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1.  Activation of lysosomal function in the course of autophagy via mTORC1 suppression and autophagosome-lysosome fusion 
Cell Research  2013;23(4):508-523.
Lysosome is a key subcellular organelle in the execution of the autophagic process and at present little is known whether lysosomal function is controlled in the process of autophagy. In this study, we first found that suppression of mammalian target of rapamycin (mTOR) activity by starvation or two mTOR catalytic inhibitors (PP242 and Torin1), but not by an allosteric inhibitor (rapamycin), leads to activation of lysosomal function. Second, we provided evidence that activation of lysosomal function is associated with the suppression of mTOR complex 1 (mTORC1), but not mTORC2, and the mTORC1 localization to lysosomes is not directly correlated to its regulatory role in lysosomal function. Third, we examined the involvement of transcription factor EB (TFEB) and demonstrated that TFEB activation following mTORC1 suppression is necessary but not sufficient for lysosomal activation. Finally, Atg5 or Atg7 deletion or blockage of the autophagosome-lysosome fusion process effectively diminished lysosomal activation, suggesting that lysosomal activation occurring in the course of autophagy is dependent on autophagosome-lysosome fusion. Taken together, this study demonstrates that in the course of autophagy, lysosomal function is upregulated via a dual mechanism involving mTORC1 suppression and autophagosome-lysosome fusion.
PMCID: PMC3616426  PMID: 23337583
autophagy; lysosome; mTORC1; autophagosome
2.  Mst1 and Mst2 Are Essential Regulators of Trophoblast Differentiation and Placenta Morphogenesis 
PLoS ONE  2014;9(3):e90701.
The placenta is essential for survival and growth of the fetus because it promotes the delivery of nutrients and oxygen from the maternal circulation as well as fetal waste disposal. Mst1 and Mst2 (Mst1/2), key components of the mammalian hpo/Mst signaling pathway, encode two highly conserved Ser/Thr kinases and play important roles in the prevention of tumorigenesis and autoimmunity, control of T cell development and trafficking, and embryonic development. However, their functions in placental development are not fully understood, and the underlying cellular and molecular mechanisms remain elusive. Here, we investigated the functions of Mst1/2 in mouse placental development using both conventional and conditional (endothelial) Mst1/2 double knockout mice. We found that the number of trophoblast giant cells dramatically increased while spongiotrophoblast cells almost completely disappeared in Mst1/2 deficient placentas. We showed that Mst1/2 deficiency down regulated the expression of Mash2, which is required for suppressing the differentiation of trophoblast giant cells. Furthermore, we demonstrated that endothelial-specific deletion of Mst1/2 led to impaired placental labyrinthine vasculature and embryonic lethality at E11.5, but neither affected vasculature in yolk sac and embryo proper nor endocardium development. Collectively, our findings suggest that Mst1/2 regulate placental development by control of trophoblast cell differentiation and labyrinthine vasculature at midgestation and Mst1/2 control labyrinth morphogenesis in trophoblast- and fetal endothelial-dependent manners. Thus, our studies have defined novel roles of Mst1/2 in mouse placental development.
PMCID: PMC3942462  PMID: 24595170
3.  A Master-Slave Surveillance System to Acquire Panoramic and Multiscale Videos 
The Scientific World Journal  2014;2014:491549.
This paper describes a master-slave visual surveillance system that uses stationary-dynamic camera assemblies to achieve wide field of view and selective focus of interest. In this system, the fish-eye panoramic camera is capable of monitoring a large area, and the PTZ dome camera has high mobility and zoom ability. In order to achieve the precise interaction, preprocessing spatial calibration between these two cameras is required. This paper introduces a novel calibration approach to automatically calculate a transformation matrix model between two coordinate systems by matching feature points. In addition, a distortion correction method based on Midpoint Circle Algorithm is proposed to handle obvious horizontal distortion in the captured panoramic image. Experimental results using realistic scenes have demonstrated the efficiency and applicability of the system with real-time surveillance.
PMCID: PMC3960552
4.  Molecular analysis of hyperthermophilic endoglucanase Cel12B from Thermotoga maritima and the properties of its functional residues 
Although many hyperthermophilic endoglucanases have been reported from archaea and bacteria, a complete survey and classification of all sequences in these species from disparate evolutionary groups, and the relationship between their molecular structures and functions are lacking. The completion of several high-quality gene or genome sequencing projects provided us with the unique opportunity to make a complete assessment and thorough comparative analysis of the hyperthermophilic endoglucanases encoded in archaea and bacteria.
