Background and Objective
Of three first-line anti-tuberculosis (anti-TB) drugs, isoniazid is most commonly associated with hepatotoxicity. Differences in INH-induced toxicity have been attributed to genetic variability at several loci, NAT2, CYP2E1, GSTM1and GSTT1, that code for drug-metabolizing enzymes. This study evaluated whether the polymorphisms in these enzymes were associated with an increased risk of anti-TB drug-induced hepatitis in patients and could potentially be used to identify patients at risk of liver injury.
Methods and Design
In a cross-sectional study, 2244 tuberculosis patients were assessed two months after the start of treatment. Anti-TB drug-induced liver injury (ATLI) was defined as an ALT, AST or bilirubin value more than twice the upper limit of normal. NAT2, CYP2E1, GSTM1 and GSTT1 genotypes were determined using the PCR/ligase detection reaction assays.
2244 patients were evaluated, there were 89 cases of ATLI, a prevalence of 4% 9 patients (0.4%) had ALT levels more than 5 times the upper limit of normal. The prevalence of ATLI was greater among men than women, and there was a weak association with NAT2*5 genotypes, with ATLI more common among patients with the NAT2*5*CT genotype. The sensitivity of the CT genotype for identifying patients with ATLI was 42% and the positive predictive value 5.9%. CT ATLI was more common among slow acetylators (prevalence ratio 2.0 (95% CI 0.95,4.20) )compared to rapid acetylators. There was no evidence that ATLI was associated with CYP2E1 RsaIc1/c1genotype, CYP2E1 RsaIc1/c2 or c2/c2 genotypes, or GSTM1/GSTT1 null genotypes.
In Xinjiang Uyghur TB patients, liver injury was associated with the genetic variant NAT2*5, however the genetic markers studied are unlikely to be useful for screening patients due to the low sensitivity and low positive predictive values for identifying persons at risk of liver injury.
Sterol C24-methyltransferases (SMTs) constitute a group of sequence-related proteins that catalyze the pattern of sterol diversity across eukaryotic kingdoms. The only gene for sterol alkylation in green algae was identified and the corresponding catalyst from Chlamydomonas reinhardtii (Cr) was characterized kinetically and for product distributions. The properties of CrSMT were similar to those predicted for an ancient SMT expected to possess broad C3-anchoring requirements for substrate binding and formation of 24β-methyl/ethyl Δ25(27)-olefin products typical of primitive organisms. Unnatural Δ24(25)-sterol substrates, missing a C4β-angular methyl group involved with binding orientation, convert to product ratios in favor of Δ24(28)-products. Remodeling the active site to alter the electronics of Try110 (to Leu) results in delayed timing of the hydride migration from methyl attack of the Δ24-bond, that thereby produces metabolic switching of product ratios in favor of Δ25(27)-olefins or impairs the second C1-transfer activity. Incubation of [27-13C]lanosterol or [methyl-2H3]SAM as co-substrates established the CrSMT catalyzes a sterol methylation pathway by the “algal” Δ25(27)-olefin route, where methylation proceeds by a conserved SN2 reaction and de-protonation proceeds from the pro-Z methyl group on lanosterol corresponding to C27. This previously unrecognized catalytic competence for an enzyme of sterol biosynthesis, together with phylogenomic analyses, suggest that mutational divergence of a promiscuous SMT produced substrate- and phyla-specific SMT1 (catalyzes first biomethylation) and SMT2 (catalyzes second biomethylation) isoforms in red and green algae, respectively, and in the case of SMT2 selection afforded modification in reaction channeling necessary for the switch in ergosterol (24β-methyl) biosynthesis to stigmasterol (24α-ethyl) biosynthesis during the course of land plant evolution.
Chlamydmonas reinhardtii green algae; Sterol evolution; Sterol C24-methyltransferase; Ergosterol; Cholesterol; SMT2; SMT1
Polo-like kinase 1 (Plk1) is pivotal for proper mitotic progression, its targeting activity is regulated by precise subcellular positioning and phosphorylation. Here we assessed the protein expression, subcellular localization and possible functions of phosphorylated Plk1 (pPlk1Ser137 and pPlk1Thr210) in mouse oocytes during meiotic division. Western blot analysis revealed a peptide of pPlk1Ser137 with high and stable expression from germinal vesicle (GV) until metaphase II (MII), while pPlk1Thr210 was detected as one large single band at GV stage and 2 small bands after germinal vesicle breakdown (GVBD), which maintained stable up to MII. Immunofluorescence analysis showed pPlk1Ser137 was colocalized with microtubule organizing center (MTOC) proteins, γ-tubulin and pericentrin, on spindle poles, concomitantly with persistent concentration at centromeres and dynamic aggregation between chromosome arms. Differently, pPlk1Thr210 was persistently distributed across the whole body of chromosomes after meiotic resumption. The specific Plk1 inhibitor, BI2536, repressed pPlk1Ser137 accumulation at MTOCs and between chromosome arms, consequently disturbed γ-tubulin and pericentrin recruiting to MTOCs, destroyed meiotic spindle formation, and delayed REC8 cleavage, therefore arresting oocytes at metaphase I (MI) with chromosome misalignment. BI2536 completely reversed the premature degradation of REC8 and precocious segregation of chromosomes induced with okadaic acid (OA), an inhibitor to protein phosphatase 2A. Additionally, the protein levels of pPlk1Ser137 and pPlk1Thr210, as well as the subcellular distribution of pPlk1Thr210, were not affected by BI2536. Taken together, our results demonstrate that Plk1 activity is required for meiotic spindle assembly and REC8 cleavage, with pPlk1Ser137 is the action executor, in mouse oocytes during meiotic division.
