bluuebunny/crossref_metadata_2025 huggingface dataset:
Continuous hydrogen production during fermentation of α‐cellulose by the thermophillic bacterium Clostridium thermocellum
Authors: Magnusson Lauren , Cicek Nazim , Sparling Richard , Levin David
Published: September 2008

Continuous hydrogen (H2) production during fermentation of α‐cellulose was established using the thermophillic, anaerobic bacterium Clostridium thermocellum ATCC 27405. The objectives of this work were to characterize growth of C. thermocellum, quantify H2 production and determine soluble end‐product synthesis patterns during fermentation of a cellulosic substrate under continuous culture conditions. A 5 L working volume fermentor was established and growth experiments were maintained for over 3,000 h. Substrate concentrations were varied from 1 to 4 g/L and the feed was introduced with continuous nitrogen gas sparging to prevent clogging of the feed‐line. The pH and temperature of the reactor were maintained at 7.0 and 600°C, respectively, throughout the study. At concentrations above 4 g/L, the delivery of α‐cellulose was impaired due to feed‐line clogging and it became difficult to maintain a homogenous suspension. The highest total gas (H2 plus CO2) production rate, 56.6 mL L−1 h−1, was observed at a dilution rate of 0.042 h−1 and substrate concentration of 4 g/L. Under these conditions, the H2 production rate was 5.06 mmol h−1. Acetate and ethanol were the major soluble end‐products, while lactate and formate were greatly reduced compared to production in batch cultures. Concentrations of all metabolites increased with increasing substrate concentration, with the exception of lactate. Despite a number of short‐term electrical and mechanical failures during the testing period, the system recovered quickly, exhibiting substantial robustness. A carbon balance was completed to ensure that all end‐products were accounted for, with final results indicating near 100% carbon recovery. This study shows that long‐term, stable H2 production can be achieved during direct fermentation of an insoluble cellulosic substrate under continuous culture conditions. Biotechnol. Bioeng. 2009; 102: 759–766. © 2008 Wiley Periodicals, Inc.
Cell cycle progression inEscherichia coliB/r affects transcription of certain genes: Implications for synthetic genome design
Authors: Echtenkamp Patricia L. , Wilson David B. , Shuler Michael L.
Published: September 2008

We propose that transcript levels for some genes are affected by the bacterial cell division cycle and this may be an important factor to consider when designing synthetic bacterial genomes. To test this hypothesis, transcript levels of 58 genes inEscherichia coliB/r A were determined at five times during the cell division cycle. A two‐step ANOVA technique was used to analyze data from custom oligonucleotide microarrays containing genes involved in important cellular processes including central metabolism, macromolecular synthesis, and transport and secretion. Consistent with results previously found inCaulobacter, approximately 17% of the transcript levels were cell cycle dependent. Cell cycle regulation can be divided into two classes: genes displaying increased transcript concentrations following gene replication and genes displaying an increased transcript concentration prior to replication initiation. Transcripts levels forhns,uspA, andzwfwere affected by the cell division cycle, but did not fit well into either class. These results indicate that transcription of a significant fraction of the genome is affected by replication cycle progression. The results also show that both physical gene position and the physiological function of a gene affect when it is transcribed. In addition to the simple association with replication fork progression, other phenomena must be occurring to account for some of our observations. In conclusion, gene position, with regard to the C period, and gene function are important factors to incorporate into design criteria for synthetic bacterial genomes. Biotechnol. Bioeng. 2009; 102: 902–909. © 2008 Wiley Periodicals, Inc.
Biocatalytic polyester acrylation—process optimization and enzyme stability
Authors: Hagström Anna E.V. , Nordblad Mathias , Adlercreutz Patrick
Published: September 2008

An OH‐functional polyester has been acrylated via transesterification of ethyl acrylate, catalyzed by Candida antarctica lipase B (CalB) in two different preparations: Novozym® 435 and immobilized on Accurel® MP1000. The batch process resulted in incomplete acrylation as well as severe degradation of the polyester. A high degree of acrylation was achieved by optimization through the application of low pressure (15 kPa), continuous inflow of ethyl acrylate and continuous distillation to evaporate the by‐product, ethanol. The enzyme preparations displayed good stability with half‐lives of 180 and 324 h for Novozym® 435 and CalB/MP1000, respectively. This translates into product yields of 3600 and 6200 times the weight of the catalyst, indicating that the enzyme will have a marginal impact on the total process cost. Biotechnol. Bioeng. 2009; 102: 693–699. © 2008 Wiley Periodicals, Inc.
Enrichment of fermentation media and optimization of expression conditions for the production of EAK16 peptide as fusions with SUMO
Authors: Satakarni Makkapati , Koutinas Apostolis A. , Webb Colin , Curtis Robin
Published: September 2008

