan-induced diabetic mice. Glibenclamide was administrated as the positive control. Our results showed that low-dose group, middle-dose group, and high-dose group of VOCCA and VOCCH could significantly reduce the levels of blood glucose (P < 0.05) compared with untreated group, but not in normal mice. Besides, high-dose groups of VOCCA and VOCCH exhibited more significant hypoglycemic activities (P < 0.01). After treated with VOCCH, the oral glucose tolerance of high-dose group of VOCCH was improved compared with model control group (P < 0.05).
Myc activation has been implicated in the pathogenesis of hepatoblastoma (HB), a rare embryonal neoplasm derived from liver progenitor cells. Here, microRNA (miR) expression profiling of 65 HBs evidenced differential patterns related to developmental stage and Myc activity. Undifferentiated aggressive HBs overexpressed the miR-371-3 cluster with concomitant down-regulation of the miR-100/let-7a-2/miR-125b-1 cluster, evoking an ES cell expression profile. ChIP and Myc inhibition assays in hepatoma cells demonstrated that both miR clusters are regulated by Myc in an opposite manner. We show that the two miR clusters exert antagonistic effects on cell proliferation and tumorigenicity. Moreover, their combined deregulation cooperated in modulating the hepatic tumor phenotype, implicating stem cell-like regulation of Myc-dependent miRs in poorly differentiated HBs. Importantly, a four-miR signature representative of these clusters efficiently stratified HB patients, and when applied to 241 hepatocellular carcinomas (HCCs), it identified invasive tumors with a poor prognosis. Our data argue that Myc-driven reprogramming of miR expression patterns contributes to the aggressive phenotype of liver tumors originating from hepatic progenitor cells.
Cathelicidins were initially characterized as a family of antimicrobial peptides. Now it is clear that they fulfill several immune functions in addition to their antimicrobial activity. In the current work, three cDNA sequences encoding pheasant cathelicidins were cloned from a constructed bone marrow cDNA library of Phasianus colchicus, using a nested-PCR-based cloning strategy. The three deduced mature antimicrobial peptides, Pc-CATH1, 2 and 3 are composed of 26, 32, and 29 amino acid residues, respectively. Unlike the mammalian cathelicidins that are highly divergent even within the same genus, Pc-CATHs are remarkably conserved with chicken fowlicidins with only a few of residues mutated according to the phylogenetic analysis result. Synthetic Pc-CATH1 exerted strong antimicrobial activity against most of bacteria and fungi tested, including the clinically isolated (IS) drug-resistant strains. Most MIC values against Gram-positive bacteria were in the range of 0.09-2.95 ?M in the presence of 100mM NaCl. Pc-CATH1 displayed a negligible hemolytic activity against human erythrocytes, lysing 3.6% of erythrocytes at 3.15 ?M (10 ?g/ml), significantly higher than the corresponding MIC. Pc-CATH1 was stable in the human serum for up to 72 h, revealing its extraordinary serum stability. These specific features of Pc-CATH1 may make its applications much wider given the potency and breadth of the peptides bacteriocidal capacity and its resistance towards serum and high-salt environments.
Tau aggregation is a hallmark of several neurodegenerative diseases, including AD (Alzheimers disease), although the mechanism underlying tau aggregation remains unclear. Recent studies show that the proteolysis of tau plays an important role in both tau aggregation and neurodegeneration. On one hand, truncation of tau may generate amyloidogenic tau fragments that initiate the aggregation of tau, which in turn can cause toxicity. On the other hand, truncation of tau may result in tau fragments which induce neurodegeneration through unknown mechanisms, independently of tau aggregation. Blocking the truncation of tau thus may represent a promising therapeutic approach for AD or other tauopathies. In the present paper, we summarize our data on tau cleavage in a cell model of tauopathy and major results on tau cleavage reported in the literature.