mouse; MTOC; oocytes; Plk1 phosphorylation; REC8 cleavage; spindle formation
Haspin-catalyzed histone H3 threonine 3 (Thr3) phosphorylation facilitates chromosomal passenger complex (CPC) docking at centromeres, regulating indirectly chromosome behavior during somatic mitosis. It is not fully known about the expression and function of H3 with phosphorylated Thr3 (H3T3-P) during meiosis in oocytes. In this study, we investigated the expression and sub-cellular distribution of H3T3-P, as well as its function in mouse oocytes during meiotic division. Western blot analysis revealed that H3T3-P expression was only detected after germinal vesicle breakdown (GVBD), and gradually increased to peak level at metaphase I (MI), but sharply decreased at metaphase II (MII). Immunofluorescence showed H3T3-P was only brightly labeled on chromosomes after GVBD, with relatively high concentration across the whole chromosome axis from pro-metaphase I (pro-MI) to MI. Specially, H3T3-P distribution was exclusively limited to the local space between sister centromeres at MII stage. Haspin inhibitor, 5-iodotubercidin (5-ITu), dose- and time-dependently blocked H3T3-P expression in mouse oocytes. H3T3-P inhibition delayed the resumption of meiosis (GVBD) and chromatin condensation. Moreover, the loss of H3T3-P speeded up the meiotic transition to MII of pro-MI oocytes in spite of the presence of non-aligned chromosomes, even reversed MI-arrest induced with Nocodazole. The inhibition of H3T3-P expression distinguishably damaged MAD1 recruitment on centromeres, which indicates the spindle assembly checkpoint was impaired in function, logically explaining the premature onset of anaphase I. Therefore, Haspin-catalyzed histone H3 phosphorylation is essential for chromatin condensation and the following timely transition from meiosis I to meiosis II in mouse oocytes during meiotic division.
GVBD; H3T3-P; meiotic progression; MAD1; oocytes
Automatic and accurate segmentation of the prostate and rectum in planning CT images is a challenging task due to low image contrast, unpredictable organ (relative) position, and uncertain existence of bowel gas across different patients. Recently, regression forest was adopted for organ deformable segmentation on 2D medical images by training one landmark detector for each point on the shape model. However, it seems impractical for regression forest to guide 3D deformable segmentation as a landmark detector, due to large number of vertices in the 3D shape model as well as the difficulty in building accurate 3D vertex correspondence for each landmark detector. In this paper, we propose a novel boundary detection method by exploiting the power of regression forest for prostate and rectum segmentation. The contributions of this paper are as follows: 1) we introduce regression forest as a local boundary regressor to vote the entire boundary of a target organ, which avoids training a large number of landmark detectors and building an accurate 3D vertex correspondence for each landmark detector; 2) an auto-context model is integrated with regression forest to improve the accuracy of the boundary regression; 3) we further combine a deformable segmentation method with the proposed local boundary regressor for the final organ segmentation by integrating organ shape priors. Our method is evaluated on a planning CT image dataset with 70 images from 70 different patients. The experimental results show that our proposed boundary regression method outperforms the conventional boundary classification method in guiding the deformable model for prostate and rectum segmentations. Compared with other state-of-the-art methods, our method also shows a competitive performance.