EAK16 (AEAEAKAKAEAKAEAK) belongs to a novel class of self‐assembling peptides, which is being investigated in research and industry. SUMO belongs to the ubiquitin class of proteins and is a promising fusion partner currently in use. In this study, EAK16 peptide fusions with hexa‐histidine tagged SUMO have been constructed using Escherichia coli based pET expression vector. Intracellular expression of the SUMO–EAK16 fusion using LB media has been optimized. Low‐cost complex media (fungal autolysates, wheat and gluten hydrolysates) produced via a novel wheat‐based biorefinery have been used as alternative fermentation media to LB. Shake flask cultures using either enriched LB or complex wheat‐derived media containing 2 g/L of glucose resulted in intracellular SUMO–EAK16 fusion protein production of approximately 250 mg/L fermentation volume which corresponded to 30–35% of the total bacterial protein expressed being the fusion protein. Fusion protein productivities up to five times higher were achieved when using a bioreactor. Biotechnol. Bioeng. 2009; 102: 725–735. © 2008 Wiley Periodicals, Inc.
High‐throughput lensfree imaging and characterization of a heterogeneous cell solution on a chip
Authors: Su Ting‐Wei , Seo Sungkyu , Erlinger Anthony , Ozcan Aydogan
Published: September 2008

A high‐throughput on‐chip imaging platform that can rapidly monitor and characterize various cell types within a heterogeneous solution over a depth‐of‐field of ∼4 mm and a field‐of‐view of ∼10 cm2 is introduced. This powerful system can rapidly image/monitor multiple layers of cells, within a volume of ∼4 mL all in parallel without the need for any lenses, microscope‐objectives or any mechanical scanning. In this high‐throughput lensless imaging scheme, the classical diffraction pattern (i.e., the shadow) of each micro‐particle within the entire sample volume is detected in less than a second using an opto‐electronic sensor chip. The acquired shadow image is then digitally processed using a custom developed “decision algorithm” to enable both the identification of the particle location in 3D and the characterization of each micro‐particle type within the sample volume. Through experimental results, we show that different cell types (e.g., red blood cells, fibroblasts, etc.) or other micro‐particles all exhibit uniquely different shadow patterns and therefore can be rapidly identified without any ambiguity using the developed decision algorithm, enabling high‐throughput characterization of a heterogeneous solution. This lensfree on chip cell imaging platform shows a significant promise especially for medical diagnostic applications relevant to global health problems, where compact and cost‐effective diagnostic tools are urgently needed in resource limited settings. Biotechnol. Bioeng. 2009; 102: 856–868. © 2008 Wiley Periodicals, Inc.
Rapid and quantitative quality control of microarrays using cationic nanoparticles
Authors: Sun Ye , Fan Wenhua , McCann Michael P. , Golovlev Val
Published: September 2008

The fabrication quality of microarrays significantly influences the accuracy and reproducibility of microarray experiments. In this report, we present a simple and fast quality control (QC) method for spotted oligonucleotide and cDNA microarrays. It employs a nonspecific electrostatic interaction of colloidal gold nanoparticles with the chemical groups of DNA molecules and other biomolecules immobilized on the microarray surface that bear positive or negative charges. An inexpensive flatbed scanner is used to visualize and quantify the binding of cationic gold particles to the anionic DNA probes on the microarray surface. An image analysis software was designed to assess the various parameters of the array spots including spot intensity, shape and array homogeneity, calculate the overall array quality score, and save the detailed array quality report in an Excel file. The gold staining technique is fast and sensitive. It can be completed in 10 min and detect less than 1% of the probe amount commonly recommended for microarrays. Compared to the current microarray QC method that utilizes the hybridization of probes with short random sequence oligonucleotides labeled with fluorophore, our gold staining method requires less time for the analysis, reduces the reagent cost, and eliminates the need for the expensive laser scanner. Biotechnol. Bioeng. 2009; 102: 960–964. © 2008 Wiley Periodicals, Inc.
A model of Agrobacterium tumefaciens vacuum infiltration into harvested leaf tissue and subsequent in planta transgene transient expression
Authors: Simmons Christopher W. , VanderGheynst Jean S. , Upadhyaya Shrinivasa K.
Published: September 2008

Agrobacterium‐mediated gene transfer, or agroinfiltration, can be a highly efficient method for transforming and inducing transient transgene expression in plant tissue. The technique uses the innate DNA secretion pathway of Agrobacterium tumefaciens to vector a particular plasmid‐encoded segment of DNA from the bacteria to plant cells. Vacuum is often applied to plant tissue submerged in a suspension of A. tumefaciens to improve agroinfiltration. However, the effects of vacuum application on agroinfiltration and in planta transient transgene expression have not been well quantified. Here we show that vacuum application and release act to drive A. tumefaciens suspension into the interior of leaf tissue. Moreover, the amount of suspension that enters leaves can be predicted based on the vacuum intensity and duration. Furthermore, we show that transient expression levels of an agroinfiltrated reporter gene vary in response to the amount of A. tumefaciens vacuum infiltrated into leaf tissue, suggesting that vacuum infiltration conditions can be tailored to achieve optimal transient transgene expression levels after agroinfiltration. Biotechnol. Bioeng. 2009; 102: 965–970. © 2008 Wiley Periodicals, Inc.
Determination of rank by median absolute deviation (DRMAD): a simple method for determining the number of principal factors responsible for a data matrix
Authors: Malinowski Edmund R.
Published: September 2008