Structure alignment of the 19 hyperthermophilic endoglucanases from archaea and bacteria which grow above 80°C revealed that Gly30, Pro63, Pro83, Trp115, Glu131, Met133, Trp135, Trp175, Gly227 and Glu229 are conserved amino acid residues. In addition, the average percentage composition of residues cysteine and histidine of 19 endoglucanases is only 0.28 and 0.74 while it is high in thermophilic or mesophilic one. It can be inferred from the nodes that there is a close relationship among the 19 protein from hyperthermophilic bacteria and archaea based on phylogenetic analysis. Among these conserved amino acid residues, as far as Cel12B concerned, two Glu residues might be the catalytic nucleophile and proton donor, Gly30, Pro63, Pro83 and Gly227 residues might be necessary to the thermostability of protein, and Trp115, Met133, Trp135, Trp175 residues is related to the binding of substrate. Site-directed mutagenesis results reveal that Pro63 and Pro83 contribute to the thermostability of Cel12B and Met133 is confirmed to have role in enhancing the binding of substrate.
The conserved acids have been shown great importance to maintain the structure, thermostability, as well as the similarity of the enzymatic properties of those proteins. We have made clear the function of these conserved amino acid residues in Cel12B protein, which is helpful in analyzing other undetailed molecular structure and transforming them with site directed mutagenesis, as well as providing the theoretical basis for degrading cellulose from woody and herbaceous plants.
PMCID: PMC3936955  PMID: 24529187
Cellulose; Conserved amino acid residues; Endoglucanase; Phylogenetic analysis; Thermostability
5.  Spectral CT Imaging of Laryngeal and Hypopharyngeal Squamous Cell Carcinoma: Evaluation of Image Quality and Status of Lymph Nodes 
PLoS ONE  2013;8(12):e83492.
The purpose of this study was to evaluate image quality and status of lymph nodes in laryngeal and hypopharyngeal squamous cell carcinoma (SCC) patients using spectral CT imaging.
Materials and Methods
Thirty-eight patients with laryngeal and hypopharyngeal SCCs were scanned with spectral CT mode in venous phase. The conventional 140-kVp polychromatic images and one hundred and one sets of monochromatic images were generated ranging from 40 keV to 140 keV. The mean optimal keV was calculated on the monochromatic images. The image quality of the mean optimal keV monochromatic images and polychromatic images was compared with two different methods including a quantitative analysis method and a qualitative analysis method. The HU curve slope (λHU) in the target lymph nodes and the primary lesion was calculated respectively. The ratio of λHU was studied between metastatic and non-metastatic lymph nodes group.
A total of 38 primary lesions were included. The mean optimal keV was obtained at 55±1.77 keV on the monochromatic images. The image quality evaluated by two different methods including a quantitative analysis method and a qualitative analysis method was obviously increased on monochromatic images than polychromatic images (p<0.05). The ratio of λHU between metastatic and non-metastatic lymph nodes was significantly different in the venous phase images (p<0.05).
The monochromatic images obtained with spectral CT can be used to improve the image quality of laryngeal and hypopharyngeal SCC and the N-staging accuracy. The quantitative ratio of λHU may be helpful for differentiating between metastatic and non-metastatic cervical lymph nodes.
PMCID: PMC3875441  PMID: 24386214
6.  A novel model mouth system for evaluation of In Vitro release of nicotine from moist snuff 
Pouch moist snuff, as a form of oral smokeless tobacco products, is becoming increasingly popular in North America, Scandinavia (where it is known as Snus), South Asia and parts of Africa. User usually places a pouch between the upper jaw and cheek to obtain euphoria from tobacco, leading to partial intake of tobacco constituents. To evaluate user exposure to tobacco, an approach with a novel model mouth system was developed and applied to evaluate release of nicotine from the pouch.
A novel model mouth system has been developed to evaluate release behavior of tobacco constituents in pouch moist snuff. The system consists of the release medium reservoir module, the flow speed control module, the temperature control module, nicotine release module, and release solution collection module, and simulates buccal condition in terms of temperature, saliva compositions, and the rate of saliva production, etc. Artificial saliva was used as the release medium to evaluate release of nicotine in pouch moist snuff. The optimized test condition was that the release temperature of 37°C and the flow rate performed at 0.2 mL min-1 in the first 5 min and 0.1 mL min-1 in the next 55 min. The performance of the model mouth system was compared with in vivo data of nicotine release in human volunteers. Data from 23 brands of moist snuff indicated that nicotine release rates increased with extraction time and approximately 60-90% of nicotine was released after 30 min of extraction in most of the samples, and the release behavior of nicotine was affected by product weights, nicotine concentration, and product pH, etc.