Tissue samples from many diseases have been used for gene expression profiling studies, but these samples often vary widely in the cell types they contain. Such variation could confound efforts to correlate expression with clinical parameters. In principle, the proportion of each major tissue component can be estimated from the profiling data and used to triage samples before studying correlations with disease parameters. Four large gene expression microarray data sets from prostate cancer, whose tissue components were estimated by pathologists, were used to test the performance of multivariate linear regression models for in silico prediction of major tissue components. Ten-fold cross-validation within each data set yielded average differences between the pathologists predictions and the in silico predictions of 8% to 14% for the tumor component and 13% to 17% for the stroma component. Across independent data sets that used similar platforms and fresh frozen samples, the average differences were 11% to 12% for tumor and 12% to 17% for stroma. When the models were applied to 219 arrays of "tumor-enriched" samples in the literature, almost one quarter were predicted to have 30% or less tumor cells. Furthermore, there was a 10.5% difference in the average predicted tumor content between 37 recurrent and 42 nonrecurrent cancer patients. As a result, genes that correlated with tissue percentage generally also correlated with recurrence. If such a correlation is not desired, then some samples might be removed to rebalance the data set or tissue percentages might be incorporated into the prediction algorithm. A web service, "CellPred," has been designed for the in silico prediction of sample tissue components based on expression data.
Epigenetic changes play a crucial role in leukemogenesis. HDACs are frequently recruited to target gene promoters by balanced translocation derived oncogenic fusion proteins. As important epigenetic effector mechanisms, histone deacetylases (HDAC) have emerged as potential therapeutic targets. However, the patterns of HDAC1 localization and the role of HDACs in leukemia pathogenesis remain to be elucidated. Using ChIP-Chip analyses we analyzed HDAC1 deposition patterns at more than 10,000 gene promoters in a large cohort of leukemia patients and CD34+ controls. HDAC1 binding was significantly increased in AML blasts compared to CD34+ progenitor cells at 130 gene promoters whereas decreased binding was observed at 66 gene promoters. Distinct HDAC1 binding patterns occurred in AML subtypes with balanced translocations t(15;17), t(8;21) and inv(16). In addition, a more generalized signature was established, that revealed an AML specific pattern of HDAC1 distribution. Many of the HDAC1-binding altered promoters regulate genes involved in hematopoiesis, transcriptional regulation and signal transduction. HDAC1 binding patterns were associated with patients event free survival. This is the first study to determine HDAC1 modification patterns in a large number of AML and ALL specimens. Our findings suggest that dyslocalization of HDAC1 is a common feature in AML. Importantly, HDAC1 modifications possess prognostic power for patient survival. Our findings suggest that altered HDAC1 localization is an explanation for the observed benefit of HDAC inhibitors in AML therapy.
The amyloid cascade hypothesis of Alzheimers disease (AD) posits that the generation of ?-amyloid (A?) triggers Tau neurofibrillary pathology. Recently a "17 kD" calpain-induced Tau fragment, comprising residues 45-230 (molecular weight [MW], 18.7 kD), was proposed to mediate A?-induced toxicity. Here, we demonstrate that the "17 kD" fragment is actually much smaller, containing residues 125-230 (molecular weight, 10.7 kD). Inducing Tau phosphorylation by okadaic acid or mimicking phosphorylation by Glu mutations at the epitopes of Alzheimer-diagnostic antibodies AT100/AT8/PHF1 could not prevent the generation of this fragment. The fragment can be induced not only by A? oligomers, but also by other cell stressors, e.g., thapsigargin (a Ca(2+)-ATPase inhibitor) or glutamate (an excitatory neurotransmitter). However, overexpression of neither Tau(45-230) nor Tau(125-230) fragment is toxic to Chinese hamster ovary (CHO) cells, neuroblastoma cells (N2a) or primary hippocampal neurons. Finally, the calpain-induced fragment can be observed both in Alzheimers disease brains and in control normal human brains. We conclude that the 17 kD Tau fragment is not a mediator of A?-induced toxicity, leaving open the possibility that upstream calpain activation might cause both Tau fragmentation and toxicity.
