bluuebunny/crossref_metadata_2025 huggingface dataset:
Kinetic study on the thermal dehydration of CaCO3 · H2O by the master plots method
Authors: Wanjun Tang , Donghua Chen , Cunxin Wang
Published: March 2006

Thermal dehydration process of calcium oxalate monohydrate, CaC2O4 · H2O, was reinvestigated from a viewpoint of reaction kinetics. On the basis of data of thermogravimetry, kinetic analysis was performed under nonisothermal conditions using an integral composite procedure, which includes a integral isoconversional method and an integral master plots method. The results of the integral isoconversional method of TGA data at various heating rates suggested that dehydration of CaC2O4 · H2O followed a single step with an activation energy of 99.02 ± 2.61 kJ/mol, and from the master plots method, the reaction was described by an Rn model. Finally, it was estimated that the pre‐exponential factor A = (2.82 ± 1.81) ×109 s−1, and the kinetic exponent n = 2.3 ± 0.2. The results of nonisothermal kinetic analysis of the thermal dehydration reactions of CaC2O4 · H2O suggested that the integral composite procedure be very successful in evaluating kinetic parameters and describing kinetic model of solid‐state reactions. © 2006 American Institute of Chemical Engineers AIChE J, 2006
Research on stored biological samples: Views of African American and White American cancer patients
Authors: Pentz Rebecca D. , Billot Laurent , Wendler David
Published: March 2006

Proposals on consent for research with biological samples should be informed by empirical studies of individuals' views. Studies to date queried mostly white research subjects. The aim of this study was to compare the views of two groups of patients: cancer patients at a university clinic (Winship Cancer Institute at Emory Healthcare) and cancer patients at an inner city county hospital (Grady) who were given the option of tissue banking. Overall, 315/452 (70%) patients completed the survey. The Grady cohort was 86% African American; the Winship cohort was 82% White. The vast majority (95%) of individuals in both cohorts agreed to provide a biological sample for future research. Both cohorts were willing for their samples to be used to study cancer and other diseases, including Alzheimer disease. Few participants preferred to control the disease to be studied (10%) or wished to be contacted again for consent for each future research project (11%). In our sample, almost all clinical patients, regardless of site of care, ethnicity or socioeconomic status, were willing to provide a biological sample for research purposes and allow investigators to determine the research to be done without contacting the patients again. These findings support the recommendation to offer individuals a simplified consent with a one‐time binary choice whether to provide biological samples for future research. © 2006 Wiley‐Liss, Inc.
The CACNA1A and ATP1A2 genes are not involved in dominantly inherited migraine with aura
Authors: Kirchmann Malene , Thomsen Lise Lykke , Olesen Jes
Published: February 2006

Epidemiological studies indicate that migraine with typical aura (MA) has a major genetic component but the genes for MA have not been identified. However, the autosomal dominantly inherited familial hemiplegic migraine (FHM) is often caused by mutations in the CACNA1A or ATP1A2 genes. The aim of the study was to investigate if the CACNA1A or ATP1A2 genes are involved in MA with an apparently autosomal dominant mode of inheritance. From a clinic population diagnosed by a trained physician we recruited 34 extended families (comprising 174 MA patients) with an apparently autosomal dominant mode of inheritance of MA. We performed a linkage analysis of 161 of 174 MA patients and 79 unaffected relatives using a framework marker set of 44 markers for chromosome 1 and 22 markers for chromosome 19. Linkage analysis was made with a non‐parametric or autosomal dominant parametric model, either allowing for heterogeneity or not, using an affected only analysis. We identified no linkage to CACNA1A and ATP1A2 loci on chromosome 19 or 1, respectively. Additionally, at least two patients from each family and 92 healthy, unrelated controls were selected for a sequence analysis. We sequenced the 48 exons of CACNA1A and the 23 exons of ATP1A2, including promoter and flanking intron sequences. No polymorphism was identified in the CACNA1A or ATP1A2 genes with a strong correlation to MA. Our study shows that the CACNA1A or ATP1A2 genes are probably not involved in MA. To identify the genes involved in the common forms of migraine, future genetic studies should focus on MA and migraine without aura (MO) and not FHM. © 2006 Wiley‐Liss, Inc.
Failure to confirm genetic association between schizophrenia and markers on chromosome 1q23.3 in the region of the gene encoding the regulator of G‐protein signaling 4 protein (RGS4)
Authors: Rizig Mie A. , McQuillin Andrew , Puri Vinay , Choudhury Khalid , Datta Susmita , Thirumalai Srinivasa , Lawrence Jacob , Quested Digby , Pimm Jonathan , Bass Nicholas , Lamb Graham , Moorey Helen , Badacsonyi Allison , Kelly Katie , Morgan Jenny , Punukollu Bhaskar , Kandasami Gomathinayagam , Curtis David , Gurling Hugh
Published: February 2006