Median absolute deviation (MAD) is a well‐established statistical method for determining outliers. This simple statistic can be used to determine the number of principal factors responsible for a data matrix by direct application to the residual standard deviation (RSD) obtained from principal component analysis (PCA). Unlike many other popular methods the proposed method, called determination of rank by MAD (DRMAD), does not involve the use of pseudo degrees of freedom, pseudo F‐tests, extensive calibration tables, time‐consuming iterations, nor empirical procedures. The method does not require strict adherence to normal distributions of experimental uncertainties. The computations are direct, simple to use and extremely fast, ideally suitable for online data processing. The results obtained using various sets of chemical data previously reported in the chemical literature agree with the early work. Limitations of the method, determined from model data, are discussed. An algorithm, written in MATLAB format, is presented in the Appendix. Copyright © 2008 John Wiley & Sons, Ltd.
Use of cluster separation indices and the influence of outliers: application of two new separation indices, the modified silhouette index and the overlap coefficient to simulated data and mouse urine metabolomic profiles
Authors: Dixon Sarah J. , Heinrich Nina , Holmboe Maria , Schaefer Michele L. , Reed Randall R. , Trevejo Jose , Brereton Richard G.
Published: September 2008

To quantify separate classes, four indices are compared namely the Davies Bouldin index, the silhouette width and two new approaches described in this paper, the modified silhouette width index based on the proportion of objects with a positive silhouette width and the Overlap Coefficient. Four sets of simulated datasets are described, each in turn, consisting of 15 sets of data of varying degrees of overlap, and differing in the nature of outliers. Three experimental datasets consisting of the gas chromatography mass spectrometry of extracts from mouse urine obtained to study the effect of different environmental (stress), physiological (diet) and developmental (age) factors on their metabolic profiles are also described. The paper discusses the robustness of each approach to outliers, and to allow assessment of class separation for each index. The two modifications protect against outliers. Copyright © 2008 John Wiley & Sons, Ltd.
Accuracy of magnetic resonance imaging in predicting residual disease in patients treated for stage IB2/II cervical carcinoma with chemoradiation therapy
Authors: Vincens Etienne , Balleyguier Corinne , Rey Annie , Uzan Catherine , Zareski Elise , Gouy Sebastien , Pautier Patricia , Duvillard Pierre , Haie‐Meder Christine , Morice Philippe
Published: September 2008

BACKGROUND

The evaluation of residual disease (RD) after chemoradiation therapy (CRT) in stage IB2/II cervical carcinoma conventionally is based on a clinical examination and magnetic resonance imaging (MRI) performed 3 to 8 weeks after the end of treatment. Very few studies have correlated MRI and histologic findings specifically in cervix cancer. This was the objective of the current study.

METHODS

A retrospective review was undertaken of patients who fulfilled the following inclusion criteria: 1) stage IB2/II cervical cancer, 2) external radiotherapy (45 grays [Gy]) was given with concomitant chemotherapy followed by uterovaginal brachytherapy (15 Gy), 3) MRI studies were obtained between 3 weeks and 8 weeks after brachytherapy, and 4) completion surgery included at least a hysterectomy. Postsurgical histologic findings and MRI results were compared.

RESULTS

Forty‐four patients who were treated between 2003 and 2006 fulfilled all inclusion criteria. Twelve patients (27%) had achieved a radiologic complete response, 16 patients (36.5%) had unclassified lesions (RD or ‘fibrosis’), and 16 patients (36.5%) had radiologic residual tumor. According to the histologic results, 19 patients (43%) had no RD, 10 patients (23%) had inframillimeter RD, 2 patients (5%) had RD that measured <1 cm, and 13 patients (29%) had RD that measured >1 cm. The sensitivity and specificity of MRI in evaluating RD were 80% and 55%, respectively.

CONCLUSIONS

The current results indicated that the evaluation of RD 3 to 8 weeks after CRT with MRI is difficult, and the risk of false‐positive results is high. Another radiologic procedure or a more technologically advanced MRI technique, such as diffusion‐weighted MRI, should be evaluated. Cancer 2008. © 2008 American Cancer Society.
Multidisciplinary treatment of brain metastases derived from clear cell renal cancer incorporating stereotactic radiosurgery
Authors: Samlowski Wolfram E. , Majer Martin , Boucher Kenneth M. , Shrieve Annabelle F. , Dechet Christopher , Jensen Randy L. , Shrieve Dennis C.
Published: September 2008

BACKGROUND

Brain metastases are a frequent complication in patients with metastatic clear cell renal cancer. Survival after whole‐brain radiotherapy (WBRT) is disappointing. A retrospective analysis of multimodality treatment was performed in patients who had received linear accelerator (LINAC)‐based stereotactic radiosurgery (SRS).

METHODS

Thirty‐two patients underwent SRS‐based treatment for 71 metastatic foci between 2000 and 2006. All patients had a Karnofsky performance status ≥70 and all 32 patients had extracranial metastatic disease (Radiation Therapy Oncology Group recursive partitioning analysis [RPA] Class 2). Survival was calculated from the time of diagnosis of brain metastases. The minimum potential follow‐up was 1 year after SRS. Univariate and multivariate analysis of potential prognostic factors affecting survival was performed.

RESULTS

Twenty‐six patients required only 1 SRS treatment (84%) to achieve central nervous system (CNS) control, whereas 5 patients received 2 to 3 treatments (16%). The median survival of renal cancer patients from the diagnosis of brain metastases was 10.1 months (95% confidence interval, 6.4‐14.8 months). One‐year and 3‐year survival rates were 43% and 16%, respectively. The addition of surgery or WBRT did not appear to prolong survival. Immunotherapy after control of brain metastases with SRS appeared to result in significantly improved survival. Survival was also found to be strongly influenced by prognostic stratification of metastatic disease using Motzer or modified risk criteria.