CT image; prostate segmentation; rectum segmentation; regression forest; local boundary regression; deformable segmentation
Platelet glycoprotein Ib‐IX complex is affixed to the membrane skeleton through interaction with actin binding protein 280 (ABP‐280). We find that removal of the ABP‐280 binding sites in GP Ibα cytoplasmic tail has little impact on the complex clustering induced by antibody crosslinking. However, large truncation of the GP Ibα cytoplasmic tail allows the formation of larger patches of the complex, suggesting that an ABP‐280 independent force may exist. Besides, we observe that the signaling upon GP Ib‐IX clustering is elicited in both membrane lipid domain dependent and independent manner, a choice that relies on how the membrane skeleton interacts with the complex. Our findings suggest a more complex mechanism for how the membrane skeleton regulates the GP Ib‐IX function. © 2016 The Authors IUBMB Life published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology, 68(10):823–829, 2016
platelet glycoprotein Ib‐IX complex; membrane skeleton; clustering; membrane lipid domain; signaling
The aim of this meta‐analysis and systematic review of randomized controlled trials (RCTs) was to evaluate the efficacy and safety of adductor canal block (ACB) for early postoperative pain management in patients undergoing total knee arthroplasty (TKA). Relevant manuscripts comparing ACB with saline or femoral nerve block (FNB) in TKA patients were searched for in the databases of PubMed, EMBASE, and Cochrane library. The outcomes assessed included cumulative analgesic consumption, pain at rest or during movement, ability to ambulate, quadriceps strength, and complications (nausea, vomiting or sedation). For continuous outcomes, pooled effects were measured using weighted mean difference (WMD) or standard mean difference (SMD), together with 95% confidence intervals (CIs). For outcomes without sufficient data for synthesis, qualitative interpretation of individual studies was summarized. Finally, 11 RCTs involving 675 patients met the inclusion criteria. The pooled results showed that ACB resulted in less postoperative analgesic consumption than saline (WMD, −12.84 mg; 95% CI, −19.40 mg to −6.27 mg; P < 0.001) and less pain at rest or during activity. No conclusions could be drawn regarding ability to ambulate and quadriceps strength, because only one study reported these variables. Most studies comparing ACB and FNB reported similar effects on postoperative analgesic consumption (WMD, −0.56 mg; 95% CI, −8.05 mg to 6.93 mg; P = 0.884) and pain; however, ability to ambulate and quadriceps strength were significantly better with ACB (SMD, 0.99; 95% CI, 0.04–1.94; P = 0.041). Additionally, ACB did not increase the rate of complications. Our results suggest that, compared with saline, ACB decreases analgesic consumption and offers short‐term advantages in terms of pain relief. Compared with FNB, ACB was associated with better ability to ambulate and quadriceps strength.
Adductor canal block; Analgesia; Meta analysis; Randomized controlled trials; Total knee arthroplasty
Microbial antimonite [Sb(III)] oxidation converts toxic Sb(III) into less toxic antimonate [Sb(V)] and plays an important role in the biogeochemical Sb cycle. Currently, little is known about the mechanisms underlying bacterial Sb(III) resistance and oxidation.
In this study, Tn5 transposon mutagenesis was conducted in the Sb(III)-oxidizing strain Pseudomonas stutzeri TS44 to isolate the genes responsible for Sb(III) resistance and oxidation. An insertion mutation into gshA, encoding a glutamate cysteine ligase involved in glutathione biosynthesis, generated a strain called P. stutzeri TS44-gshA540. This mutant strain was complemented with a plasmid carrying gshA to generate strain P. stutzeri TS44-gshA-C. The transcription of gshA, the two superoxide dismutase (SOD)-encoding genes sodB and sodC as well as the catalase-encoding gene katE was monitored because gshA-encoded glutamate cysteine ligase is responsible for the biosynthesis of glutathione (GSH) and involved in the cellular stress defense system as are superoxide dismutase and catalase responsible for the conversion of ROS. In addition, the cellular content of total ROS and in particular H2O2 was analyzed. Compared to the wild type P. stutzeri TS44 and TS44-gshA-C, the mutant P. stutzeri TS44-gshA540 had a lower GSH content and exhibited an increased content of total ROS and H2O2 and increased the Sb(III) oxidation rate. Furthermore, the transcription of sodB, sodC and katE was induced by Sb(III). A positive linear correlation was found between the Sb(III) oxidation rate and the H2O2 content (R
2 = 0.97), indicating that the accumulated H2O2 is correlated to the increased Sb(III) oxidation rate.
Based on the results, we propose that a disruption of the pathway involved in ROS-protection allowed H2O2 to accumulate. In addition to the previously reported enzyme mediated Sb(III) oxidation, the mechanism of bacterial oxidation of Sb(III) to Sb(V) includes a non-enzymatic mediated step using H2O2 as the oxidant.
Electronic supplementary material
The online version of this article (doi:10.1186/s12866-016-0902-5) contains supplementary material, which is available to authorized users.
Pseudomonas stutzeri; Sb(III) oxidation; H2O2; Transposon mutagenesis; gshA; Reactive oxygen species (ROS)
Homologous recombination-mediated genome engineering has been broadly applied in prokaryotes with high efficiency and accuracy. However, this method is limited in realizing larger-scale genome editing with numerous genes or large DNA fragments because of the relatively complicated procedure for DNA editing template construction. Here, we describe a CRISPR-Cas9 assisted non-homologous end-joining (CA-NHEJ) strategy for the rapid and efficient inactivation of bacterial gene (s) in a homologous recombination-independent manner and without the use of selective marker. Our study show that CA-NHEJ can be used to delete large chromosomal DNA fragments in a single step that does not require homologous DNA template. It is thus a novel and powerful tool for bacterial genomes reducing and possesses the potential for accelerating the genome evolution.
