Hemodynamic stresses are presumed to play an important role in the development of calcific aortic valve disease (CAVD). The elucidation of the shear stress mechanisms involved in the pathogenesis of CAVD has been hampered by the complexity of the native unsteady and side-specific valvular flow environment. To address this gap, this article describes the design and validation of a novel device to expose leaflet samples to time-dependent side-specific shear stress. The device built on a double cone-and-plate geometry was dimensioned based on our previous single-sided shear stress device that minimizes secondary flow effects inherent to this geometry. A fluid–structure interaction (FSI) model was designed to predict the actual shear stress produced on a tissue sample mounted in the new device. Staining was performed on porcine leaflets conditioned in the new bioreactor to assess endothelial integrity and cellular apoptosis. The FSI results demonstrated good agreement between the target (native) and the actual side-specific shear stress produced on a tissue sample. No significant difference in endothelial integrity and cellular apoptosis was detected between samples conditioned for 96 h and fresh controls. This new device will enable the investigation of valvular response to normal and pathologic hemodynamics and the potential mechano-etiology of CAVD.
Mechanobiology; Signal transduction; Hemodynamics; Flow; Remodeling; Inflammation
Calcific aortic valve disease (CAVD) is an active process presumably triggered by interplays between cardiovascular risk factors, molecular signaling networks and hemodynamic cues. While earlier studies demonstrated that alterations in fluid shear stress (FSS) on the fibrosa could trigger inflammation, the mechanisms of CAVD pathogenesis secondary to side-specific FSS abnormalities are poorly understood. This knowledge could be critical to the elucidation of key CAVD risk factors such as congenital valve defects, aging and hypertension, which are known to generate FSS disturbances. The objective of this study was to characterize ex vivo the contribution of isolated and combined abnormalities in FSS magnitude and frequency to early valvular pathogenesis. The ventricularis and fibrosa of porcine aortic valve leaflets were exposed simultaneously to different combinations of sub-physiologic/physiologic/supra-physiologic levels of FSS magnitude and frequency for 24, 48 and 72 hours in a double cone-and-plate device. Endothelial activation and paracrine signaling were investigated by measuring cell-adhesion molecule (ICAM-1, VCAM-1) and cytokine (BMP-4, TGF-β1) expressions, respectively. Extracellular matrix (ECM) degradation was characterized by measuring the expression and activity of the proteases MMP-2, MMP-9, cathepsin L and cathepsin S. The effect of the FSS treatment yielding the most significant pathological response was examined over a 72-hour period to characterize the time-dependence of FSS mechano-transduction. While cytokine expression was stimulated under elevated FSS magnitude at normal frequency, ECM degradation was stimulated under both elevated FSS magnitude at normal frequency and physiologic FSS magnitude at abnormal frequency. In contrast, combined FSS magnitude and frequency abnormalities essentially maintained valvular homeostasis. The pathological response under supra-physiologic FSS magnitude peaked at 48 hours but was then maintained until the 72-hour time point. This study confirms the sensitivity of valve leaflets to both FSS magnitude and frequency and suggests the ability of supra-physiologic FSS levels or abnormal FSS frequencies to initiate CAVD mechanisms.
Gene amplified in squamous cell carcinoma 1 (GASC1) is a member of Jumonji C-domain containing histone demethylases that play an essential role in affecting chromatin architecture and gene expression. The purpose of this study was to determine the expression features and the clinical significance of GASC1 in esophageal squamous cell carcinoma (ESCC). GASC1 expression was detected on tissue microarrays of ESCC samples in 185 cases using immunohistochemical staining. Strong nuclear staining for GASC1 was observed in a subset of ESCC samples. The nuclear expression of GASC1 was significantly associated with lymph node metastasis (P=0.030) and tumor-node metastasis stages (P=0.013). Kaplan-Meier survival analysis showed a tendency that high expression of GASC1 in the nucleus was associated with poor survival of ESCC patients, with a 5-year survival rate of 26.5%, as compared to 43.7% for patients with GASC1-negative/low expression. Furthermore, multivariate analysis revealed that high expression of GASC1 likely acts as a predictive factor for overall survival of ESCC patients, despite the P-value failing to reach significance (P=0.059). The findings indicate that histone demethylase GASC1 may play an important role in promoting cancer metastasis, and shed new light on the importance of targeting GASC1 to suppress metastatic disease in various tumor types, including ESCC.
