Expression, purification, and characterization of c-FLIP tandem death effector domains from Escherichia coli.

Cellular FLICE-like inhibitory protein (c-FLIP) regulates extrinsic apoptosis by controlling procaspase-8 activation through its tandem N-terminal death effector domains (DEDs). Here, we present the expression and purification of c-FLIP tandem DEDs (tDED) from Escherichia coli. We observed that the c-FLIPtDED maintains monomeric form under near-physiological pH condition in vitro. Our results also reveal a significant correlation between the pH conditions and the structure of c-FLIPtDED (F114A). The described methods and results would be helpful for follow-up study on the structural and functional of c-FLIP.

Solution structure of c-FLIP death effector domains.

The formation of death-inducing signaling complex (DISC) and death effector domain (DED) filament initiates extrinsic apoptosis. Recruitment and activation of procaspase-8 at the DISC are regulated by c-FLIP. The interaction between c-FLIP and procaspase-8 is mediated by their tandem DEDs (tDED). However, the structure of c-FLIPtDED and how c-FLIP interferes with procaspase-8 activation at the DISC remain elusive. Here, we solved the monomeric structure of c-FLIPtDED (F114G) at near physiological pH by solution nuclear magnetic resonance (NMR). Structural superimposition reveals c-FLIPtDED (F114G) adopts a structural topology similar to that of procaspase-8tDED. Our results provide a structural basis for understanding how c-FLIP interacts with procaspase-8 and the molecular mechanisms of c-FLIP in regulating cell death.

Backbone and side-chain chemical shift assignments of a cellular FLICE-inhibitory protein (c-FLIPS).

Cellular FLICE-inhibitory protein (c-FLIP), which is involved in regulating the apoptosis of the extrinsic cell death pathway contains two death effector domains (DED). There are several splicing variants including short-form (c-FLIPS) and long-form (c-FLIPL). The death-inducing signaling complex (DISC) initiates apoptosis and programmed necrosis, DISC assembly and activation are regulated by c-FLIP. Here we report the NMR chemical shift assignments of c-FLIPs, which pave the way for investigating the molecular basis of the anti-apoptotic function of c-FLIPS.

The 90-day prognostic value of serum cyclophilin A in traumatic brain injury.

BACKGROUND: Cyclophilin A is involved in many inflammatory diseases and its expression is up-regulated after brain injury. We determined if serum cyclophilin A could be used as a marker for severity and 90-day outcome in patients with traumatic brain injury (TBI). METHODS: Serum cyclophilin A concentrations were quantified in 105 severe TBI patients and 105 healthy individuals. Its association with Glasgow Coma Scale (GCS) score, 90-day mortality and 90-day poor outcome (Glasgow Outcome Scale score of 1-3) were investigated. RESULTS: Serum cyclophilin A concentrations were significantly higher in TBI patients than in healthy individuals. Cyclophilin A concentrations had a close relation to GCS scores and showed a high discriminatory ability for 90-day mortality and poor outcome according to area under receiver operating characteristic curve (AUC). Its AUC was in the range of GCS scores. Moreover, its combination with GCS scores significantly improved the predictive performance of GCS scores alone. In addition, serum cyclophilin A emerged as an independent predictor for 90-day mortality, overall survival and poor outcome. CONCLUSIONS: Increased serum cyclophilin A concentrations could reflect trauma severity and unfavorable outcome after head trauma, substantializing cyclophilin A as a potential biomarker for prognostic prediction of TBI.

New meroterpenoids from the endophytic fungus Aspergillus flavipes AIL8 derived from the mangrove plant Acanthus ilicifolius.

Four new meroterpenoids (2-5), along with three known analogues (1, 6, and 7) were isolated from mangrove plant Acanthus ilicifolius derived endophytic fungus Aspergillus flavipes. The structures of these compounds were elucidated by NMR and MS analysis, the configurations were assigned by CD data, and the stereochemistry of 1 was confirmed by X-ray crystallography analysis. A possible biogenetic pathway of compounds 1-7 was also proposed. All compounds were evaluated for antibacterial and cytotoxic activities.

New phenyl derivatives from endophytic fungus Aspergillus flavipes AIL8 derived of mangrove plant Acanthus ilicifolius.

Two new aromatic butyrolactones, flavipesins A (1) and B (2), two new natural products (3 and 4), and a known phenyl dioxolanone (5) were isolated from marine-derived endophytic fungus Aspergillus flavipes. The structures of compounds 1-5 were elucidated by 1D- and 2D-NMR and MS analysis, the absolute configurations were assigned by optical rotation and CD data, and the stereochemistry of 1 was determined by X-ray crystallography analysis. 1 demonstrated lower MIC values against Staphylococcus aureus (8.0 µg/mL) and Bacillus subtillis (0.25 µg/mL). 1 also showed the unique antibiofilm activity of penetration through the biofilm matrix and kills live bacteria inside mature S. aureus biofilm.