Tobacco stalk is one kind of abundant crop residues in China. The high lignification of tobacco stalk increases its reusing cost and the existing of nicotine will cause serious pollution. The biodegradation of lignocellulosic biomass has been demonstrated to be an environmental and economical approach for the utilization of plant stalk. Meanwhile, many nicotine-degrading microorganisms were found in nature. However, microorganisms which could degraded both nicotine and lignin haven’t been reported. Therefore, it’s imperative to find some suitable microorganisms to break down lignin and simultaneously remove nicotine in tobacco stalk.
The nicotine in tobacco stalk could be degraded effectively by Trametes versicolor, Trametes hirsute and Phanerochaete chrysosporium. The nicotine content in tobacco stalk was lowered to below 500 mg/kg (a safe concentration to environment) after 10 days of fermentation with Phanerochaete chrysosporium and Trametes versicolor, and 15 days with Trametes hirsute. The degradation rate of lignin in the fermented tobacco stalk was 37.70, 51.56 and 53.75% with Trametes versicolor, Trametes hirsute and Phanerochaete chrysosporium, respectively. Meanwhile, 24.28% hemicellulose was degraded by Phanerochaete chrysosporium and 28.19% cellulose was removed by Trametes hirsute. Through the enzyme activity analysis, the main and highest ligninolytic enzymes produced by Phanerochaete chrysosporium, Trametes hirsute and Trametes versicolor were lignin peroxidase (88.62 U · L−1), manganese peroxidase (100.95 U · L−1) and laccase (745.65 U · L−1). Meanwhile, relatively high and stable cellulase activity was also detected during the fermentation with Phanerochaete chrysosporium, and the highest endoglucanase, exoglucanase and filter paper enzyme activities were 0.38 U · mL−1, 0.45 U · mL−1 and 0.35U · mL−1, respectively. Moreover, the products in the fermentation of tobacco stalk with P. chrysosporium were identified with GC-MS, besides the chemicals produced in the degradation of lignin and nicotine, some small molecular valuable chemicals and fatty acid were also detected.
Our study developed a new method for the degradation and detoxification of tobacco stalk by fermentation with white rot fungi Phanerochaete chrysosporium and Trametes hirsute. The different oxidative enzymes and chemical products detected during the degradation indicated a possible pathway for the utilization of tobacco stalk.
Electronic supplementary material
The online version of this article (doi:10.1186/s12896-016-0311-8) contains supplementary material, which is available to authorized users.
Tobacco stalk; Nicotine degradation; Delignification; Phanerochaete chrysosporium; Lignocellulolytic enzymes
The intriguing properties of phosphorene motivate scientists to further explore the structures and properties of phosphorus materials. Here, we report a new allotrope named K4 phosphorus composed of three-coordinated phosphorus atoms in non-layered structure which is not only dynamically and mechanically stable, but also possesses thermal stability comparable to that of the orthorhombic black phosphorus (A17). Due to its unique configuration, K4 phosphorus exhibits exceptional properties: it possesses a band gap of 1.54 eV which is much larger than that of black phosphorus (0.30 eV), and it is stiffer than black phosphorus. The band gap of the newly predicted phase can be effectively tuned by appling hydrostastic pressure. In addition, K4 phosphorus exibits a good light absorption in visible and near ultraviolet region. These findings add additional features to the phosphorus family with new potential applications in nanoelectronics and nanomechanics.
Background. Despite the great achievements in the treatment of advanced-stage ovarian cancer, it is still a severe condition with an unfavorable 5-year survival rate. Statins have been suggested to reduce the risk of several cancers beyond their cholesterol-lowing effects. However, the prognostic significance of statins in patients with advanced-stage ovarian cancer remains controversial. Methods. A retrospective study was performed to evaluate the association between statin intake and overall survival (OS) among patients with advanced-stage ovarian cancer. Patients who underwent cytoreductive surgery followed by courses of intravenous chemotherapy were matched through a propensity score analysis. Results. A total of 60 propensity-matched patients were included. Women in statin group showed a similar OS than the nonstatin counterparts (P = 0.966), whereas residual tumor was significantly associated with better OS (P = 0.013) and was an independent factor that associated with OS (P = 0.002, hazard ratio = 5.460, and 95% confidence interval: 1.894 to 15.742) in multivariable analysis. Conclusions. Our results suggested that statin usage was not associated with improved OS in patients with advanced-stage ovarian cancer undergoing surgery and chemotherapy. Considering the retrospective nature and the relative small sample size of the study, further prospective studies and random control trials are needed.