Histone demethylase; GASC1; lymph node metastasis; immunohistochemistry; esophageal squamous cell carcinoma
A combination of molecular-targeted cancer imaging and therapy is an emerging strategy to improve cancer diagnosis and minimize the side effects of conventional treatments. Here, we generated a recombinant protein, EC1-GLuc-p53C, by fusing EC1 peptide, an artificial ligand of ErbB2, with Gaussia luciferase (GLuc) and a p53-activating peptide, p53C. EC1-GLuc-p53C was expressed and purified from E. coli BL21. In vitro experiments showed that EC1-GLuc-p53c was stable in luminescent activity and selectively targeted ErbB2-overexpressing BT474 cells for bioluminescence imaging. Moreover, the internalized EC1-GLuc-p53C in BT474 cells exerted its function to reactivate p53 and significantly inhibited cellular proliferation. In tumor-bearing mice, the ErbB2-targeted bioluminescence imaging and therapeutic effect of EC1-GLuc-p53C were also observed specifically in BT474 tumors but not in MCF7 tumors, which does not overexpress ErbB2. Thus, the present study demonstrates EC1-GLuc-p53C to be an effective theranostic reagent targeting ErbB2 for bioluminescence imaging and cancer therapy.
Objective: The expression of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein and its regulation by chemotherapeutics were analyzed in primary acute leukemic cells.
Materials and Methods: Peripheral blood was collected from 16 patients with acute leukemia on days 0, 1, 3, and 5 of chemotherapy. The mononuclear cells were separated from the peripheral blood, and TRAIL expression was assessed by flow cytometry. The bone marrow mononuclear cells of patients with acute leukemia were separated before chemotherapy and cultured in vitro with VP-16 and/or interferon (IFN). The TRAIL expression level was detected after the cell culture.
Results: TRAIL expression in the mononuclear cells of peripheral blood was significantly upregulated on day 1 (p<0.05) and then significantly decreased on day 5 after chemotherapy (p<0.05). Results from the in vitro culture revealed that VP-16 upregulated TRAIL expression in the bone marrow mononuclear cells of patients with acute leukemia, but the binding of VP-16 to IFN did not enhance TRAIL expression as compared with VP-16 alone (p>0.05).
onclusion: OA single chemotherapy mechanism for leukemia may suffice to induce TRAIL expression and promote the apoptosis of leukemic cells.
Conflict of interest:None declared.
TRAIL; Acute leukemia; chemotherapeutics
Macroautophagy (autophagy) is crucial for cell survival during starvation and plays important roles in animal development and human diseases. Molecular understanding of autophagy has mainly come from the budding yeast Saccharomyces cerevisiae, and it remains unclear to what extent the mechanisms are the same in other organisms. Here, through screening the mating phenotype of a genome-wide deletion collection of the fission yeast Schizosaccharomyces pombe, we obtained a comprehensive catalog of autophagy genes in this highly tractable organism, including genes encoding three heretofore unidentified core Atg proteins, Atg10, Atg14, and Atg16, and two novel factors, Ctl1 and Fsc1. We systematically examined the subcellular localization of fission yeast autophagy factors for the first time and characterized the phenotypes of their mutants, thereby uncovering both similarities and differences between the two yeasts. Unlike budding yeast, all three Atg18/WIPI proteins in fission yeast are essential for autophagy, and we found that they play different roles, with Atg18a uniquely required for the targeting of the Atg12–Atg5·Atg16 complex. Our investigation of the two novel factors revealed unforeseen autophagy mechanisms. The choline transporter-like protein Ctl1 interacts with Atg9 and is required for autophagosome formation. The fasciclin domain protein Fsc1 localizes to the vacuole membrane and is required for autophagosome-vacuole fusion but not other vacuolar fusion events. Our study sheds new light on the evolutionary diversity of the autophagy machinery and establishes the fission yeast as a useful model for dissecting the mechanisms of autophagy.