Alterations in DNA methylation offer unique prospects as tumor markers. The big limitation in cervical cancer research is that it is too hard to obtain the pure normal tissue from a cervical cancer mass. So, we first profile type-specific DNA methylation of major two types of human uterine cervical cancer, adenocarcinoma (ACA) and squamous cell carcinoma (SCC), to establish a precise source of marker research. To assess the DNA methylation status of promoter regions in human uterine cervical ACAs and SCCs, fresh frozen tissues were obtained from bulky tumor masses to minimize the contamination from normal tissues and two array platforms using digestion with methylation-sensitive restriction-enzyme HpaII, ligation, and PCR were performed: an array of 11,994 (approximately 1.5 kb) PCR products from 10,445 promoter regions, and an array of 355,264 oligonucleotides for 18,212 HpaII fragments in 12,617 promoter regions. Loci near 21 genes showed significant differences between six ACA and four SCC from the analysis of two array data. Real-time PCR-based validation was performed on 13 loci using other nearby candidate methylation targets in the same promoter. Methylation patterns of 11 of 13 linked loci concurred with the microarray results. Four loci were further studied using tissues from additional patients (23 ACA and 24 SCC). Hypermethylation of loci in PAK6 and NOGOR most strongly correlated with ACA. Therefore, we have identified the 21 genes with differential methylation pattern between ACA and SCC and, furthermore, we found that PAK6 and NOGOR could be useful markers of ACA to be distinct from SCC.
Acute myeloid leukemia (AML) is commonly associated with alterations in transcription factors because of altered expression or gene mutations. These changes might induce leukemia-specific patterns of histone modifications. We used chromatin-immunoprecipitation on microarray to analyze histone 3 lysine 9 trimethylation (H3K9me3) patterns in primary AML (n = 108), acute lymphoid leukemia (n = 28), CD34(+) cells (n = 21) and white blood cells (n = 15) specimens. Hundreds of promoter regions in AML showed significant alterations in H3K9me3 levels. H3K9me3 deregulation in AML occurred preferentially as a decrease in H3K9me3 levels at core promoter regions. The altered genomic regions showed an overrepresentation of cis-binding sites for ETS and cyclic adenosine monophosphate response elements (CREs) for transcription factors of the CREB/CREM/ATF1 family. The decrease in H3K9me3 levels at CREs was associated with increased CRE-driven promoter activity in AML blasts in vivo. AML-specific H3K9me3 patterns were not associated with known cytogenetic abnormalities. But a signature derived from H3K9me3 patterns predicted event-free survival in AML patients. When the H3K9me3 signature was combined with established clinical prognostic markers, it outperformed prognosis prediction based on clinical parameters alone. These findings demonstrate widespread changes of H3K9me3 levels at gene promoters in AML. Signatures of histone modification patterns are associated with patient prognosis in AML.
In the present study, EA-CATH1 and EA-CATH2 were identified from a constructed lung cDNA library of donkey (Equus asinus) as members of cathelicidin-derived antimicrobial peptides, using a nested PCR-based cloning strategy. Composed of 25 and 26 residues, respectively, EA-CATH1 and EA-CATH2 are smaller than most other cathelicidins and have no sequence homology to other cathelicidins identified to date. Chemically synthesized EA-CATH1 exerted potent antimicrobial activity against most of the 32 strains of bacteria and fungi tested, especially the clinically isolated drug-resistant strains, and minimal inhibitory concentration values against Gram-positive bacteria were mostly in the range of 0.3-2.4 microg mL(-1). EA-CATH1 showed an extraordinary serum stability and no haemolytic activity against human erythrocytes in a dose up to 20 microg mL(-1). CD spectra showed that EA-CATH1 mainly adopts an alpha-helical conformation in a 50% trifluoroethanol/water solution, but a random coil in aqueous solution. Scanning electron microscope observations of Staphylococcus aureus (ATCC2592) treated with EA-CATH1 demonstrated that EA-CATH could cause rapid disruption of the bacterial membrane, and in turn lead to cell lysis. This might explain the much faster killing kinetics of EA-CATH1 than conventional antibiotics revealed by killing kinetics data. In the presence of CaCl(2), EA-CATH1 exerted haemagglutination activity, which might potentiate an inhibition against the bacterial polyprotein interaction with the host erythrocyte surface, thereby possibly restricting bacterial colonization and spread.