It is well established that the RpoN-RpoS sigma factor (σ54-σS) cascade plays an essential role in differential gene expression during the enzootic cycle of Borrelia burgdorferi, the causative agent of Lyme disease. The RpoN-RpoS pathway is activated by the response regulator/σ54-dependent activator (also called bacterial enhancer-binding protein [bEBP]) Rrp2. One unique feature of Rrp2 is that this activator is essential for cell replication, whereas RpoN-RpoS is dispensable for bacterial growth. How Rrp2 controls cell replication, a function that is independent of RpoN-RpoS, remains to be elucidated. In this study, by generating a series of conditional rrp2 mutant strains, we demonstrated that the N-terminal receiver domain of Rrp2 is required for spirochetal growth. Furthermore, a D52A point mutation at the phosphorylation site within the N terminus of Rrp2 abolished cell replication. Mutation of the ATPase motif within the central domain of Rrp2 did not affect spirochetal replication, indicating that phosphorylation-dependent ATPase activity of Rrp2 for σ54 activation is not required for cell growth. However, deletion of the C-terminal domain or a 16-amino-acid truncation of the helix-turn-helix (HTH) DNA-binding motif within the C-terminal domain of Rrp2 abolished spirochetal replication. It was shown that constitutive expression of rpoS is deleterious to borrelial growth. We showed that the essential nature of Rrp2 is not due to an effect on rpoS. These data suggest that phosphorylation-dependent oligomerization and DNA binding of Rrp2 likely function as a repressor, independently of the activation of σ54, controlling an essential step of cell replication in B. burgdorferi.
IMPORTANCE Bacterial enhancer-binding proteins (bEBPs) are a unique group of transcriptional activators specifically required for σ54-dependent gene transcription. This work demonstrates that the B. burgdorferi bEBP, Rrp2, has an additional function that is independent of σ54, that of its essentiality for spirochetal growth, and such a function is dependent on its N-terminal signal domain and C-terminal DNA-binding domain. These findings expand our knowledge on bEBP and provide a foundation to further study the underlying mechanism of this new function of bEBP.
Epithelial ovarian cancer (EOC) is the most lethal tumor of all gynecologic tumors. There is no curative therapy for EOC thus far. The tumor-homing ability of adult mesenchymal stem cells (MSCs) provide the promising potential to use them as vehicles to transport therapeutic agents to the site of tumor. Meanwhile, studies have showed the intrinsic anti-tumor properties of MSCs against various kinds of cancer, including epithelial ovarian cancer. Human endometrial mesenchymal stem cells (EnSCs) derived from menstrual blood are a novel source for adult MSCs and exert restorative function in some diseases. Whether EnSCs endow innate anti-tumor properties on EOC cells has never been reported. By using tumor-bearing animal model and ex vivo experiments, we found that EnSCs attenuated tumor growth by inducing cell cycle arrest, promoting apoptosis, disturbing mitochondria membrane potential and decreasing pro-angiogenic ability in EOC cells in vitro and/or in vivo. Furthermore, EnSCs decreased AKT phosphorylation and promoted nuclear translocation of Forkhead box O-3a (FoxO3a) in EOC cells. Collectively, our findings elucidated the potential intrinsic anti-tumor properties of EnSCs on EOC cells in vivo and in vitro. This research provides a potential strategy for EnSC-based anti-cancer therapy against epithelial ovarian cancer.
To explore whether polycomb repressor Bmi1 plays an important role in dentin and mandible development homeostasis by maintaining redox balance, 3-week-old Bmi1 gene knockout (Bmi1-/-) mice were treated with the antioxidant N-acetylcysteine (NAC) for 2 weeks in their drinking water and phenotypes of the tooth and mandibles were compared with vehicle-treated Bmi1-/- mice and wild-type mice by radiograph, histochemistry and immunohistochemistry. Alterations of oxidative stress, DNA damage, cell proliferation and cell cycle-related parameters were also examined in mandibles. Results showed that the tooth volume and the dentin sialoprotein immunopositive areas, the cortical thickness, alveolar bone volume, osteoblast number and activity, and mRNA expression levels of Runx2, alkaline phosphatase and type I collagen were all reduced significantly in Bmi1-/- mice compared with their wild-type littermates, whereas these parameters were increased significantly in NAC-treated Bmi1-/- mice compared with vehicle-Bmi1-/- mice, although they were not normalized. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were reduced, DNA damage markers including γ-H2AX and 8-oxoguanine levels were increased, the number of Ki67 positive cells was decreased, whereas protein expression levels of p16, p19, p21, p27 and p53 were up-regulated in mandibles from Bmi1-/- mice compared with those from wild-type mice; alterations of these antioxidative enzyme activities, DNA damage markers, cell proliferation and cell cycle-related parameters were all partially rescued by the treatment with antioxidant NAC in Bmi1 deficient mice. These results demonstrated that Bmi1 deficiency resulted in defects in dentin and alveolar bone formation, while the treatment with antioxidant could improve these defects obviously. Therefore, our results indicate that Bmi1 plays an important role in stimulating dentin formation and alveolar bone formation by maintaining redox homeostasis, preventing DNA damage and inhibiting cyclin-dependent kinase inhibitors.