Autophagy is a eukaryotic cellular process that transports cytoplasmic contents into lysosomes/vacuoles for degradation. It has been linked to multiple human diseases, including cancer and neurodegenerative disorders. The molecular machinery of autophagy was first identified and has been best characterized in the budding yeast Saccharomyces cerevisiae, but little is known about the autophagy machinery in another important unicellular model organism, the fission yeast Schizosaccharomyces pombe. In this study, we performed an unbiased and comprehensive screening of the fission yeast autophagy genes by profiling the mating phenotypes of nearly 3000 deletion strains. Following up on the screening results, we systematically characterized both previously known and newly identified fission yeast autophagy factors by examining their localization and the phenotype of their mutants. Our analysis increased the number of experimentally defined fission yeast autophagy factors from 14 to 23, including two novel factors that act in ways different from all previously known autophagy proteins. Together, our data reveal unexpected evolutionary divergence of autophagy mechanisms and establish a new model system for unraveling the molecular details of the autophagy process.
We introduce a technique for measuring the conductivity of individual hybrid metal, semiconducting core-shell and full-metal conducting particles by a microscopic four-point probe (μ-4PP) method. The four-point probe geometry allows for minimizing contact resistances between electrodes and particles. By using a focused ion beam we fabricate platinum nanoleads between four microelectrodes on a silicon chip and an individual particle, and determine the particle's conductivity via sensitive current and voltage measurements. Up to sixteen particles can be taken up by each chip, which allows for multiple conductivity measurements by simply multiplexing the electric contacts connected to a multimeter. Although, for demonstration, we used full Au (conducting) and Ag-coated latex particles (semiconducting) of a few micrometers in diameter, the method can be applied to other types of conducting or semiconducting particles of different diameters.
The Sedum alfredii Hance hyperaccumulating ecotype (HE) has the ability to hyperaccumulate cadmium (Cd), as well as zinc (Zn) and lead (Pb) in above-ground tissues. Although many physiological studies have been conducted with these plants, the molecular mechanisms underlying their hyper-tolerance to heavy metals are largely unknown. Here we report on the generation of 9.4 gigabases of adaptor-trimmed raw sequences and the assembly of 57,162 transcript contigs in S. alfredii Hance (HE) shoots by the combination of Roche 454 and Illumina/Solexa deep sequencing technologies. We also have functionally annotated the transcriptome and analyzed the transcriptome changes upon Cd hyperaccumulation in S. alfredii Hance (HE) shoots. There are 110 contigs and 123 contigs that were up-regulated (Fold Change ≧2.0) and down-regulated (Fold Change ≦0.5) by chronic Cd treatment in S. alfredii Hance (HE) at q-value cutoff of 0.005, respectively. Quantitative RT-PCR was employed to compare gene expression patterns between S. alfredii Hance (HE) and non-hyperaccumulating ecotype (NHE). Our results demonstrated that several genes involved in cell wall modification, metal translocation and remobilization were more induced or constitutively expressed at higher levels in HE shoots than that in NHE shoots in response to Cd exposure. Together, our study provides large-scale expressed sequence information and genome-wide transcriptome profiling of Cd responses in S. alfredii Hance (HE) shoots.
Kras; Pten; Murine model; T-ALL
Kawasaki disease (KD) is a complex disease, leading to the damage of multisystems. The pathogen that triggers this sophisticated disease is still unknown since it was first reported in 1967. To increase our knowledge on the effects of genes in KD, we extracted statistically significant genes so far associated with this mysterious illness from candidate gene studies and genome-wide association studies. These genes contributed to susceptibility to KD, coronary artery lesions, resistance to initial IVIG treatment, incomplete KD, and so on. Gene ontology category and pathways were analyzed for relationships among these statistically significant genes. These genes were represented in a variety of functional categories, including immune response, inflammatory response, and cellular calcium ion homeostasis. They were mainly enriched in the pathway of immune response. We further highlighted the compelling immune pathway of NF-AT signal and leukocyte interactions combined with another transcription factor NF-κB in the pathogenesis of KD. STRING analysis, a network analysis focusing on protein interactions, validated close contact between these genes and implied the importance of this pathway. This data will contribute to understanding pathogenesis of KD.