CONCLUSIONS

The results of the current study demonstrated that SRS‐based treatment of patients with up to 5 brain metastases from clear cell renal cancer is feasible and results in excellent CNS control. Survival beyond 3 years from the time of diagnosis of brain metastases was achievable in 16% of patients and was associated with the use of systemic immunotherapy with interleukin‐2 and interferon but not antiangiogenic agents. Cancer 2008. © 2008 American Cancer Society.
Targeted therapy in prostate cancer—Are we our own worst enemy?
Authors: Dawson Nancy A.
Published: September 2008

Targeted therapies are being assessed in castrate‐resistant prostate cancer, but their development is slow, and the correct targets are not always clear. Given the propensity for prostate cancer to metastasize to bone, therapies that target the bone microenvironment are a logical choice.
Surgical resection of the primary tumor, chest wall control, and survival in women with metastatic breast cancer
Authors: Hazard Hannah W. , Gorla Seema R. , Scholtens Denise , Kiel Krystyna , Gradishar William J. , Khan Seema A.
Published: September 2008

BACKGROUND

Among women presenting with de novo stage IV breast cancer, 35% to 60% undergo local therapy, presumably to avoid uncontrolled chest wall disease. Several studies suggest that resection of the primary tumor may prolong survival, but chest wall outcome data are notably lacking. The authors reviewed chest wall status, time to first progression (TTFP), and overall survival (OS) in this group of women.

METHODS

Women presenting at the Lynn Sage Breast Center (1995‐2005) with an intact primary tumor and stage IV breast cancer or postoperative diagnosis of distant metastases were identified. Logistic regression and Cox proportional hazards models, adjusted for relevant covariates, were used to examine associations between surgical treatment and chest wall status, TTFP, and OS.

RESULTS

Of 111 eligible women, 47 (42%) underwent early resection of the primary tumor. Chest wall status was available for 103 women. Local control was maintained in 36 of 44 (82%) patients in the surgical group versus 20 of 59 (34%) patients without surgery (P = .001). TTFP was prolonged in the surgical group (adjusted hazards ratio [HR], 0.493; P = .015). The adjusted HR for OS in the surgical group was 0.798 (P = .520). Chest wall control was associated with improved OS regardless of whether surgical resection of the tumor was performed (HR, 0.415; P < .0002).

CONCLUSIONS

These data support the notion that improved local control may play a role in improving outcomes in women with stage IV breast cancer, and resection of in‐breast tumors can help to achieve this. A randomized trial is needed to rule out selection bias as an explanation for these findings. Cancer 2008. © 2008 American Cancer Society.
Parallels between the development of therapeutic drugs and cancer health disparity programs
Authors: Chu Kenneth C. , Chen Moon S. , Dignan Mark B. , Taylor Emmanuel , Partridge Edward ,   Community Network Program Principal Investigators
Published: September 2008

BACKGROUND

There are analogies between the development of therapeutic drugs for cancer and the development of interventions for reducing cancer health disparities. In both cases, it can take between 12 and 15 years for the benefits to become apparent.

METHODS

The initial preclinical phase of drug development is analogous to the development of community partnerships and helping the community learn about cancer. The preclinical phase of in vitro and in vivo testing is analogous to identifying the disparities in the community. Then clinical testing begins with phase 1, toxicity, and dose‐establishing studies. Analogously, community‐based participatory research is used to develop disparities–reducing interventions (DRIs) within the community.

RESULTS

The phase 2 clinical studies to determine whether the drug has activity are analogous to the DRI being implemented in the community to determine whether it can cause behavioral changes that will reduce cancer health disparities. If a drug passes phase 1 and 2 studies, phase 3 clinical trials are initiated. These are controlled studies to examine the efficacy of the drug. The similar activity for disparities research is to determine whether the DRI is better than the current standard/usual practice in controlled trials. If the drug is beneficial, the final phase is the dissemination and adoption of the drug. Analogously in disparities, if the DRI is beneficial, it is disseminated and is culturally adapted to other racial/ethnic groups and finally adopted as standard practice.

CONCLUSIONS

The process of creating an effective DRI can be envisioned to have 4 stages, which can be used to aid in measuring the progress being made in reducing cancer health disparities. Cancer 2008. Published 2008 by the American Cancer Society.
Acute Renal Failure After Biopsy of a Transplant Kidney
Authors: Burhan Umber , Schleifer Charles R.
Published: September 2008

Percutaneous biopsies of renal allografts are performed frequently, mostly for clinical indications and occasionally as protocol biopsies in some transplant centers. A rare complication of percutaneous needle biopsy is the development of a subcapsular hematoma, also known as Page kidney. The authors report such a case in a renal allograft following biopsy. What makes this case remarkable is the spontaneous resolution of the hematoma without surgical intervention, with reasonable recovery of allograft function, allowing cessation of dialysis.
Assessment of Potential Living Kidney Donors: Options for Increasing Donation
Authors: Akoh Jacob A. , Stacey Sarah
Published: September 2008

OBJECTIVE

The aim of this study was to report the outcome of assessments of potential donors within a single center and to identify what factors can be modified to increase donation.