Antioxidant; Bmi1; DNA damage; dentin; mandible
Risk assessment of congestive heart failure (CHF) is essential for detection, especially helping patients make informed decisions about medications, devices, transplantation, and end-of-life care. The majority of studies have focused on disease detection between CHF patients and normal subjects using short-/long-term heart rate variability (HRV) measures but not much on quantification. We downloaded 116 nominal 24-hour RR interval records from the MIT/BIH database, including 72 normal people and 44 CHF patients. These records were analyzed under a 4-level risk assessment model: no risk (normal people, N), mild risk (patients with New York Heart Association (NYHA) class I-II, P1), moderate risk (patients with NYHA III, P2), and severe risk (patients with NYHA III-IV, P3). A novel multistage classification approach is proposed for risk assessment and rating CHF using the non-equilibrium decision-tree–based support vector machine classifier. We propose dynamic indices of HRV to capture the dynamics of 5-minute short term HRV measurements for quantifying autonomic activity changes of CHF. We extracted 54 classical measures and 126 dynamic indices and selected from these using backward elimination to detect and quantify CHF patients. Experimental results show that the multistage risk assessment model can realize CHF detection and quantification analysis with total accuracy of 96.61%. The multistage model provides a powerful predictor between predicted and actual ratings, and it could serve as a clinically meaningful outcome providing an early assessment and a prognostic marker for CHF patients.
We have previously described the existence of two phenotypically distinct cell subsets in ALK-positive anaplastic large cell lymphoma (ALK + ALCL) based on their differential responsiveness to a Sox2 reporter (SRR2), with reporter-responsive (RR) cells being more tumorigenic and chemoresistant than reporter-unresponsive (RU) cells. However, the regulator(s) of RU/RR dichotomy are not identified. In this study, we aim to delineate the key regulator(s) of RU/RR dichotomy.
JASPER motif match analysis was used to identify the putative factors binding to SRR2 sequence. SRR2 probe pull-down assay and quantitate real-time PCR were performed to analyze the regulation of Sox2 transcriptional activity by MYC. Methylcellulose colony formation assay, chemoresistance to doxorubicin and mouse xenograft study were performed to investigate the biological functions of MYC. PCR array and western blotting were executed to study related signaling pathways that regulate MYC expression. Immunofluorescence and immunohistochemistry assay were initiated to evaluate the expression of MYC and its correlation with its regulator by chi-square test analysis in human primary tumor cells.
We identified MYC as a potential regulator of RU/RR dichotomy. In support of its role, MYC was highly expressed in RR cells compared to RU cells, and inhibition of MYC substantially decreased the Sox2/SRR2 binding, Sox2 transcriptional activity, chemoresistance, and methylcellulose colony formation. In contrast, enforced expression of MYC in RU cells conferred the RR phenotype. The Wnt/β-catenin pathway, a positive regulator of MYC, was highly active in RR but not RU cells. While inhibition of this pathway in RR cells substantially decreased MYC expression and SRR2 reporter activity, experimental activation of this pathway led to the opposite effects in RU cells. Collectively, our results support a model in which a positive feedback loop involving Wnt/β-catenin/MYC and Sox2 contributes to the RR phenotype. In a mouse xenograft model, RU cells stably transfected with MYC showed upregulation of the Wnt/β-catenin/MYC/Sox2 axis and increased tumorigenecity. Correlating with these findings, there was a significant correlation between the expression of active β-catenin and MYC in ALK + ALCL primary tumor cells.
A positive feedback loop involving the Wnt/β-catenin/MYC/Sox2 axis defines a highly tumorigenic cell subset in ALK + ALCL.
Electronic supplementary material
The online version of this article (doi:10.1186/s13045-016-0349-z) contains supplementary material, which is available to authorized users.
Intra-tumoral heterogeneity; MYC; Sox2; Wnt/β-catenin; Cancer stemness; ALK-positive anaplastic large cell lymphoma
A Gram-negative bacterial strain, Comamonas aquatica CJG, absorbs low-density lipoprotein but not high-density lipoprotein in serum. Here, we report its draft genomic sequence of 3,764,434 bp, containing total 3,425 genes, 27% of which encode proteins for metabolism and energy conversion, and it is 30% identical to the genome of Comamonas testosteroni.