The model mouth system can be used to evaluate the release behavior of constituents in pouch moist snuff, especially those directly related to human health such as nicotine and tobacco specific nitrosamine (TSNA), etc. This indicated that the system is an alternative tool to evaluate user exposure to tobacco. With further testing and validation, the model mouth system can be applied in risk evaluation of smokeless tobacco products.
PMCID: PMC3829806  PMID: 24215662
Model mouth system; Nicotine; In vitro; Release; Moist snuff; Smokeless tobacco products
7.  Association between RASSF1A Promoter Methylation and Ovarian Cancer: A Meta-Analysis 
PLoS ONE  2013;8(10):e76787.
The RAS association domain family protein 1a gene (RASSF1A) is one of the tumor suppressor genes (TSG). Inactivation of RASSF1A is critical to the pathogenesis of cancer. Aberrant TSG methylation was considered an important epigenetic silencing mechanism in the progression of ovarian cancer. A number of studies have discussed association between RASSF1A promoter methylation and ovarian cancer. However, they were mostly based on a small number of samples and showed inconsist results, Therefore, we conducted a meta-analysis to better identify the association.
Eligible studies were identified by searching the PubMed, EMBASE, Web of Science, and CNKI databases using a systematic searching strategy. We pooled the odds ratio (ORs) from individual studies using a fixed-effects model. We performed heterogeneity and publication bias analysis simultaneously.
Thirteen studies, with 763 ovarian cancer patients and 438 controls were included in the meta-analysis. The frequencies of RASSF1A promoter methylation ranged from 30% to 58% (median is 48%) in the cancer group and 0 to 21% (median is 0) in the control group. The frequencies of RASSF1A promoter methylation in the cancer group were significantly higher than those in the control group. The pooled odds ratio was 11.17 (95% CI = 7.51–16.61) in the cancer group versus the corresponding control group under the fixed-effects model.
The results suggested that RASSF1A promoter methylation had a strong association with ovarian cancer.
PMCID: PMC3792894  PMID: 24116157
8.  High-level expression of a novel thermostable and mannose-tolerant β-mannosidase from Thermotoga thermarum DSM 5069 in Escherichia coli 
BMC Biotechnology  2013;13:83.
Mannan is one of the primary polysaccharides in hemicellulose and is widely distributed in plants. β-Mannosidase is an important constituent of the mannan-degrading enzyme system and it plays an important role in many industrial applications, such as food, feed and pulp/paper industries as well as the production of second generation bio-fuel. Therefore, the mannose-tolerant β-mannosidase with high catalytic efficiency for bioconversion of mannan has a great potential in the fields as above.
A β-mannosidase gene (Tth man5) of 1,827 bp was cloned from the extremely thermophilic bacterium Thermotoga thermarum DSM 5069 that encodes a protein containing 608 amino acid residues, and was over-expressed in Escherichia coli BL21 (DE3). The results of phylogenetic analysis, amino acid alignment and biochemical properties indicate that the Tth Man5 is a novel β-mannosidase of glycoside hydrolase family 5. The optimal activity of the Tth Man5 β-mannosidase was obtained at pH 5.5 and 85°C and was stable over a pH range of 5.0 to 8.5 and exhibited 2 h half-life at 90°C. The kinetic parameters Km and Vmax values for p-nitrophenyl-β-D-mannopyranoside and 1,4-β-D-mannan were 4.36±0.5 mM and 227.27±1.59 μmol min-1 mg-1, 58.34±1.75 mg mL-1 and 285.71±10.86 μmol min-1 mg-1, respectively. The kcat/Km values for p-nitrophenyl-β-D-mannopyranoside and 1,4-β-D-mannan were 441.35±0.04 mM-1 s-1 and 41.47±1.58 s-1 mg-1 mL, respectively. It displayed high tolerance to mannose, with a Ki value of approximately 900 mM.
This work provides a novel and useful β-mannosidase with high mannose tolerance, thermostability and catalytic efficiency, and these characteristics constitute a powerful tool for improving the enzymatic conversion of mannan through synergetic action with other mannan-degrading enzymes.