Introduction of bifunctional groups into mesoporous silica for enhancing uptake of thorium(IV) from aqueous solution.

The potential industrial application of thorium (Th), as well as the environmental and human healthy problems caused by thorium, promotes the development of reliable methods for the separation and removal of Th(IV) from environmental and geological samples. Herein, the phosphonate-amino bifunctionalized mesoporous silica (PAMS) was fabricated by a one-step self-assembly approach for enhancing Th(IV) uptake from aqueous solution. The synthesized sorbent was found to possess ordered mesoporous structures with uniform pore diameter and large surface area, characterized by SEM, XRD, and N2 sorption/desorption measurements. The enhancement of Th(IV) uptake by PAMS was achieved by coupling of an access mechanism to a complexation mechanism, and the sorption can be optimized by adjusting the coverage of the functional groups in the PAMS sorbent. The systemic study on Th(IV) sorption/desorption by using one coverage of PAMS (PAMS12) shows that the Th(IV) sorption by PAMS is fast with equilibrium time of less than 1 h, and the sorption capacity is more than 160 mg/g at a relatively low pH. The sorption isotherm has been successfully modeled by the Langmuir isotherm and D-R isotherm, which reveals a monolayer homogeneous chemisorption of Th(IV) in PAMS. The Th(IV) sorption by PAMS is pH dependent but ionic strength independent. In addition, the sorbed Th(IV) can be completely desorbed using 0.2 mol/L or more concentrated nitric acid solution. The sorption test performed in the solution containing a range of competing metal ions suggests that the PAMS sorbent has a desirable selectivity for Th(IV) ions.

MOF-76: from a luminescent probe to highly efficient U(VI) sorption material.

MOF-76 exhibits not only high sensitivity for the detection of U(vi), but also high adsorption capacity of 298 mg g(-1) at a low pH value of ∼3.0. Furthermore, the high selectivity for uranium adsorption over a series of competing metal ions is also illustrated.

[Mechanism of heavy-metal tolerance in Pseudomonas aeruginosa ZGKD2].

The cadmium-resistant bacterium Pseudomonas aeruginosa strain ZGKD2 exhibiting tolerance to various heavy-metals was isolated from gangue pile of coal area in our laboratory. This bacterium could serve as an effective metal sequestering and growth-promoting bioinoculant for plants grown in metal-contaminated soil. However, the mechanism of heavy-metal tolerance is still unclear. When the beef extract-peptone medium was supplemented with 200-3 000 micromol x L(-1) Cd2+, Cu2+, Zn2+, Ni2+, Pb2+ or Mn2+, the maximum biomass of strain ZGKD2 decreased with the increase of heavy-metal concentrations, while different concentrations of heavy-metals had no significant effect on its alkaline production. Stationary-phase cells of strain ZGKD2 were exposed to 0, 200, 600 and 1 000 micromol x L(-1) of Cd2+ , Cu2+, Zn2+ and Ni2+ or 0, 1 000, 2 000 and 3 000 micromol x L(-1) of Pb2+ and Mn2+ for 2 h, respectively. The activity of SOD and CAT increased in a heavy-metal-concentration-dependent manner, especially in the Cd2+ and Cu2+ treatments. The siderophore production of strain ZGKD2 in modified sugar-aspartic acid medium was enhanced by 200- 1 000 micromol x L(-1) of various heavy-metals. Cd2+ and Zn2+ strongly induced the siderophore production of strain ZGKD2, Ni2+ and Mn2+ had little effect, whereas Cu2+ led to significant inhibition. The siderophore production of strain ZGKD2 was positively related with its metal tolerance. These results indicated that alkaline production, siderophore production, and the increase of antioxidant enzyme activities in strain ZGKD2 might be the main mechanisms of heavy-metal tolerance.

Pituitary adenoma biomarkers identified using proteomic fingerprint technology.

OBJECTIVE: To determine whether pituitary adenomas can be diagnosed by identifying protein biomarkers in the serum. METHODS: We compared serum proteins from 65 pituitary adenoma patients and 90 healthy donors using proteomic fingerprint technology combining magnetic beads with matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS). RESULTS: A total of 42 M/Z peaks were identified as related to pituitary adenoma (P<0.01). A diagnostic model established based on three biomarkers (3382.0, 4601.9, 9191.2) showed that the sensitivity of diagnosing pituitary adenoma was 90.0% and the specificity was 88.3%. The model was further tested by blind analysis showing that the sensitivity was 88.0% and the specificity was 83.3%. CONCLUSIONS: These results suggest that proteomic fingerprint technology can be used to identify pituitary adenoma biomarkers and the model based on three biomarkers (3382.0, 4601.9, 9191.2) provides a powerful and reliable method for diagnosing pituitary adenoma.