The bicuspid aortic valve (BAV) is the most common congenital cardiac anomaly and is frequently associated with calcific aortic valve disease (CAVD). The most prevalent type-I morphology, which results from left-/right-coronary cusp fusion, generates different hemodynamics than a tricuspid aortic valve (TAV). While valvular calcification has been linked to genetic and atherogenic predispositions, hemodynamic abnormalities are increasingly pointed as potential pathogenic contributors. In particular, the wall shear stress (WSS) produced by blood flow on the leaflets regulates homeostasis in the TAV. In contrast, WSS alterations cause valve dysfunction and disease. While such observations support the existence of synergies between valvular hemodynamics and biology, the role played by BAV WSS in valvular calcification remains unknown. The objective of this study was to isolate the acute effects of native BAV WSS abnormalities on CAVD pathogenesis. Porcine aortic valve leaflets were subjected ex vivo to the native WSS experienced by TAV and type-I BAV leaflets for 48 hours. Immunostaining, immunoblotting and zymography were performed to characterize endothelial activation, pro-inflammatory paracrine signaling, extracellular matrix remodeling and markers involved in valvular interstitial cell activation and osteogenesis. While TAV and non-coronary BAV leaflet WSS essentially maintained valvular homeostasis, fused BAV leaflet WSS promoted fibrosa endothelial activation, paracrine signaling (2.4-fold and 3.7-fold increase in BMP-4 and TGF-β1, respectively, relative to fresh controls), catabolic enzyme secretion (6.3-fold, 16.8-fold, 11.7-fold, 16.7-fold and 5.5-fold increase in MMP-2, MMP-9, cathepsin L, cathepsin S and TIMP-2, respectively) and activity (1.7-fold and 2.4-fold increase in MMP-2 and MMP-9 activity, respectively), and bone matrix synthesis (5-fold increase in osteocalcin). In contrast, BAV WSS did not significantly affect α-SMA and Runx2 expressions and TIMP/MMP ratio. This study demonstrates the key role played by BAV hemodynamic abnormalities in CAVD pathogenesis and suggests the dependence of BAV vulnerability to calcification on the local degree of WSS abnormality.
Chemical investigation of the ethanol extract of soft coral Sinularia sp. collected from the South China Sea led to the isolation of three new polyoxygenated sterols, (3S,23R,24S)-ergost-5-ene-3β,23α,25-triol (1), (24S)-ergostane-6-acetate-3β,5α,6β,25-tetraol (2), (24S)-ergostane-6-acetate-3β,6β,12β,25-tetraol (3) together with three known ones (4–6). The structures, including relative configurations of the new compounds (1–3), were elucidated by detailed analysis of spectroscopic data (IR, UV, NMR, MS) and by comparison with related reported compounds. The absolute configuration of 1 was further determined by modified Mosher’s method. Compound 5 exhibited moderate cytotoxicity against K562 cell line with an IC50 value of 3.18 μM, but also displayed strong lethality toward the brine shrimp Artemia salina with a LC50 value of 0.96 μM.
soft coral; Sinularia sp.; polyoxygenated sterols; cytotoxicity
The ability of the pathogenic fungus Candida albicans to switch cellular morphologies is important for infection and virulence. Recent studies have revealed that C. albicans yeast cells can switch to filamentous growth under genotoxic stress in a manner dependent on the DNA replication/damage checkpoint. Here, we have investigated the functions of Pph3 (orf19.4378) and Psy2 (orf19.3685), whose orthologues in Saccharomyces cerevisiae mediate the dephosphorylation of the DNA damage checkpoint kinase Rad53 and the histone variant H2AX during recovery from DNA damage. Deleting PPH3 or PSY2 causes hypersensitivity to DNA-damaging agents, including cisplatin, methylmethane sulfonate (MMS), and UV light. In addition, pph3Δ and psy2Δ cells exhibit strong filamentous growth under genotoxic stress. Flow cytometry analysis shows that the mutant cells have lost the ability to adapt to genotoxic stress and remain arrested even after the stress is withdrawn. Furthermore, we show that Pph3 and Psy2 are required for the dephosphorylation of Rad53, but not H2AX, during DNA damage recovery. Taken together, these results show that C. albicans Pph3 and Psy2 have important roles in mediating genotoxin-induced filamentous growth and regulating Rad53 dephosphorylation.