METHODS

Between January 2003 and February 2008, 364 potential donors (237 related, 120 unrelated, and 7 altruistic) were referred to the stages assessment process for living kidney donation. Records of assessment and outcomes were entered prospectively into a spreadsheet maintained by the Living Donor Transplant Coordinator.

RESULTS

Of the 143 potential donors proceeding to stage 2 assessments, 25 were excluded for medical and other reasons, and 46 are currently being assessed. Sixty‐five donor nephrectomies were performed, with 7 awaiting surgery, giving a donation rate of 21.3% (65 of 305) and a potential donation rate of 24% (72 of 305) if all 7 successfully donate. The reasons for exclusion from stage 1 assessment included immunological considerations, multiple donors, and cadaveric transplantation of an intended recipient.

DISCUSSION

Introduction of paired exchange and non‐directed donation, ABO incompatible transplantation, or desensitization of potential recipients will improve living donation rates. Potential donors withdrawing either because of another more suitable donor or their recipient being transplanted could be made aware (no obligations) of the option of altruistic non‐directed donation.
Prevalence of Occult Hepatitis B Infection in Hemodialysis Patients
Authors: Ersoy Osman , Yilmaz Rahmi , Arici Mustafa , Turgan Cetin , Bayraktar Yusuf
Published: September 2008

BACKGROUND

The presence of low‐level hepatitis B virus (HBV) DNA has been demonstrated in some hepatitis B surface antigen (HBsAg)–negative patients. This HBsAg‐negative but HBV DNA–positive case has been named occult hepatitis B virus (OHBV) infection. The aim of this study was to determine the prevalence of OHBV in patients on maintenance hemodialysis.

METHODS

Eighty patients who were on dialysis and were previously detected as HBsAg negative were recruited in the study. Thirty‐two patients (40%) were female, and the mean age was 51.56 ± 14.16 years (21–78 years). HBV DNA was searched for by the polymerase chain reaction method in the sera of all patients.

RESULTS

Sixty‐eight patients (85%) positive for at least 1 of the serological markers used for hepatitis B, 3 patients (3.75%) positive for only antibody to hepatitis B core antigen, and 33 patients (41.25%) positive for only antibody to HBsAg were identified. The prevalence of OHBV infection was found to be 1.25%.

CONCLUSIONS

Even though the prevalence of OHBV infection was found to be low in patients undergoing dialysis, searching for HBV DNA, if possible, will be useful because there is contagiousness in OHBV infection, and activation of OHBV infection has been known to occur after immunosuppressive treatment.
Seismic design and response evaluation of unbonded post‐tensioned hybrid coupled wall structures
Authors: Kurama Yahya C. , Shen Qiang
Published: September 2008

This paper presents an analytical investigation on the seismic design and response of coupled wall structures that use unbonded post‐tensioned steel coupling beams. Both monolithic cast‐in‐place reinforced concrete wall piers and precast concrete wall piers are considered. Steel top and seat angles are used at the coupling beam ends for energy dissipation. The seismic design of prototype structures to achieve target displacement‐based performance objectives is evaluated based on nonlinear static and dynamic time history analyses. Additional recommendations are provided on shear design. Comparisons with ‘conventional’ structures that use embedded steel coupling beams as well as isolated walls with no coupling are provided. The results indicate that while the peak lateral displacements of unbonded post‐tensioned coupled wall structures are larger than the peak displacements of structures with embedded beams, the residual displacements are significantly reduced as a result of the restoring effect of the post‐tensioning steel. Copyright © 2008 John Wiley & Sons, Ltd.
Evaluation on in vitro biocompatibility of silk fibroin‐based biomaterials with primarily cultured hippocampal neurons
Authors: Tang Xin , Ding Fei , Yang Yumin , Hu Nan , Wu Hong , Gu Xiaosong
Published: September 2008

Silk fibroin‐based biomaterials have recently found increasing applications in the tissue‐engineering field including the generation of artificial nerve guides for peripheral nerve repair. The aim of this study was to investigate the suitability of silk fibroin as a candidate biomaterial for central nervous system (CNS) therapy. We found that substrates made up of silk fibroin fibers supported the survival and growth of the attached hippocampal neurons by using morphological observation. We also cultured the hippocampal neurons in silk fibroin extract for different times, and observed no significant difference occurring in their morphology, cell viability for these cultured hippocampal neurons as compared to those cultured in plain neuronal culture medium. Moreover, immunocytochemistry, RT‐PCR, and Western blot analysis revealed that no significant difference was found in mRNA or protein levels of the growth‐associated molecules, such as brain‐derived neurotrophic factor, growth‐associated protein‐43, neurofilament, nerve growth factor, and nerve growth factor‐receptor P75, between the hippocampal neurons cultured in the silk fibroin extract and in plain neuronal culture medium. Taken together, all the results demonstrate that silk fibroin has good biocompatibility with primarily cultured hippocampal neurons without any significant cytotoxic effects on their cell phenotype and functions, suggesting a potential possible use of silk fibroin for preparing the tissue‐engineered nerve guides or drug delivery vehicles to treat CNS injuries or diseases. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res, 2009
Macroporous hydroxyapatite scaffolds for bone tissue engineering applications: Physicochemical characterization and assessment of rat bone marrow stromal cell viability
Authors: Oliveira Joaquim M. , Silva Simone S. , Malafaya Patricia B. , Rodrigues Marcia T. , Kotobuki Noriko , Hirose Motohiro , Gomes Manuela E. , Mano Joao F. , Ohgushi Hajime , Reis Rui L.
Published: September 2008