[Geographic variation of seed morphological traits of Picea schrenkiana var. tianschanica in Tianshan Mountains, Xinjiang of Northwest China].

Taking the Picea schrenkiana var. tianschanica forests at three sites with different longitudes (Zhaosu, Tianchi, and Qitai) in Tianshan Mountains as the objects, the cones were collected along an altitudinal gradient to analyze the variation of their seed morphological traits (seed scale length and width, seed scale length/width ratio, seed wing length and width, seed wing length/ width ratio, seed length and width, and seed length/width ratio). All the seed traits except seed width tended to decrease with increasing altitude. The seed traits except seed wing width, seed width, and seed length/width ratio all had significant negative correlations with altitude. Seed scale length and width and seed scale length/width ratio had significant positive correlations with longitude. Seed scale length, seed scale length/width ratio, and seed wing length/width ratio had significant negative correlations with slope degree. No significant correlations were observed between the seed traits except seed wing width and the slope aspect. Altitude was the main factor affecting the seed scale length, seed scale length/width ratio, and seed wing length/width ratio.

Decreased plasma gelsolin is associated with 1-year outcome in patients with traumatic brain injury.

PURPOSE: Decreased plasma gelsolin level has been associated with 1-month mortality after traumatic brain injury (TBI). Thus, we investigated the ability of gelsolin to predict 1-year mortality and functional outcome in these patients. METHODS: One hundred fourteen healthy controls and 114 patients with acute severe TBI were included in this study. Plasma gelsolin concentration on admission was measured by ELISA. RESULTS: Fifty-five patients (48.2%) had unfavorable outcome (Glasgow Outcome Scale score of 1-3) and 38 patients (33.3%) died in 1 year after TBI. Upon admission, plasma gelsolin level in patients was substantially lower than that in healthy controls. A multivariate analysis selected plasma gelsolin level as an independent predictor for 1-year unfavorable outcome and mortality of patients. A receiver operating characteristic curve analysis showed plasma gelsolin level predicted 1-year unfavorable outcome and mortality statistically significantly. The predictive value of the gelsolin concentration was thus similar to that of Glasgow Coma Scale (GCS) score. In a combined logistic-regression model, gelsolin did not statistically significantly improve the area under curve of GCS score. CONCLUSIONS: Plasma gelsolin level is a useful, complementary tool to predict functional outcome and mortality 1 year after TBI.

[Distribution patterns of Picea schrenkiana var. tianschanica population in Tianshan Mountains].

A vertical transect investigation on Picea schrenkiana var. tianschanica forests was conducted at five different longitudinal sites (Zhaosu, Gongliu, Wusu, Urumqi, and Hami) in Tianshan Mountains, and the distribution pattern of P. schrenkiana var. tianschanica population at each site was analyzed based on theoretical distribution model and aggregation intensity index. On the whole, the P. schrenkiana var. tianschanica population in Tianshan Mountains presented a clumped distribution, and the distribution pattern and clustering intensity were affected by the developmental stages of stem and the ranges of altitude to some degree. The clustering intensity increased with the increasing size (DBH) or developmental stage of stem, and had the highest values at high altitudes.

Effect of human cytomegalovirus infection on nerve growth factor expression in human glioma U251 cells.

OBJECTIVE: To explore the change of endogenic nerve growth factor (NGF) expression in human glioma cells infected with human cytomegalovirus (HCMV). METHODS: U251 cells were cultured in RPMI 1640 culture medium and infected with HCMV AD169 strain in vitro to establish a cell model of viral infection. Morphologic changes of U251 cells were observed under inverted microscope before and after infection with HCMV. Expression of NGF gene and protein of cells was detected by RT-PCR and Western blotting before and after infection with HCMV. RESULTS: The cytopathic effects of HCMV-infected cells appeared on day 5 after infection. However, differential NGF expression was evident on day 7. NGF expression was decreased significantly in U251 cells on day 7 after infection in comparison with control group (P < 0.05). CONCLUSION: HCMV can down-regulate endogenous NGF levels in human glioma cell line U251.

Nasal cavity-maxillary sinus-pterygopalatine fossa-Meckel's cave: a preliminary anatomic study of an endoscopy-based operative approach.