SLE is an autoimmune inflammatory disease in which various pro- and anti-inflammatory cytokines, including TGF-β, IL-10, BAFF, IL-6, IFN-α, IFN-γ, IL-17, and IL-23, play crucial pathogenic roles. Virtually, all these cytokines can be generated by both innate and adaptive immune cells and exert different effects depending on specific local microenvironment. They can also interact with each other, forming a complex network to maintain delicate immune homeostasis. In this paper, we elaborate on the abnormal secretion and functions of these cytokines in SLE, analyze their potential pathogenic roles, and probe into the possibility of them being utilized as targets for therapy.
Years of education are inversely related to the prevalence of major depressive disorder (MDD), but the relationship between the clinical features of MDD and educational status is poorly understood. We investigated this in 1970 Chinese women with recurrent MDD identified in a clinical setting.
Clinical and demographic features were obtained from 1970 Han Chinese women with DSM-IV major depression between 30 and 60 years of age across China. Analysis of linear, logistic and multiple logistic regression models were used to determine the association between educational level and clinical features of MDD.
Subjects with more years of education are more likely to have MDD, with an odds ratio of 1.14 for those with more than ten years. Low educational status is not associated with an increase in the number of episodes, nor with increased rates of co-morbidity with anxiety disorders. Education impacts differentially on the symptoms of depression: lower educational attainment is associated with more biological symptoms and increased suicidal ideation and plans to commit suicide.
Findings may not generalize to males or to other patient populations. Since the threshold for treatment seeking differs as a function of education there may an ascertainment bias in the sample.
The relationship between symptoms of MDD and educational status in Chinese women is unexpectedly complex. Our findings are inconsistent with the simple hypothesis from European and US reports that low levels of educational attainment increase the risk and severity of MDD.
Major depressive disorder; Education; Socio-economic status; Symptom
The current recommended therapy for Kawasaki disease (KD) is the combination of intravenous immunoglobulin (IVIG) and aspirin. However, the role of corticosteroid therapy in KD remains controversial. Using meta-analysis, this study aimed to investigate the efficacy of corticosteroid therapy in KD by comparing it with standard IVIG and aspirin therapy. We included all related randomized and quasi-randomized controlled trials by searching Medline, the Cochrane Central Register of Controlled Trials, EMBASE, Pub Med, Chinese BioMedical Literature Database, China National Knowledge Infrastructure, and the Japanese database (Japan Science and Technology) as well as hand searches of selected references. Data collection and meta-analysis were performed to evaluate the effect of corticosteroids. Our search yielded 11 studies; 7 of which evaluated the effect of corticosteroid for primary therapy in KD, and 4 investigated the effect of corticosteroid therapy in IVIG-resistant patients. Meta-analysis of these studies revealed a significant reduction in the rates of initial treatment failure among patients who received corticosteroid therapy in combination with IVIG compared to IVIG alone (odds ratio (OR) = 0.50; 95% CI, 0.32~0.79; p = 0.003). Furthermore, the use of corticosteroids reduced the duration of fever and the time required for C-reactive protein to return to normal. Our data did not show any significant increase in the incidence of coronary artery lesions or coronary aneurysms (OR = 0.67; 95% CI, 0.35~1.28; p = 0.23) in the corticosteroid group. Conclusion. Corticosteroid combined with IVIG in primary treatment or as treatment of IVIG-resistant patients improved clinical course without increasing coronary artery lesions in children with acute KD.
Kawasaki disease; Coronary artery lesions; Coronary artery aneurysm; Corticosteroids; Meta-analysis
AIM: To describe survival trends in patients in Northeast China diagnosed as gastric cancer.
METHODS: A review of all inpatient and outpatient records of gastric cancer patients was conducted in the First Affiliated Hospital of China Medical University. All the gastric cancer patients who satisfied the inclusion criteria from January 1, 1980 through December 31, 2003 were included in the study. The main outcomes were based on median survival and 3-year and 5-year survival rates, by decade of diagnosis.