PMCID: PMC3852774  PMID: 24099409
Thermotoga thermarum; β-mannosidase; Mannose-tolerant; Mannan; Thermostability; Mannooligosaccharides
9.  Association between MGMT Promoter Methylation and Non-Small Cell Lung Cancer: A Meta-Analysis 
PLoS ONE  2013;8(9):e72633.
O6-methylguanine-DNA methyltransferase (MGMT) is one of most important DNA repair enzyme against common carcinogens such as alkylate and tobacco. Aberrant promoter methylation of the gene is frequently observed in non-small cell lung cancer (NSCLC). However, the importance of epigenetic inactivation of the gene in NSCLC published in the literature showed inconsistence. We quantified the association between MGMT promoter methylation and NSCLC using a meta-analysis method.
We systematically reviewed studies of MGMT promoter methylation and NSCLC in PubMed, EMBASE, Ovid, ISI Web of Science, Elsevier and CNKI databases and quantified the association between MGMT promoter methylation and NSCLC using meta-analysis method. Odds ratio (OR) and corresponding 95% confidence interval (CI) were calculated to evaluate the strength of association. Potential sources of heterogeneity were assessed by subgroup analysis and meta-regression.
A total of 18 studies from 2001 to 2011, with 1, 160 tumor tissues and 970 controls, were involved in the meta-analysis. The frequencies of MGMT promote methylation ranged from 1.5% to 70.0% (median, 26.1%) in NSCLC tissue and 0.0% to 55.0% (median, 2.4%) in non-cancerous control, respectively. The summary of OR was 4.43 (95% CI: 2.85, 6.89) in the random-effects model. With stratification by potential source of heterogeneity, the OR was 20.45 (95% CI: 5.83, 71.73) in heterogeneous control subgroup, while it was 4.16 (95% CI: 3.02, 5.72) in the autologous control subgroup. The OR was 5.31 (95% CI: 3.00, 9.41) in MSP subgroup and 3.06 (95% CI: 1.75, 5.33) in Q-MSP subgroup.
This meta-analysis identified a strong association between methylation of MGMT gene and NSCLC. Prospective studies should be required to confirm the results in the future.
PMCID: PMC3784462  PMID: 24086261
10.  Improved regenerative myogenesis and muscular dystrophy in mice lacking Mkp5  
The Journal of Clinical Investigation  2013;123(5):2064-2077.
Duchenne muscular dystrophy (DMD) is a degenerative skeletal muscle disease caused by mutations in dystrophin. The degree of functional deterioration in muscle stem cells determines the severity of DMD. The mitogen-activated protein kinases (MAPKs), which are inactivated by MAPK phosphatases (MKPs), represent a central signaling node in the regulation of muscle stem cell function. Here we show that the dual-specificity protein phosphatase DUSP10/MKP-5 negatively regulates muscle stem cell function in mice. MKP-5 controlled JNK to coordinate muscle stem cell proliferation and p38 MAPK to control differentiation. Genetic loss of Mkp5 in mice improved regenerative myogenesis and dystrophin-deficient mdx mice lacking Mkp5 exhibited an attenuated dystrophic muscle phenotype. Hence, enhanced promyogenic MAPK activity preserved muscle stem cell function even in the absence of dystrophin and ultimately curtailed the pathogenesis associated with DMD. These results identify MKP-5 as an essential negative regulator of the promyogenic actions of the MAPKs and suggest that MKP-5 may serve as a target to promote muscle stem cell function in the treatment of degenerative skeletal muscle diseases.
PMCID: PMC3635719  PMID: 23543058
11.  Biochemical properties of a novel thermostable and highly xylose-tolerant β-xylosidase/α-arabinosidase from Thermotoga thermarum 
β-Xylosidase is an important constituent of the hemicellulase system and it plays an important role in hydrolyzing xylooligosaccharides to xylose. Xylose, a useful monose, has been utilized in a wide range of applications such as food, light, chemical as well as energy industry. Therefore, the xylose-tolerant β-xylosidase with high specific activity for bioconversion of xylooligosaccharides has a great potential in the fields as above.