OBJECTIVE: To provide a new approach for the treatment of tumor in Meckel's cave, by dissecting adjacent structures of the nasal cavity-maxillary sinus-pterygopalatine fossa-Meckel's cave approach. METHODS: Fifteen adult cadaver heads (30 sides) were dissected and the correlated anatomic landmarks were observed, measured and analyzed in an operative route. RESULTS: The approach was divided into 3 steps: entering the maxillary sinus, the later pterygopalatine fossa and the final Meckel's cave. Safe access to Meckel's cave could be achieved by tracing the vidian neurovascular bundles and dissecting the quadrangular space (QS). The distances from the nasal columella to the apertura maxillaries, the sphenopalatine foramen, and the anterior foramen of the pterygoid canal were (44.08+/-2.61) mm, (64.83+/-2.42) mm, and (70.43+/-2.94) mm, respectively. The angles between the horizontal plate of the palatine bone and the link from nasal columella to apertura maxillaries, between the horizontal plate of the palatine bone and the link from nasal columella to sphenopalatine foramen were (38.10+/-2.46) degrees and (26.15+/-2.26) degrees , respectively. CONCLUSION: The endoscopic approach of transnasal maxillary sinus-pterygopalatine fossa-Meckel's cave (ENMPA) is a safe and direct way to access Meckel's cave, and could be employed for the treatment of tumor in Meckel's cave.

[The expression and anti-apoptotic function of HCMV IE2 protein controlled by Tet-On system].

During the infection of host cells, IE2 protein is one of the first and most abundantly expressed products of HCMV genome, which plays an important role in the controlling of cell cycle and apoptosis. But the correlation between expression level and anti-apoptotic activity of IE2 protein is still not clear. In this study, we successfully established a HCMV IE2 protein expression cell line that was controlled by Tet-On system. The effect of IE2 protein on cell apoptosis and the expression of p53 was detected under different condition of induction. Our results showed that the IE2 protein could inhibit cell apoptosis induced by TNF-alpha. Additionally, the anti-apoptotic activity of IE2 protein seemed to be relevant to its expression level. However, we failed to detect any difference of p53 expression between the IE2 protein expression and non-expression cells. These data indicated that the IE2 protein might inhibit cell apoptosis through regulating different signal pathways.

[Human cytomegalovirus inhibits the differentiation of human hippocampus neural stem cells].

The objective of present study is to investigate the effect of human cytomegalovirus (HCMV) infection on human hippocampus neural stem cells NSCs differentiation in vitro, Fetal hippocampus tissue was dissociated mechanically and then cultured in proliferation medium with EGF and bFGF. Immunofluorescence method was used to detect the expression of NSCs marker-Nestin within these cells. Cultured in 10% FBS, NSCs began to differentiate. On the onset of the differentiation, HCMV AD169 (MOI=5) was added into the differentiation medium. After 7 days differentiation, the effect of HCMV infection on NSCs differentiation was observed by detecting the rate of nestin, GFAP and HCMV immediate-early (IE) positive cells with confocal microscopy and immunofluorescence method. The resucts showed most of the cells (passage 4-6 ) were Nestin positive and could differentiate into NSE-positive neurons and GFAP-positive astrocytes. On day 7 postinfection, 86% +/- 12% of infected cells were IE positive. The percentage of Nestin-positive cells was 50% +/- 19% and 93% +/- 10% (t= 6.03, P<0.01)and those of GFAP-positive cells was 81% +/- 11% and 55 +/- 17% (t=3.77, P<0.01) in uninfected and infected cells respectively. These findings indicated that NSCs were HCMV permissive cell and HCMV AD 169 infection suppressed the differentiation of Hippocampus-genetic human neural stem cells into astrocytes.

[Species richness patterns of Picea schrenkiana var. tianschanica communities along an altitudinal gradient at different longitudes in Xinjiang of Northwest China].

The Picea schrenkiana var. tianschanica forest in Zhaosu, Gongliu, Wusu, Urumqi and Hami that covered about 12 longitudes (81 degrees 05'-93 degrees 41.5'E) along the Tianshan Mountains was investigated by using vertical transects to analyze the species richness of the forest along an altitudinal gradient at different longitudes. The results showed that at most longitudinal positions, P. schrenkiana var. tianschanica forest was basically of pure forest, with P. schrenkiana var. tianschanica as the dominant species, very few of other arbor and shrub species, and the species richness of herbs being controlled by the growth of P. schrenkiana var. tianschanica. This forest had the widest distribution, highest canopy density, best growth, and highest stand volume at middle altitudes, resulting in the lowest species richness of herbs. The herb species richness in P. schrenkiana var. tianschanica forest in Tianshan Mountains showed an inverse hump-shaped variation along the altitude, which could be described by the quadratic equation y = ax2 - bx + c (a, b, c > 0).

Identification and Function of MicroRNAs Encoded by Herpesviruses.

MicroRNAs (miRNAs) play important roles in eukaryotes, plants and some viruses. It is increasingly clear that miRNAs-encoded by viruses can affect the viral life cycle and host physiology. Viral miRNAs could repress the innate and adaptive host immunity, modulate cellular signaling pathways, and regulate the expression of cellular and viral genes. These functions facilitate viral acute and persistent infections, and have profound effects on the host cell survival and disease progression. Here, we discuss the miRNAs encoded by herpesviruses, and their regulatory roles involved in virus-host interactions.