In this work, a new methodology is reported for developing hydroxyapatite (HA) scaffolds using an organic sacrifice template. The novelty of work consists of possibility of obtaining porous and highly interconnected scaffolds mimicking the sacrificial component. Our purpose consisted of evaluating the physicochemical properties of the HA scaffolds by means of Fourier transform infra‐red spectroscopy, X‐ray diffraction analysis, and scanning electron microscopy (SEM) attached with an X‐ray detector. The HA scaffolds obtained possess a porosity of ∼70%, and macropores diameter in the range of 50–600 μm. In contrast, results regarding the microcomputed tomography analysis have demonstrated both high pore uniformity and interconnectivity across the scaffolds. The compressive strength of the HA scaffolds was found to be 30.2 ± 6.0 MPa. Bioactivity of the HA scaffolds was assessed by immersion into a simulated body fluid solution, in vitro. SEM observations have showed a deposition of apatite on the surface of the HA scaffolds, with a “cauliflower‐like” morphology after 1 day, and tend to be more pronounced with the immersion time. The changes in calcium and phosphorus concentration were monitored by inductively‐coupled plasma optical emission spectrometry. Cytotoxicity of the HA scaffolds was preliminarily investigated by carrying direct observation of mouse fibroblasts cells (L929 cell‐line) death in the inverted microscope, and then cell viability was determined by means of carrying out a MTS assay. Complementarily, a luminescent cell viability assay based on the quantification of adenosine triphosphate was performed using rat bone marrow stromal cells (RBMSCs). A LIVE/DEAD assay and SEM analysis allowed the visualization of the RBMSCs adhesion and proliferation on the surface of the HA scaffolds. According to the results obtained from 3D architecture, mechanical properties, biocompatibility, and adhesion tests, it is suggested that HA scaffolds has potential to find applications in bone tissue engineering scaffolding. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res, 2009
Synthesis and characterization of semi‐interpenetrating polymer networks based on polyurethane and N‐isopropylacrylamide for wound dressing
Authors: Thimma Reddy Thatiparti , Kano Arihiro , Maruyama Atsushi , Hadano Michiko , Takahara Atsushi
Published: September 2008

Thermosensitive semi‐interpenetrating polymer networks (semi‐IPNs) composed of crosslinked poly(N‐isopropylacrylamide) (PNiPAAm) and linear segmented polyurethane urea (SPUU) were synthesized via thermal initiated free radical polymerization. Synthesized semi‐IPNs of various compositions were characterized by Fourier transform infrared spectroscopy, water equilibrium swelling at different temperatures, drug lading, drug release, cell adhesion, and detachment. The semi‐IPN films of all the compositions were transparent in dry state and negative thermosensitivity in their swelling ratio, that is, lower swelling levels with increasing temperature. The drug release study revealed that the rate of drug release is fast in case of pure SPUU compared to PNiPAAm and semi‐IPN film. Drug release depended mainly on solubility of the drugs and physical networks between SPUU and PNiPAAm. Finally NIH3T3 cells were seeded on the semi‐IPN films and found that cells were securely attached and proliferated to confluence. Upon cooling, cells were detached from the semi‐IPN films. Therefore, the semi‐IPN films may be good candidate materials for wound dressing applications. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009
Demineralized bone matrix as an osteoinductive biomaterial andin vitropredictors of its biological potential
Authors: Katz Jordan M. , Nataraj Chandra , Jaw Rebecca , Deigl Elizabeth , Bursac Predrag
Published: September 2008

The osteoinductivity of demineralized bone matrix (DBM) varies from donor to donor as a result of varying levels of multiple growth factors, matrix integrity, and artifacts from material processing. Manyin vitroassays are currently used for screening the osteoinductivity of DBM. The objectives of this study were to determine the correlation of specific growth factors andin vitromitotic stimulation toin vivoectopic bone formation capacity with a large number of DBM samples. Samples were assayed using ELISA methods for BMP‐2/4 and TGF‐β1 (n= 304) and cell proliferation using SAOS‐2 osteoblasts (n= 239). All samples were then implanted intramuscularly in the abdomen of nude rats. Allin vitroassays showed significant variability for any particular level of ostoinductivity determined byin vivomodel. A significant, but only very weak, positive correlation toin vivoresults was found for TGF‐β1 (r2= 0.016), BMP 2/4 (r2= 0.065), and SAOS‐2 cell proliferation (r2= 0.053). The results of this study amplify the notion that a multitude of factors and their relative interplay, rather than a single factor are likely to determine the potency of any particular lot of DBM. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009
In vivo evaluation of anodic TiO2 nanotubes: An experimental study in the pig
Authors: von Wilmowsky Cornelius , Bauer Sebastian , Lutz Rainer , Meisel Mark , Neukam Friedrich Wilhelm , Toyoshima Takeshi , Schmuki Patrik , Nkenke Emeka , Schlegel Karl Andreas
Published: September 2008