A β-xylosidase gene (Tth xynB3) of 2,322 bp was cloned from the extremely thermophilic bacterium Thermotoga thermarum DSM 5069 that encodes a protein containing 774 amino acid residues, and was expressed in Escherichia coli BL21 (DE3). The phylogenetic trees of β-xylosidases were constructed using Neighbor-Joining (NJ) and Maximum-Parsimony (MP) methods. The phylogeny and amino acid analysis indicated that the Tth xynB3 β-xylosidase was a novel β-xylosidase of GH3. The optimal activity of the Tth xynB3 β-xylosidase was obtained at pH 6.0 and 95°C and was stable over a pH range of 5.0-7.5 and exhibited 2 h half-life at 85°C. The kinetic parameters Km and Vmax values for p-nitrophenyl-β-D-xylopyranoside and p-nitrophenyl-α-L-arabinofuranoside were 0.27 mM and 223.3 U/mg, 0.21 mM and 75 U/mg, respectively. The kcat/Km values for p-nitrophenyl-β-D-xylopyranoside and p-nitrophenyl-α-L-arabinofuranoside were 1,173.4 mM-1 s-1 and 505.9 mM-1 s-1, respectively. It displayed high tolerance to xylose, with Ki value approximately 1000 mM. It was stimulated by xylose at higher concentration up to 500 mM, above which the enzyme activity of Tth xynB3 β-xylosidase was gradually decreased. However, it still remained approximately 50% of its original activity even if the concentration of xylose was as high as 1000 mM. It was also discovered that the Tth xynB3 β-xylosidase exhibited a high hydrolytic activity on xylooligosaccharides. When 5% substrate was incubated with 0.3 U Tth xynB3 β-xylosidase in 200 μL reaction system for 3 h, almost all the substrate was biodegraded into xylose.
The article provides a useful and novel β-xylosidase displaying extraordinary and desirable properties: high xylose tolerance and catalytic activity at temperatures above 75°C, thermally stable and excellent hydrolytic activity on xylooligosaccharides.
PMCID: PMC3621209  PMID: 23422003
Thermotoga thermarum; β-xylosidase; α-arabinosidase; Xylose tolerant; Hemicellulose; Thermostability; Xylooligosaccharides
12.  A novel highly thermostable xylanase stimulated by Ca2+ from Thermotoga thermarum: cloning, expression and characterization 
Xylanase is an important component of hemicellulase enzyme system. Since it plays an important role in the hydrolysis of hemicellulose into xylooligosaccharides (XOs), high thermostable xylanase has been the focus of much recent attention as powerful enzyme as well as in the field of biomass utilization.
A xylanase gene (xyn10A) with 3,474 bp was cloned from the extremely thermophilic bacterium Thermotoga thermarum that encodes a protein containing 1,158 amino acid residues. Based on amino acid sequence homology, hydrophobic cluster and three dimensional structure analyses, it was attested that the xylanase belongs to the glycoside hydrolase (GH) families 10 with five carbohydrate binding domains. When the xylanase gene was cloned and expressed in Escherichia coli BL21 (DE3), the specific enzyme activity of xylanase produced by the recombinant strain was up to 145.8 U mg-1. The xylanase was optimally active at 95°C, pH 7.0. In addition, it exhibited high thermostability over broad range of pH 4.0-8.5 and temperature 55-90°C upon the addition of 5 mM Ca2+. Confirmed by Ion Chromatography System (ICS) analysis, the end products of the hydrolysis of beechwood xylan were xylose, xylobiose, xylotriose, xylotetraose, xylopentaose and xylohexaose.
The xylanase from T. thermarum is one of the hyperthermophilic xylanases that exhibits high thermostability, and thus, is a suitable candidate for generating XOs from cellulosic materials such as agricultural and forestry residues for the uses as prebiotics and precursors for further preparation of furfural and other chemicals.
PMCID: PMC3598563  PMID: 23418789
Xylanase; Xylan; Thermostability; Beechwood; Oat spelt; Birchwood
13.  Thermoanaerobacterium thermosaccharolyticum β-glucosidase: a glucose-tolerant enzyme with high specific activity for cellobiose 
β-Glucosidase is an important component of the cellulase enzyme system. It does not only participate in cellulose degradation, it also plays an important role in hydrolyzing cellulose to fermentable glucose by relieving the inhibition of exoglucanase and endoglucanase from cellobiose. Therefore, the glucose-tolerant β-glucosidase with high specific activity for cellobiose might be a potent candidate for industrial applications.