Most current microarray oligonucleotide probe design strategies are based on probe design factors (PDFs), which include probe hybridization free energy (PHFE), probe minimum folding energy (PMFE), dimer score, hairpin score, homology score and complexity score. The impact of these PDFs on probe performance was evaluated using four sets of microarray comparative genome hybridization (aCGH) data, which included two array manufacturing methods and the genomes of two species. Since most of the hybridizing DNA is equimolar in CGH data, such data are ideal for testing the general hybridization properties of almost all candidate oligonucleotides. In all our data sets, PDFs related to probe secondary structure (PMFE, hairpin score and dimer score) are the most significant factors linearly correlated with probe hybridization intensities. PHFE, homology and complexity score are correlating significantly with probe specificities, but in a non-linear fashion. We developed a new PDF, pseudo probe binding energy (PPBE), by iteratively fitting dinucleotide positional weights and dinucleotide stacking energies until the average residue sum of squares for the model was minimized. PPBE showed a better correlation with probe sensitivity and a better specificity than all other PDFs, although training data are required to construct a PPBE model prior to designing new oligonucleotide probes. The physical properties that are measured by PPBE are as yet unknown but include a platform-dependent component. A practical way to use these PDFs for probe design is to set cutoff thresholds to filter out bad quality probes. Programs and correlation parameters from this study are freely available to facilitate the design of DNA microarray oligonucleotide probes.
Ticks are blood-feeding arthropods that may secrete immunosuppressant molecules, which inhibit host inflammatory and immune responses and provide survival advantages to pathogens at tick bleeding sites in hosts. In the current work, two families of immunoregulatory peptides, hyalomin-A and -B, were first identified from salivary glands of hard tick Hyalomma asiaticum asiaticum. Three copies of hyalomin-A are encoded by an identical gene and released from the same protein precursor. Both hyalomin-A and -B can exert significant anti-inflammatory functions, either by directly inhibiting host secretion of inflammatory factors such as tumor necrosis factor-alpha, monocyte chemotectic protein-1, and interferon-gamma or by indirectly increasing the secretion of immunosuppressant cytokine of interleukin-10. Hyalomin-A and -B were both found to potently scavenge free radical in vitro in a rapid manner and inhibited adjuvant-induced inflammation in mouse models in vivo. The JNK/SAPK subgroup of the MAPK signaling pathway was involved in such immunoregulatory functions of hyalomin-A and -B. These results showed that immunoregulatory peptides of tick salivary glands suppress host inflammatory response by modulating cytokine secretion and detoxifying reactive oxygen species.
Aggregation and cleavage are two hallmarks of Tau pathology in Alzheimer disease (AD), and abnormal fragmentation of Tau is thought to contribute to the nucleation of Tau paired helical filaments. Clearance of the abnormally modified protein could occur by the ubiquitin-proteasome and autophagy-lysosomal pathways, the two major routes for protein degradation in cells. There is a debate on which of these pathways contributes to clearance of Tau protein and of the abnormal Tau aggregates formed in AD. Here, we demonstrate in an inducible neuronal cell model of tauopathy that the autophagy-lysosomal system contributes to both Tau fragmentation into pro-aggregating forms and to clearance of Tau aggregates. Inhibition of macroautophagy enhances Tau aggregation and cytotoxicity. The Tau repeat domain can be cleaved near the N terminus by a cytosolic protease to generate the fragment F1. Additional cleavage near the C terminus by the lysosomal protease cathepsin L is required to generate Tau fragments F2 and F3 that are highly amyloidogenic and capable of seeding the aggregation of Tau. We identify in this work that components of a selective form of autophagy, chaperone-mediated autophagy, are involved in the delivery of cytosolic Tau to lysosomes for this limited cleavage. However, F1 does not fully enter the lysosome but remains associated with the lysosomal membrane. Inefficient translocation of the Tau fragments across the lysosomal membrane seems to promote formation of Tau oligomers at the surface of these organelles which may act as precursors of aggregation and interfere with lysosomal functioning.