By electrospinning of polycaprolactone (PCL) solutions containing N-(benzoylthio)benzamide (NSHD1), a H2S donor, fibrous scaffolds with hydrogen sulfide (H2S) releasing capability (H2S-fibers) are fabricated. The resultant microfibers are capable of releasing H2S upon immersion in aqueous solution containing biological thiols under physiological conditions. The H2S release peaks of H2S-fibers appeared at 2~4 hours, while the peak of donor alone showed at 45 minutes. H2S release half-lives of H2S-fibers were 10–20 times longer than that of donor alone. Furthermore, H2S-fibers can protect cells from H2O2 induced oxidative damage by significantly decreasing the production of intracellular reactive oxygen species (ROS). Finally, we investigated the H2S-fibers application as a wound dressing in vitro. Given that H2S has a broad range of physiological functions, H2S-fibers hold great potential for various biomedical applications.
hydrogen sulfide; controlled release; electrospinning; NSHD1; ischemia reperfusion injury
Porphyromonas gingivalis is a prominent periodontal, and emerging systemic, pathogen that redirects host cell signalling pathways and modulates innate immune responses. In this study, we show that P. gingivalis infection induces the dephosphorylation and activation of forkhead box-O (FOXO)1, 3 and 4 in gingival epithelial cells. In addition, immunofluorescence showed that FOXO1 accumulated in the nucleus of P. gingivalis-infected cells. Quantitative reverse transcription PCR demonstrated that transcription of genes involved in protection against oxidative stress (Cat, Sod2, Prdx3), inflammatory responses (IL1β) and anti-apoptosis (Bcl-6) was induced by P. gingivalis, while small-interfering RNA (siRNA)-mediated knockdown of FOXO1 suppressed the transcriptional activation of these genes. P. gingivalis-induced secretion of interleukin (IL)-1β and inhibition of apoptosis were also impeded by FOXO1 knockdown. Neutralization of reactive oxygen species (ROS) by N-acetyl-l-cysteine blocked the activation of FOXO1 by P. gingivalis and concomitantly suppressed the activation of oxidative stress responses, anti-apoptosis programmes and IL-β production. Inhibition of c-Jun-N-terminal kinase (JNK) either pharmacologically or by siRNA, reduced FOXO1 activation and downstream FOXO1-dependent gene regulation in response to P. gingivalis. The results indicate that P. gingivalis-induced ROS activate FOXO transcription factors through JNK signalling, and that FOXO1 controls oxidative stress responses, inflammatory cytokine production and cell survival. These data position FOXO as an important signalling node in the epithelial cell–P. gingivalis interaction, with particular relevance to cell fate and dysbiotic host responses.
Although some existing epidemiological observations and molecular experiments suggested that brain disorders in the realm of psychiatry may be influenced by immune dysregulation, the degree of genetic overlap between psychiatric disorders and immune disorders has not been well established. We investigated this issue by integrative analysis of genome-wide association studies of 18 complex human traits/diseases (five psychiatric disorders, seven immune disorders, and others) and multiple genome-wide annotation resources (Central nervous system genes, immune-related expression-quantitative trait loci (eQTL) and DNase I hypertensive sites from 98 cell-lines). We detected pleiotropy in 24 of the 35 psychiatric-immune disorder pairs. The strongest pleiotropy was observed for schizophrenia-rheumatoid arthritis with MHC region included in the analysis (p = 3.9 × 10−285), and schizophrenia-Crohns disease with MHC region excluded (p = 1.1 × 10−36). Significant enrichment (>1.4 fold) of immune-related eQTL was observed in four psychiatric disorders. Genomic regions responsible for pleiotropy between psychiatric disorders and immune disorders were detected. The MHC region on chromosome 6 appears to be the most important with other regions, such as cytoband 1p13.2, also playing significant roles in pleiotropy. We also found that most alleles shared between schizophrenia and Crohns disease have the same effect direction, with similar trend found for other disorder pairs, such as bipolar-Crohn’s disease. Our results offer a novel birds-eye view of the genetic relationship and demonstrate strong evidence for pervasive pleiotropy between psychiatric disorders and immune disorders. Our findings might open new routes for prevention and treatment strategies for these disorders based on a new appreciation of the importance of immunological mechanisms in mediating risk of many psychiatric diseases.
GWAS; Pleiotropy; Psychiatric disorder; Immune-mediated disease
5-Aminolevulinic acid (ALA), a nonprotein amino acid involved in tetrapyrrole synthesis, has been widely applied in agriculture, medicine, and food production. Many engineered metabolic pathways have been constructed; however, the production yields are still low. In this study, several 5-aminolevulinic acid synthases (ALASs) from different sources were evaluated and compared with respect to their ALA production capacities in an engineered Corynebacterium glutamicum CgS1 strain that can accumulate succinyl-coenzyme A (CoA). A codon-optimized ALAS from Rhodobacter capsulatus SB1003 displayed the best potential. Recombinant strain CgS1/pEC-SB produced 7.6 g/liter ALA using a mineral salt medium in a fed-batch fermentation mode. Employing two-stage fermentation, 12.46 g/liter ALA was produced within 17 h, with a productivity of 0.73 g/liter/h, in recombinant C. glutamicum. Through overexpression of the heterologous nonspecific ALA exporter RhtA from Escherichia coli, the titer was further increased to 14.7 g/liter. This indicated that strain CgS1/pEC-SB-rhtA holds attractive industrial application potential for the future.