The β-glucosidase gene bgl that encodes a 443-amino-acid protein was cloned and over-expressed from Thermoanaerobacterium thermosaccharolyticum DSM 571 in Escherichia coli. The phylogenetic trees of β-glucosidases were constructed using Neighbor-Joining (NJ) and Maximum-Parsimony (MP) methods. The phylogeny and amino acid analysis indicated that the BGL was a novel β-glucosidase. By replacing the rare codons for the N-terminal amino acids of the target protein, the expression level of bgl was increased from 6.6 to 11.2 U/mg in LB medium. Recombinant BGL was purified by heat treatment followed by Ni-NTA affinity. The optimal activity was at pH 6.4 and 70°C. The purified enzyme was stable over pH range of 5.2–7.6 and had a 1 h half life at 68°C. The activity of BGL was significantly enhanced by Fe2+ and Mn2+. The Vmax of 64 U/mg and 120 U/mg were found for p-nitrophenyl-β-D-glucopyranoside (Km value of 0.62 mM) and cellobiose (Km value of 7.9 mM), respectively. It displayed high tolerance to glucose and cellobiose. The Kcat for cellobiose was 67.7 s-1 at 60°C and pH 6.4, when the concentration of cellobiose was 290 mM. It was activated by glucose at concentrations lower that 200 mM. With glucose further increasing, the enzyme activity of BGL was gradually inhibited, but remained 50% of the original value in even as high as 600 mM glucose.
The article provides a useful novel β-glucosidase which displayed favorable properties: high glucose and cellobiose tolerance, independence of metal ions, and high hydrolysis activity on cellobiose.
PMCID: PMC3395577  PMID: 22571470
β-glucosidase; Glucose tolerance; Thermoanaerobacterium thermosaccharolyticum; Over-expression; Phylogeny
14.  Predicting Progression of IgA Nephropathy: New Clinical Progression Risk Score 
PLoS ONE  2012;7(6):e38904.
IgA nephropathy (IgAN) is a common cause of end-stage renal disease (ESRD) in Asia. In this study, based on a large cohort of Chinese patients with IgAN, we aim to identify independent predictive factors associated with disease progression to ESRD. We collected retrospective clinical data and renal outcomes on 619 biopsy-diagnosed IgAN patients with a mean follow-up time of 41.3 months. In total, 67 individuals reached the study endpoint defined by occurrence of ESRD necessitating renal replacement therapy. In the fully adjusted Cox proportional hazards model, there were four baseline variables with a significant independent effect on the risk of ESRD. These included: eGFR [HR = 0.96(0.95–0.97)], serum albumin [HR = 0.47(0.32–0.68)], hemoglobin [HR = 0.79(0.72–0.88)], and SBP [HR = 1.02(1.00–1.03)]. Based on these observations, we developed a 4-variable equation of a clinical risk score for disease progression. Our risk score explained nearly 22% of the total variance in the primary outcome. Survival ROC curves revealed that the risk score provided improved prediction of ESRD at 24th, 60th and 120th month of follow-up compared to the three previously proposed risk scores. In summary, our data indicate that IgAN patients with higher systolic blood pressure, lower eGFR, hemoglobin, and albumin levels at baseline are at a greatest risk of progression to ESRD. The new progression risk score calculated based on these four baseline variables offers a simple clinical tool for risk stratification.
PMCID: PMC3375310  PMID: 22719981
15.  9-(2,4-Dichloro­phen­yl)-3,3,6,6-tetra­methyl-3,4,5,6-tetra­hydro-9H-xanthene-1,8(2H,7H)-dione 
The title compound, C23H24Cl2O3, was synthesized by reaction of 2,4-dichloro­benzaldehyde and 5,5-dimethyl­cyclo­hexane-1,3-dione in ethyl­ene glycol. The central ring of the xanthene moiety is almost planar (with an r.m.s. deviation of 0.0268 Å from the least-squares plane) while the two outer rings, in a cis arrangement, display envelope conformations. The ring of the 2,4-dichloro­phenyl substituent is nearly perpendicular [85.89 (4)°] to the xanthene ring system.
PMCID: PMC3011743  PMID: 21589494
16.  Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling 
PLoS ONE  2010;5(5):e10845.