RESULTS: From 1980 through 2003, the median survival for patients with gastric cancer (n = 1604) increased from 33 mo to 49 mo. The decade of diagnosis was not significantly associated with patient survival for gastric cancer (P = 0.084 for overall survival, and P = 0.150 for 5-year survival); however, the survival rate of the 2000s was remarkably higher than that of the 1980s (P = 0.019 for overall survival, and P = 0.027 for 5-year survival).
CONCLUSION: There was no significant difference of survival among each period; however, the survival rate of the 2000s was remarkably higher than that of the 1980s.
Survival trends; Gastric cancer; Northeast China
Two different ligand occupancy structures of cytochrome P450 2B4 (CYP2B4) in complex with 1-biphenyl-4-methyl-1H–imidazole (1-PBI) have been solved by x-ray crystallography. 1-PBI belongs to a series of tight binding, imidazole-based CYP2B4 inhibitors. 1-PBI binding to CYP2B4 yields a type II spectrum with a Ks value of 0.23 µM and inhibits enzyme activity with an IC50 value of 0.035 µM. Previous CYP2B4 structures have shown a large degree of structural movement in response to ligand size. With two phenyl rings, 1-PBI is larger than 1-(4-chlorophenyl)imidazole (1-CPI) and 4-(4-chlorophenyl)imidazole (4-CPI) but smaller than bifonazole, which is branched and contains three phenyl rings. The CYP2B4:1-PBI complex is a structural intermediate to the closed CPI and the open bifonazole structures. The B/C loop reorganizes itself to include two short partial helices while closing one side of the active site. The F-G helix cassette pivots over the I-helix in direct response to the size of the ligand in the active site. A cluster of Phe residues at the fulcrum of this pivot point allows for dramatic repositioning of the cassette with only a relatively small amount of secondary structure rearrangement. Comparisons of ligand bound CYP2B4 structures reveal trends in plastic region mobility that could allow for predictions of their position in future structures based on ligand shape and size.
cytochrome P450 2B4; 1-biphenyl-4-methyl-1H–imidazole; 1-PBI; x-ray crystallography; plastic regions; F-G helix cassette
Insertional mutagenesis is an effective method for functional genomic studies in various organisms. It can rapidly generate easily tractable mutations. A large-scale insertional mutagenesis with the piggyBac (PB) transposon is currently performed in mice at the Institute of Developmental Biology and Molecular Medicine (IDM), Fudan University in Shanghai, China. This project is carried out via collaborations among multiple groups overseeing interconnected experimental steps and generates a large volume of experimental data continuously. Therefore, the project calls for an efficient database system for recording, management, statistical analysis, and information exchange.
This paper presents a database application called MP-PBmice (insertional mutation mapping system of PB Mutagenesis Information Center), which is developed to serve the on-going large-scale PB insertional mutagenesis project. A lightweight enterprise-level development framework Struts-Spring-Hibernate is used here to ensure constructive and flexible support to the application. The MP-PBmice database system has three major features: strict access-control, efficient workflow control, and good expandability. It supports the collaboration among different groups that enter data and exchange information on daily basis, and is capable of providing real time progress reports for the whole project. MP-PBmice can be easily adapted for other large-scale insertional mutation mapping projects and the source code of this software is freely available at http://www.idmshanghai.cn/PBmice.
MP-PBmice is a web-based application for large-scale insertional mutation mapping onto the mouse genome, implemented with the widely used framework Struts-Spring-Hibernate. This system is already in use by the on-going genome-wide PB insertional mutation mapping project at IDM, Fudan University.
Chicken type II collagen (CCII) is a protein extracted from the cartilage of chicken breast and exhibits intriguing possibilities for the treatment of autoimmune diseases by inducing oral tolerance. A 24-week, double-blind, double-dummy, randomized, methotrexate (MTX)-controlled study was conducted to evaluate the efficacy and safety of CCII in the treatment of rheumatoid arthritis (RA).