The etiology of adolescent idiopathic scoliosis is undetermined despite years of research. A number of hypotheses have been postulated to explain its development, including growth abnormalities. The irregular expression of growth hormone and insulin-like growth factor-1 (IGF-1) may disturb hormone metabolism, result in a gross asymmetry, and promote the progress of adolescent idiopathic scoliosis. Initial association studies in complex diseases have demonstrated the power of candidate gene association. Prior to our study, 1 study in this field had a negative result. A replicable study is vital for reliability. To determine the relationship of growth hormone receptor and IGF-1 genes with adolescent idiopathic scoliosis, a population-based association study was performed. Single nucleotide polymorphisms with potential function were selected from candidate genes and a distribution analysis was performed. A conclusion was made confirming the insufficiency of an association between adolescent idiopathic scoliosis and the single-nucleotide polymorphism of the growth hormone receptor and IGF-1 genes in Han Chinese.
Cross-platform microarray analysis is an increasingly important research tool, but researchers still lack open source tools for storing, integrating and analyzing large amounts of microarray data obtained from different array platforms.
Blood-feeding arthropods rely heavily on the pharmacological properties of their saliva to get a blood meal and suppress immune reactions of hosts. Little information is available on antihemostatic substances in horsefly salivary glands although their saliva has been thought to contain wide range of physiologically active molecules. In traditional Eastern medicine, horseflies are used as anti-thrombosis material for hundreds of years. By proteomics coupling transcriptome analysis with pharmacological testing, several families of proteins or peptides, which exert mainly on anti-thrombosis functions, were identified and characterized from 60,000 pairs of salivary glands of the horsefly Tabanus yao Macquart (Diptera, Tabanidae). They are: (I) ten fibrin(ogen)olytic enzymes, which hydrolyze specially alpha chain of fibrin(ogen) and are the first family of fibrin(ogen)olytic enzymes purified and characterized from arthropods; (II) another fibrin(ogen)olytic enzyme, which hydrolyzes both alpha and beta chain of fibrin(ogen); (III) ten Arg-Gly-Asp-motif containing proteins acting as platelet aggregation inhibitors; (IV) five thrombin inhibitor peptides; (V) three vasodilator peptides; (VI) one apyrase acting as platelet aggregation inhibitor; (VII) one peroxidase with both platelet aggregation inhibitory and vasodilator activities. The first three families are belonging to antigen five proteins, which show obvious similarity with insect allergens. They are the first members of the antigen 5 family found in salivary glands of blood sucking arthropods to have anti-thromobosis function. The current results imply a possible evolution from allergens of blood-sucking insects to anti-thrombosis agents. The extreme diversity of horsefly anti-thrombosis components also reveals the anti-thrombosis molecular mechanisms of the traditional Eastern medicine insect material.
Much attention has been paid on amphibian peptides for their wide-ranging pharmacological properties, clinical potential, and gene-encoded origin. More than 300 antimicrobial peptides (AMPs) from amphibians have been studied. Peptidomics and genomics analysis combined with functional test including microorganism killing, histamine-releasing, and mast cell degranulation was used to investigate antimicrobial peptide diversity. Thirty-four novel AMPs from skin secretions of Rana nigrovittata were identified in current work, and they belong to 9 families, including 6 novel families. Other three families are classified into rugosin, gaegurin, and temporin family of amphibian AMP, respectively. These AMPs share highly conserved preproregions including signal peptides and spacer acidic peptides, while greatly diversified on mature peptides structures. In this work, peptidomics combined with genomics analysis was confirmed to be an effective way to identify amphibian AMPs, especially novel families. Some AMPs reported here will provide leading molecules for designing novel antimicrobial agents.