Multiple myeloma (MM) is a malignancy of terminally-differentiated plasma cells, and the second most prevalent blood cancer. At present there is no cure for MM, and the average prognosis is only three to five years. Current treatments such as chemotherapy are able to prolong a patient's life but rarely prevent relapse of the disease. Even hematopoietic stem cell transplants and novel drug combinations are often not curative, underscoring the need for a continued search for novel therapeutics. CD137 and its ligand are members of the Tumor Necrosis Factor (TNF) receptor and TNF superfamilies, respectively. Since CD137 ligand cross-linking enhances proliferation and survival of healthy B cells we hypothesized that it would also act as a growth stimulus for B cell cancers.
Methodology/Principal Findings
Proliferation and survival of B cell lymphoma cell lines were not affected or slightly enhanced by CD137 ligand agonists in vitro. But surprisingly, they had the opposite effects on MM cells, where CD137 ligand signals inhibited proliferation and induced cell death by apoptosis. Furthermore, secretion of the pro-inflammatory cytokines, IL-6 and IL-8 were also enhanced in MM but not in non-MM cell lines in response to CD137 ligand agonists. The secretion of these cytokines in response to CD137 ligand signaling was consistent with the observed activation of the classical NF-κB pathway. We hypothesize that the induction of this pathway results in activation-induced cell death, and that this is the underlying mechanism of CD137-induced MM cell death and growth arrest.
These data point to a hitherto unrecognized role of CD137 and CD137 ligand in MM cell biology. The selective inhibition of proliferation and induction of cell death in MM cells by CD137 ligand agonists may also warrant a closer evaluation of their therapeutic potential.
PMCID: PMC2877096  PMID: 20520765
17.  Ethyl (1R,4S,5R,9S,10R,13S)-5,9,13-trimethyl-14-methyl­ene-14-oxotetra­cyclo­[,10.04,9]hexa­decane-5-carboxyl­ate 
The title compound, C22H34O3, was synthesized from isosteviol. The asymmetric unit contains of two independent mol­ecules with the same absolute configurations. In both the mol­ecules, the three six-membered rings adopt chair conformations, the stereochemistry of the A/B and B/C ring junctions are trans, and the five-membered ring D adopts an envelope conformation.
PMCID: PMC2980194  PMID: 21580043
18.  4-(4-Chloro­phen­yl)-7,7-dimethyl-7,8-dihydro-4H-1-benzopyran-2,5(3H,6H)-dione 
The title compound, C17H17ClO3, has been synthesized by the reaction of p-chloro­benzaldehyde, isopropyl­idene malonate and 5,5-dimethyl­cyclo­hexane-1,3-dione with triethyl­benzyl­ammonium chloride in water as a green solvent. The six membered pyran­one ring of the hexa­hydro­coumarin system has a screw-boat conformation while the dimethyl­cyclo­hexenone system has a distorted envelope conformation. The dihedral angle between the least-squares planes of the coumarin ring system and the benzene ring is 85.64 (9)°.
PMCID: PMC2980257  PMID: 21580158
19.  4-(4-Methoxy­phen­yl)-7,7-dimethyl-5-oxo-5,6,7,8-tetra­hydrochromene-2,5-dione 
The title compound, C18H20O4, was synthesized by the reaction of 4-methoxy­benzaldehyde, 2,2-dimethyl-1,3-dioxane-4,6-dione and 5,5-dimethyl­cyclo­hexane-1,3-dione with triethyl­benzyl­ammonium chloride in water as a green solvent. In the mol­ecule of the title compound, the six-membered pyran­one ring of the hexa­hydro­coumarin system has a screw-boat conformation while that of the dimethyl­cyclo­hexenone system has a distorted envelope conformation. The CMe2 portion of this ring is disordered over two positions with refined occupancies of 0.721 (7) and 0.279 (7).
PMCID: PMC2972097  PMID: 21578924
20.  Community-based study on CKD subjects and the associated risk factors 
Nephrology Dialysis Transplantation  2009;24(7):2117-2123.
Background. The study was performed to investigate the prevalence, awareness and the risk factors of chronic kidney disease (CKD) in the community population in Shanghai, China.
Methods. A total of 2596 residents were randomly recruited from the community population in Shanghai, China. All were screened for albuminuria, haematuria, morning spot urine albumin-to-creatinine ratio and renal function. Serum creatinine, uric acid, cholesterol, triglyceride and haemoglobin were assessed. A simplified MDRD equation was used to estimate the glomerular filtration rate (eGFR). All studied subjects were screened by kidney ultrasound. Haematuria, if present in the morning spot urine dipstick test, was confirmed by microscopy. The associations among the demographic characteristics, health characteristics and indicators of kidney damage were examined.