Five hundred three RA patients were included in the study. Patients received either 0.1 mg daily of CCII (n = 326) or 10 mg once a week of MTX (n = 177) for 24 weeks. Each patient was evaluated for pain, morning stiffness, tender joint count, swollen joint count, health assessment questionnaire (HAQ), assessments by investigator and patient, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP) by using the standard tools at baseline (week 0) and at weeks 12 and 24. Additionally, rheumatoid factor (RF) was evaluated at weeks 0 and 24. Measurement of a battery of biochemical parameters in serum, hematological parameters, and urine analysis was performed to evaluate the safety of CCII.
Four hundred fifty-four patients (94.43%) completed the 24-week follow-up. In both groups, there were decreases in pain, morning stiffness, tender joint count, swollen joint count, HAQ, and assessments by investigator and patient, and all differences were statistically significant. In the MTX group, ESR and CRP decreased. RF did not change in either group. At 24 weeks, 41.55% of patients in the CCII group and 57.86% in the MTX group met the American College of Rheumatology 20% improvement criteria (ACR-20) and 16.89% and 30.82%, respectively, met the ACR 50% improvement criteria (ACR-50). Both response rates for ACR-20 and ACR-50 in the CCII group were lower than those of the MTX group, and this difference was statistically significant (P < 0.05). The DAS28 (disease activity score using 28 joint counts) values of the two treatment groups were calculated, and there was a statistically significant difference between the two treatment groups (P < 0.05). Gastrointestinal complaints were common in both groups, but there were fewer and milder side effects in the CCII group than in the MTX group. The incidence of adverse events between the two groups was statistically significant (P < 0.05).
CCII is effective in the treatment of RA and is safe for human consumption. CCII exerts its beneficial effects by controlling inflammatory responses through inducing oral tolerance in RA patients.
Clinical trial registration number: ChiCTR-TRC-00000093.
The crystal structure of P450 2B4 bound with 1-(4-chlorophenyl)imidazole (1-CPI) has been determined to delineate the structural basis for the observed differences in binding affinity and thermodynamics relative to 4-(4-chlorophenyl)imidazole (4-CPI). Compared with the previously reported 4-CPI complex, there is a shift in the 1-CPI complex of the protein backbone in helices F and I, repositioning the side chains of Phe-206, Phe-297, and Glu-301, and leading to significant reshaping of the active site. Phe-206 and Phe-297 exchange positions, with Phe-206 becoming a ligand-contact residue, while Glu-301, rather than hydrogen bonding to the ligand, flips away from the active site and interacts with His-172. As a result the active site volume expands from 200 Å3 in the 4-CPI complex to 280 Å3 in the 1-CPI complex. Based on the two structures, it was predicted that a Phe-206->Ala substitution would alter 1-CPI but not 4-CPI binding. Isothermal titration calorimetry experiments indicated that this substitution had no effect on the thermodynamic signature of 4-CPI binding to 2B4. In contrast, relative to wild-type 1-CPI binding to F206A showed significantly less favorable entropy but more favorable enthalpy. This result is consistent with loss of the aromatic side chain and possible ordering of water molecules, now able to interact with Glu-301 and exposed residues in the I-helix. Hence, thermodynamic measurements support the active site rearrangement observed in the crystal structure of the 1-CPI complex and illustrate the malleability of the active site with the fine tuning of residue orientations and thermodynamic signatures.
Based on recent directed evolution of P450 2B1, six P450 2B11 mutants at three positions were created in an N-terminal modified construct termed P450 2B11dH and characterized for enzyme catalysis using five substrates. Mutant I209A demonstrated a 3.2-fold enhanced kcat/Km for 7-ethoxy-4-trifluoromethylcourmarin O-deethylation, largely due to a dramatic decrease in Km (0.72 vs. 18 μM). I209A also demonstrated enhanced selectivity for testosterone 16β-hydroxylation over 16α-hydroxylation. In contrast, V183L showed a 4-fold increased kcat for 7-benzyloxyresorufin debenzylation and a 4.7-fold increased kcat/Km for testosterone 16α-hydroxylation. V183L also displayed a 1.7-fold higher kcat/Km than P450 2B11dH with the anti-cancer prodrugs cyclophosphamide and ifosfamide, resulting from a ~4-fold decrease in Km. Introduction of the V183L mutation into full-length P450 2B11 did not enhance the kcat/Km. Overall, the re-engineered P450 2B11dH enzymes exhibited enhanced catalytic efficiency with several substrates including the anti-cancer prodrugs.