Classification and regression trees have long been used for cancer diagnosis and prognosis. Nevertheless, instability and variable selection bias, as well as overfitting, are well-known problems of tree-based methods. In this article, we investigate whether ensemble tree classifiers can ameliorate these difficulties, using data from two recent studies of radical prostatectomy in prostate cancer.
Lipid membranes structurally define the outer surface and internal organelles of cells. The multitude of proteins embedded in lipid bilayers are clearly functionally important, yet they remain poorly defined. Even today, integral membrane proteins represent a special challenge for current large scale shotgun proteomics methods. Here we used endothelial cell plasma membranes isolated directly from lung tissue to test the effectiveness of four different mass spectrometry-based methods, each with multiple replicate measurements, to identify membrane proteins. In doing so, we substantially expanded this membranome to 1,833 proteins, including >500 lipid-embedded proteins. The best method combined SDS-PAGE prefractionation with trypsin digestion of gel slices to generate peptides for seamless and continuous two-dimensional LC/MS/MS analysis. This three-dimensional separation method outperformed current widely used two-dimensional methods by significantly enhancing protein identifications including single and multiple pass transmembrane proteins; >30% are lipid-embedded proteins. It also profoundly improved protein coverage, sensitivity, and dynamic range of detection and substantially reduced the amount of sample and the number of replicate mass spectrometry measurements required to achieve 95% analytical completeness. Such expansion in comprehensiveness requires a trade-off in heavy instrument time but bodes well for future advancements in truly defining the ever important membranome with its potential in network-based systems analysis and the discovery of disease biomarkers and therapeutic targets. This analytical strategy can be applied to other subcellular fractions and should extend the comprehensiveness of many future organellar proteomics pursuits.
The polysaccharides in Jerusalem artichoke (JA) carry a substantial amount of energy that can be partly accessed through bioconversion into storable fuels. We review the potential for converting inulin into a variety of high value-added biorefinery products, including biofuels and biochemicals, and consider the feasibility of regarding JA as a model species of an inulin-rich crop. We discuss feedstock pretreatment, microorganisms used during fermentation, biorefinery products derived from JA, and how to enhance the economic competitiveness of JA as an energy crop.
Cyanobacteria are photoautotrophic prokaryotes with wide variations in genome sizes and ecological habitats. Peroxiredoxin (PRX) is an important protein that plays essential roles in protecting own cells against reactive oxygen species (ROS). PRXs have been identified from mammals, fungi and higher plants. However, knowledge on cyanobacterial PRXs still remains obscure. With the availability of 37 sequenced cyanobacterial genomes, we performed a comprehensive comparative analysis of PRXs and explored their diversity, distribution, domain structure and evolution.
Cerebral infarction has become one of the leading diseases and a major mortality factor around the world. Atherosclerosis is recognized as one of the important causes of ischemic stroke. Recently, accumulating evidences have indicated that the anti-inflammatory and anti-apoptotic functions of the HSP70 family play an important role in cerebral ischemia. However, the association between HSP70 SNPs and ischemic stroke was also not well established. We chose 101 cases of cerebral ischemia and 100 healthy people from the Chinese Han population as our study subjects, and PCR-RFLP was employed to analyze HSP70 polymorphisms: HSP70-1+190G/C, HSP70-2+1267A/G and HSP70-hom+2437T/C. There were no significant differences in +1267A/G allele or genotype frequencies between patients with stroke and healthy controls. However, genotypes of +190CG and +2437TT were differentially distributed between the patients and controls. A significant difference of T allele distribution in the HSP70-hom+2437T/C site was observed. Logistic regression analysis indicated that genotypes of +190CG, +2437TT and T allele in HSP70-hom were risk factors of ischemic stroke. Moreover, the study has formulated that the interactions between hypertension and +190CG or +2437TT may increase the risks of ischemic stroke. The results from this study have suggested a clinical indicator for assessing the possibilities of cerebral stroke, and supply basis to clinicians to give precaution to people who are at risk of stroke.
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