Results. Two thousand five hundred and fifty-four residents (n = 2554), after giving informed consent and with complete data, were entered into this study. Albuminuria and haematuria were detected in 6.3% and 1.2% of all the studied subjects, respectively, whereas decreased kidney function was found in 5.8% of all studied subjects. Approximately 11.8% of subjects had at least one indicator of kidney damage. The rate of awareness of CKD was 8.2%. The logistic regression model showed that age, central obesity, hypertension, diabetes, anaemia, hyperuricaemia and nephrolithiasis each contributed to the development of CKD.
Conclusion. This is the first Shanghai community-based epidemiological study data on Chinese CKD patients. The prevalence of CKD in the community population in Shanghai is 11.8%, and the rate of awareness of CKD is 8.2%. All the factors including age, central obesity, hypertension, diabetes, anaemia, hyperuricaemia and nephrolithiasis are positively correlated with the development of CKD in our studied subjects.
PMCID: PMC2698090  PMID: 19193736
awareness; chronic kidney disease; epidemiology; prevalence; risk factors
21.  (1R,4S,8R,9R,12S,13S,14R,16S,17R,19R)-17-[(Ethyl­sulfan­yl)meth­yl]-9,14-di­hydroxy-7,7-dimethyl-2,18-dioxo-3,10-dioxapenta­cyclo[,13.04,12.08,12]nona­decan-19-yl acetate acetone solvate 
The title compound, C24H32O8S·C3H6O, features three six-membered and two five-membered rings. The six-membered rings adopt chair, boat and slightly distorted boat conformations whereas one five-membered ring adopts an approximate envelope conformation and the other a twist conformation. Disorder was modelled for the ethyl­thio group with the ethyl-C atoms resolved over three positions with occupancies of 0.58 (4), 0.23 (4) and 0.19 (3). In the crystal, an O—H⋯O hydrogen bond links the molecules into chains.
PMCID: PMC2968108  PMID: 21581657
22.  (1S*,2S*,5R*,8S*,11S*,14R*,17S*,20R*)-14-Methyl-6-methyl­ene-10,16,18-trioxa­hexa­cyclo­[,8.01,11.02,8.017,20]henicosane-7,9-dione: natural diter­penoid macrocalyxoformin B 
The title compound, C20H24O5, isolated from Rabdosia var. lophanthoides Hara, is built up from six fused rings. Cyclo­hexane ring A adopts a chair conformation, ring B exists in a screw-boat conformation and ring C adopts a boat conformation; the three five-membered rings D, E and F adopt envelope conformations.
PMCID: PMC2915036  PMID: 21200956
23.  N-Alkyl Urea Hydroxamic Acids as a New Class of Peptide Deformylase Inhibitors with Antibacterial Activity 
Peptide deformylase (PDF) is a prokaryotic metalloenzyme that is essential for bacterial growth and is a new target for the development of antibacterial agents. All previously reported PDF inhibitors with sufficient antibacterial activity share the structural feature of a 2-substituted alkanoyl at the P1′ site. Using a combination of iterative parallel synthesis and traditional medicinal chemistry, we have identified a new class of PDF inhibitors with N-alkyl urea at the P1′ site. Compounds with MICs of ≤4 μg/ml against gram-positive and gram-negative pathogens, including Staphylococcus aureus, Streptococcus pneumoniae, and Haemophilus influenzae, have been identified. The concentrations needed to inhibit 50% of enzyme activity (IC50s) for Escherichia coli Ni-PDF were ≤0.1 μM, demonstrating the specificity of the inhibitors. In addition, these compounds were very selective for PDF, with IC50s of consistently >200 μM for matrilysin and other mammalian metalloproteases. Structure-activity relationship analysis identified preferred substitutions resulting in improved potency and decreased cytotoxity. One of the compounds (VRC4307) was cocrystallized with PDF, and the enzyme-inhibitor structure was determined at a resolution of 1.7 Å. This structural information indicated that the urea compounds adopt a binding position similar to that previously determined for succinate hydroxamates. Two compounds, VRC4232 and VRC4307, displayed in vivo efficacy in a mouse protection assay, with 50% protective doses of 30.8 and 17.9 mg/kg of body weight, respectively. These N-alkyl urea hydroxamic acids provide a starting point for identifying new PDF inhibitors that can serve as antimicrobial agents.
PMCID: PMC127453  PMID: 12183225

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