IMPORTANCE In this study, a two-stage fermentation strategy was used for production of the value-added nonprotein amino acid 5-aminolevulinic acid from glucose and glycine in a generally recognized as safe (GRAS) host, Corynebacterium glutamicum. The ALA titer represented the highest in the literature, to our knowledge. This high production capacity, combined with the potential easy downstream processes, made the recombinant strain an attractive candidate for industrial use in the future.
In many aromatic plants including spearmint (Mentha spicata), the sites of secondary metabolite production are tiny specialized structures called peltate glandular trichomes (PGT). Having high commercial values, these secondary metabolites are exploited largely as flavours, fragrances and pharmaceuticals. But, knowledge about transcription factors (TFs) that regulate secondary metabolism in PGT remains elusive. Understanding the role of TFs in secondary metabolism pathway will aid in metabolic engineering for increased yield of secondary metabolites and also the development of new production techniques for valuable metabolites. Here, we isolated and functionally characterized a novel MsYABBY5 gene that is preferentially expressed in PGT of spearmint. We generated transgenic plants in which MsYABBY5 was either overexpressed or silenced using RNA interference (RNAi). Analysis of the transgenic lines showed that the reduced expression of MsYABBY5 led to increased levels of terpenes and that overexpression decreased terpene levels. Additionally, ectopic expression of MsYABBY5 in Ocimum basilicum and Nicotiana sylvestris decreased secondary metabolite production in them, suggesting that the encoded transcription factor is probably a repressor of secondary metabolism.
spearmint; secondary metabolism; terpene; sweet basil; transcription factor; YABBY
The objective of this study is to observe whether cyclosporine A (CsA) inhibits the expression of dectin-1 in human corneal epithelial cells infected with Aspergillus fumigatus (A. fumigatus) and to investigate the molecular mechanisms of the inhibition.
Immortalized human corneal epithelial cells (HCECs) were pretreated with 1,25(OH)2D3 and VDR inhibitor for 1 h, and then they were pretreated with CsA for 12h. After these pretreatments, the HCECs were stimulated with A. fumigatus and curdlan respectively, and the expression of dectin-1 and proinflammatory cytokines (IL-1β and TNF-α) were detected by RT-PCR, western blot and ELISA.
Dectin-1 mRNA and dectin-1 protein expression increased when HCECs were stimulated with A. fumigatus or curdlan, and CsA inhibited the dectin-1 expression both in mRNA and protein levels specifically. Dectin-1 and proinflammatory cytokine expression levels were higher when HCECs were pretreated with VDR inhibitor and CsA compared to pretreatment with CsA alone, while dectin-1 and proinflammatory cytokine levels were lower when HCECs were pretreated with 1,25(OH)2D3 and CsA compared to pretreatment with CsA alone.
These data provide evidence that CsA can inhibit the expression of dectin-1 and proinflammatory cytokines through dectin-1 when HCECs are stimulated by A. fumigatus or curdlan. The active form of vitamin D, 1,25(OH)2D3, and VDR signaling pathway regulate the inhibition of CsA. The inhibition is enhanced by 1,25(OH)2D3, and the VDR inhibitor suppresses the inhibition.
To investigate the expression of the v-raf-1 murine leukemia viral oncogene homolog 1 (Raf-1) and its role in the innate immune response of human corneal epithelial cells (HCECs) infected by Aspergillus fumigatus.
HCECs were cultured in vitro. They were randomly divided into 4 groups, including control group, Aspergillus fumigatus group, GW5074 (an inhibitor of Raf-1) group and Laminarin [an inhibitor of Dendriti-cell-associated C-type lectin 1 (Dectin-1)] group. The protein expression level of total Raf-1 and p-Raf-1was measured by Western blot. The expression of IL-6 and IL-8 mRNA in each group was detected by real-time polymerase chain reaction.
In Aspergillus fumigatus group, total Raf-1 protein levels in HCECs remained unchanged at 5, 15, 30 and 45min after infection, while p-Raf-1 expression was significantly enhanced at 30min after infection compared with control group. However, the expression of p-Raf-1 was apparently declined after treated with GW5074 or Laminarin compared with Aspergillus fumigatus group. The expression levels of IL-6, IL-8 mRNA were significantly increased after stimulation with fumigatus compared with control group. Pre-treated with GW5074 significantly inhibited Aspergillus fumigatus-induced upregulation of IL-8 and IL-6.
Aspergillus fumigatus stimulation can elevate the expression of p-Raf-1 in HCECs in vitro. Dectin-1/Raf-1 signal pathway may play a role on regulating the expression of inflammatory cytokines, including IL-6 and IL-8.
Dectin-1/Raf-1 signal pathway; Aspergillus fumigatus; innate immune; human corneal epithelial cells