Because of their ability to mimic the dimensions of constituent components of natural bone and the possibility to serve as a gene and drug‐delivery carrier, nanotubes seem to be a promising coating for medical implants. Aim of this study was to investigate the effects of a TiO2 nanotube structured surface on periimplant bone formation in vivo when compared with an untreated standard titanium surface. Twenty‐five titanium implants covered with an ordered TiO2 nanotube layer with an individual tube diameter of 30 nm and 25 commercially pure titanium (cp‐Ti) implants were placed in the frontal skull of 25 domestic pigs. To evaluate the effects of the nanotube structured implants on the periimplant bone formation, bone‐implant contact (BIC), and immunohistochemistry analysis were performed at day 3, 7, 14, 30, and 90. Evaluating immunohistochemistry, a significantly higher collagen type‐ I expression occurred at day 7 (p = 0.003), day 14 (p = 0.016), and day 30 (p = 0.044), for the nanostructured implants in comparison with the control group. It could be found that a nanotube structured implant surface with a diameter of 30 nm does influence bone formation and bone development by enhancing osteoblast function. SEM evaluation of the specimen surfaces revealed that the nanotube coatings do resist shearing forces that evoked by implant insertion. Because of their simple, low cost, flexible manufacturing and the possibility for the usage as drug or growth factor delivery system, nanotubes seem to be a promising method for future medical implant coatings. © 2008 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2009
Theoretical studies of the reactions of CF3CHCLOCHF2/CF3CHFOCHF2 with OH radical and Cl atom and their product radicals with OH
Authors: Yang Lei , Liu Jing‐Yao , Wan Su‐Qin , Li Ze‐Sheng
Published: September 2008

The mechanisms and dynamics studies of the OH radical and Cl atom with CF3CHClOCHF2 and CF3CHFOCHF2 have been carried out theoretically. The geometries and frequencies of all the stationary points are optimized at the B3LYP/6‐311G(d,p) level, and the energy profiles are further refined by interpolated single‐point energies (ISPE) method at the G3(MP2) level of theory. For each reaction, two H‐abstraction channels are found and four products (CF3CHFOCF2, CF3CFOCHF2, and CF3CHClOCF2, CF3CClOCHF2) are produced during the above processes. The rate constants for the CF3CHClOCHF2/CF3CHFOCHF2 + OH/Cl reactions are calculated by canonical variational transition‐state theory (CVT) within 200–2000 K, and the small‐curvature tunneling is included. The total rate constants calculated from the sum of the individual rate constants and the branching ratios are in good agreement with the experimental data. The Arrhenius expressions for the reactions are obtained. Our calculation shows that the substitution of Cl by F decreases the reactivity of CF3CHClOCHF2 toward OH and Cl. In addition, the mechanisms of subsequent reactions of product radicals and OH radical are further investigated at the G3(MP2)//B3LYP/6‐311G(d,p) level, and the main products are predicted in the this article. © 2008 Wiley Periodicals, Inc. J Comput Chem 2009
Role of the variable active site residues in the function of thioredoxin family oxidoreductases
Authors: Carvalho Alexandra T. P. , Fernandes Pedro A. , Swart Marcel , Van Stralen Joost N. P. , Bickelhaupt F. Matthias , Ramos Maria J.
Published: September 2008

The enzymes of the thioredoxin family fulfill a wide range of physiological functions. Although they possess a similar CXYC active site motif, with identical environment and stereochemical properties, the redox potential and pKa of the cysteine pair varies widely across the family. As a consequence, each family member promotes oxidation or reduction reactions, or even isomerization reactions. The analysis of the three‐dimensional structures gives no clues to identify the molecular source for the different active site properties. Therefore, we carried out a set of quantum mechanical calculations in active site models to gain more understanding on the elusive molecular‐level origin of the differentiation of the properties across the family. The obtained results, together with earlier quantum mechanical calculations performed in our laboratories, gave rise to a consistent line of evidence, which points to the fact that both active site cysteines play an important role in the differentiation. In contrary to what was assumed, differentiation is not achieved through a different stabilization of the solvent exposed cysteine but, instead, through a fine tuning of the nucleophilicity of both active site cysteines. Reductant enzymes have both cysteine thiolates poorly stabilized, oxidant proteins have both cysteine thiolates highly stabilized, and isomerases have one thiolate (solvent exposed) poorly stabilized and the other (buried) thiolate highly stabilized. The feasibility of shifting the chemical equilibrium toward oxidation, reduction, or isomerization only through subtle electrostatic effects is quite unusual, and it relies on the inherent thermoneutrality of the catalytic steps carried out by a set of chemically equivalent entities all of which are cysteine thiolates. Such pattern of stabilization/destabilization, detected in our calculations is fully consistent with the observed physiological roles of this family of enzymes. © 2008 Wiley Periodicals, Inc. J Comput Chem, 2009
Refining the description of peptide backbone conformations improves protein simulations using the GROMOS 53A6 force field
Authors: Cao Zanxia , Lin Zhixiong , Wang Jun , Liu Haiyan
Published: September 2008