cytochrome P450; P450 2B11; structure-function relationships; site-directed mutagenesis; enzyme catalysis
DNA transposon piggyBac (PB) is a newly established mutagen for large-scale mutagenesis in mice. We have designed and implemented an integrated database system called PBmice (PB Mutagenesis Information CEnter) for storing, retrieving and displaying the information derived from PB insertions (INSERTs) in the mouse genome. This system is centered on INSERTs with information including their genomic locations and flanking genomic sequences, the expression levels of the hit genes, and the expression patterns of the trapped genes if a trapping vector was used. It also archives mouse phenotyping data linked to INSERTs, and allows users to conduct quick and advanced searches for genotypic and phenotypic information relevant to a particular or a set of INSERT(s). Sequence-based information can be cross-referenced with other genomic databases such as Ensembl, BLAST and GBrowse tools used in PBmice offer enhanced search and display for additional information relevant to INSERTs. The total number and genomic distribution of PB INSERTs, as well as the availability of each PB insertional LINE can also be viewed with user-friendly interfaces. PBmice is freely available at http://www.idmshanghai.cn/PBmice or http://www.scbit.org/PBmice/.
Hemodialysis (HD) and peritoneal dialysis (PD) are important renal replacement treatment in end stage renal disease (ESRD), but the comparison of quality of life (QOL) and causes of hospitalisation between the two modalities in China is lacking. In the present study, we compared the two modalities in a multi-center study.
Subjects and methods
Six hundred and fifty four HD and 408 PD patients were investigated from 10 hospitals in China from Sept, 2004 to Jan, 2005. Among the HD patients, there were 360 males and 294 females with a mean age of 57.22 ± 12.49 years (18–88 y). Among PD patients, there were 165 males and 243 females, with a mean age of 61.59 ± 12.65 years (22–89 y). Health related 36 items short form questionnaires (SF-36) were used to assess the quality of life. Hospitalisation data were collected and analyzed.
SF-36 domains of Body Pain (BP), General Health (GH), Role-Emotional (RE), Social Functioning (SF), Vitality (VT) and Mental Health (MH) were all significantly higher in the PD patients as compared to the HD patients although there was no significant difference in Physical Functioning (PF) and Role-Physical (RP) between the two groups. The two most common causes of hospitalisation in HD patients were cardiovascular disease (39.8%) and pulmonary infection (21.3%), while they were infectious peritonitis (47.6%) and cardiovascular disease (31.9%) in PD patients. The ever hospitalised patients had lower SF-36 scores in the domains of PF, BP, GH, RE, SF, VT and MH as compared to those of non-hospitalised patients.
Our study indicated that with the current practice in China, PD patients may enjoy better quality of life than their HD counterparts. Our results also showed that the most common cause of hospitalisation was cardiovascular disease in HD patients and peritonitis in PD patients.
Polycomb group (PcG) proteins are able to maintain the memory of silent transcriptional states of homeotic genes throughout development. In
Drosophila, they form multimeric complexes that bind to specific DNA regulatory elements named PcG response elements (PREs). To date, few PREs have been identified and the chromosomal distribution of PcG proteins during development is unknown. We used chromatin immunoprecipitation (ChIP) with genomic tiling path microarrays to analyze the binding profile of the PcG proteins Polycomb (PC) and Polyhomeotic (PH) across 10 Mb of euchromatin. We also analyzed the distribution of
GAGA factor (GAF), a sequence-specific DNA binding protein that is found at most previously identified PREs. Our data show that PC and PH often bind to clustered regions within large loci that encode transcription factors which play multiple roles in developmental patterning and in the regulation of cell proliferation. GAF co-localizes with PC and PH to a limited extent, suggesting that GAF is not a necessary component of chromatin at PREs. Finally, the chromosome-association profile of PC and PH changes during development, suggesting that the function of these proteins in the regulation of some of their target genes might be more dynamic than previously anticipated.
The authors map the GAGA factor and Polycomb group proteins PC and PH to
Drosophila chromosomes. PC and PH exhibit dynamic localizations over time at loci encoding transcription factors frequently important for development.