The GROMOS 53A6 parameter sets have been shown to reproduce an extensive range of thermodynamics data in condensed phase (Oostenbrink et al., J Comput Chem 2004, 25, 1656), mainly due to the reoptimized nonbonded interactions. Here, we derive refinements for the descriptions of peptide backbone conformations for thisparameter set. A two‐dimensional adaptive umbrella sampling procedure was employed to determine the free energy surfaces of model alanine and glycine dipeptides in solution to high accuracy, with sampling errors below 0.8 kJ mol−1 for relative free energies between major minima. Comparisons of these surfaces with quantum mechanical ones and conformation distributions in protein crystal structures indicated that refined treatments of backbone torsional angle terms are necessary. The high accuracy of the computed free energy surfaces allowed us to consider two types of corrections, one numerically and exactly reproducing the quantum mechanical results, and the other using small analytical terms to correct major deficiencies for the dipeptide systems. In addition, aiming at improving the directionality of backbone–backbone hydrogen bonds, we optimized and tested an off‐center charge model for the peptide backbone carbonyl oxygen. Extensive molecular dynamics simulations of five proteins and two peptides in solution indicate that refined treatments of backbone dihedral angles lead to substantial improvements of the simulations. Being much simpler, the analytical terms perform as good as or even slightly better than the exact numerical corrections. Although using off‐center charges brought some improvements, the directionality of hydrogen bonds have not been significantly improved. © 2008 Wiley Periodicals, Inc. J Comput Chem 2009
Performance of density functional models to reproduce observed 13Cα chemical shifts of proteins in solution
Authors: Vila Jorge A. , Baldoni Héctor A. , Scheraga Harold A.
Published: September 2008

The purpose of this work is to test several density functional models (namely, OPBE, O3LYP, OPW91, BPW91, OB98, BPBE, B971, OLYP, PBE1PBE, and B3LYP) to determine their accuracy and speed for computing 13Cα chemical shifts in proteins. The test is applied to 10 NMR‐derived conformations of the 76‐residue α/β protein ubiquitin (protein data bank id 1D3Z). With each functional, the 13Cα shielding was computed for 760 amino acid residues by using a combination of approaches that includes, but is not limited to, treating each amino acid X in the sequence as a terminally blocked tripeptide with the sequence Ac‐GXG‐NMe in the conformation of the regularized experimental protein structure. As computation of the 13Cα chemical shifts, not their shielding, is the main goal of this work, a computation of the 13Cα shielding of the reference, namely, tetramethylsilane, is investigated here and an effective and a computed tetramethylsilane shielding value for each of the functionals is provided. Despite observed small differences among all functionals tested, the results indicate that four of them, namely, OPBE, OPW91, OB98, and OLYP, provide the most accurate functionals with which to reproduce observed 13Cα chemical shifts of proteins in solution, and are among the faster ones. This study also provides evidence for the applicability of these functionals to proteins of any size or class, and for the validation of our previous results and conclusions, obtained from calculations with the slower B3LYP functional. © 2008 Wiley Periodicals, Inc. J Comput Chem, 2009
One‐electron spin‐orbit contribution by effective nuclear charges
Authors: Chiodo Sandro G. , Russo Nino
Published: September 2008

Effective nuclear charges of the main group elements from the second up to the fifth row have been developed for the one‐electron part of the spin‐orbit (SO) coupling Hamiltonian. These parameters, suitable to be used for SO calculations of large molecular systems, provide a useful and remarkably good approximation to the full SO Hamiltonian. We have derived atomic effective nuclear charges by fitting procedure. Computed fine‐structure splitting (FSS) of the doublet and triplet II states of AH species (A is one of the abovementioned elements) have been chosen for this purpose. We have adopted the noniterative scheme, previously reported, according to which SO contributions can be calculated through direct coupling between the II states. The latter have been optimized at B3LYP level using DZVP basis sets. As surrogates for a large number of possible applications, we have widely employed the empirical parameters to compute II‐FSSs of diatomic species for which experimental data are available. © 2008 Wiley Periodicals, Inc. J Comput Chem, 2009
Gadd45 modulation of intrinsic and extrinsic stress responses in myeloid cells
Authors: Hoffman Barbara , Liebermann Dan A.
Published: September 2008

Gadd45 proteins modulate signaling in response to physiological and environmental stressors. Expression of gadd45 genes is rapidly induced by different stressors, including differentiation‐inducing cytokines and genotoxic stress. Induction of gadd45 genes at the onset of myeloid differentiation suggested that Gadd45 protein(s) play a role in hematopoiesis, yet no apparent abnormalities were observed in either the bone marrow (BM) or peripheral blood compartments of mice deficient for either gadd45a or gadd45b. However, under conditions of hematological stress, including acute stimulation with cytokines, myelo‐ablation and inflammation, both gadd45a‐deficient and gadd45b‐deficient mice exhibited deficiencies. This is discussed within the context of what is known about Gadd45 proteins in stress signaling, hematopoietic development and the innate immune response. Furthermore, myeloid enriched BM cells from gadd45a and gadd45b deficient mice were observed to be more sensitive to ultraviolet radiation (UVC), VP‐16 and daunorubicin (DNR) induced apoptosis compared to wild‐type (WT) cells, displaying defective G2/M arrest following exposure to UVC and VP‐16, but not to DNR. Novel mechanisms that mediate the pro‐survival functions of Gadd45 in hematopoietic cells following UV irradiation were demonstrated, involving activation of the Gadd45a‐p38‐NF‐κB survival pathway and Gadd45b mediated inhibition of the stress response MKK4‐JNK apoptotic pathway. The ramifications regarding the pathogenesis of different leukemias and the response of normal and malignant hematopoietic cells to chemo‐ and radiation‐therapy, as well as other challenges to the hematopoietic compartment, are discussed. J. Cell. Physiol. 218: 26–31, 2009. © 2008 Wiley‐Liss, Inc.