The chromosome 1q23.3 region, which includes the RGS4 gene has been implicated in genetic susceptibility to schizophrenia by two linkage studies with lod scores of 6.35 and 3.20 and with positive lod between 2.00 and 3.00 scores in several other studies. Reduced post mortem RGS4 gene expression in the brain of schizophrenics was reported as well as positive allelic association between markers at the RGS4 gene locus and schizophrenia. We have attempted to replicate the finding of allelic association with schizophrenia in a UK based sample of 450 subjects with schizophrenia and 450 supernormal controls. We genotyped the same SNP marker alleles investigated in the earlier studies and also a di‐nucleotide (GT)14 repeat microsatellite marker, which was 7 kb distal to RGS4. In the new UK sample there was no evidence for allelic or haplotypic association between RGS4 markers and schizophrenia. This might reflect genetic heterogeneity between the population samples, genotyping or other methodological problems. The finding weakens the evidence that mutations or variation in the RGS4 gene have an effect on schizophrenia susceptibility. © 2006 Wiley‐Liss, Inc.
Metallvinylidene und ‐allenylidene in der Katalyse
Authors: Bruneau Christian , Dixneuf Pierre H.
Published: March 2006

Das Auftreten einer katalytischen Metallvinylidenspezies wurde erstmals im Jahre 1986 vorgeschlagen, um die direkte regioselektive Bildung von Vinylcarbamaten aus terminalen Alkinen, Kohlendioxid und Aminen zu erklären. Seit diesem ersten Bericht haben sich diverse Metallvinylidene und Metallallenylidene, die die Schlüsselintermediate bei der Aktivierung sind, als sehr nützliche Reagentien für viele Alkinreaktionen erwiesen und außerdem zum gezielten Entwurf von neuen katalytischen Reaktionen beigetragen. Derweil sich diese Entdeckung nun zum zwanzigsten Male jährt, scheint der Zeitpunkt geeignet, auf die Fortschritte zurückzublicken, die seither bei der Anwendung von metallorganischen Vinylidenen und Allenylidenen in der Katalyse erzielt worden sind.
Metal Vinylidenes and Allenylidenes in Catalysis: Applications in Anti‐Markovnikov Additions to Terminal Alkynes and Alkene Metathesis
Authors: Bruneau Christian , Dixneuf Pierre H.
Published: March 2006

The involvement of a catalytic metal vinylidene species was proposed for the first time in 1986 to explain the regioselective formation of vinyl carbamates directly from terminal alkynes, carbon dioxide, and amines. Since this initial report, various metal vinylidenes and allenylidenes, which are key activation intermediates, have proved extremely useful for many alkyne transformations. They have contributed to the rational design of new catalytic reactions. This 20th anniversary is a suitable occasion to present the advancement of organometallic vinylidenes and allenylidenes in catalysis.
Deiminated Epstein‐Barr virus nuclear antigen 1 is a target of anti–citrullinated protein antibodies in rheumatoid arthritis
Authors: Pratesi Federico , Tommasi Cristina , Anzilotti Consuelo , Chimenti Daniele , Migliorini Paola
Published: February 2006

Objective

To test the hypothesis that deimination of viral sequences containing Arg–Gly repeats could generate epitopes recognized by anti–citrullinated protein antibodies (ACPAs) that are present in rheumatoid arthritis (RA) sera.

Methods

Multiple antigen peptides derived from Epstein‐Barr virus (EBV)–encoded Epstein‐Barr nuclear antigen 1 (EBNA‐1) were synthesized, substituting the arginines with citrulline, and were used to screen RA sera. Anti–cyclic citrullinated peptide antibodies were purified by affinity chromatography and tested on a panel of in vitro deiminated proteins. Their ability to bind in vivo deiminated proteins was evaluated by immunoprecipitation, using EBV‐infected cell lines.

Results

Antibodies specific for a peptide corresponding to the EBNA‐135–58 sequence containing citrulline in place of arginine (viral citrullinated peptide [VCP]) were detected in 50% of RA sera and in <5% of normal and disease control sera. In addition, affinity‐purified anti‐VCP antibodies from RA sera reacted with filaggrin‐derived citrullinated peptides, with deiminated fibrinogen, and with deiminated recombinant EBNA‐1. Moreover, anti‐VCP antibodies immunoprecipitated, from the lysate of calcium ionophore–stimulated lymphoblastoid cell lines, an 80‐kd band that was reactive with a monoclonal anti–EBNA‐1 antibody and with anti–modified citrulline antibodies.

Conclusion

These data indicate that ACPAs react with a viral deiminated protein and suggest that EBV infection may play a role in the induction of these RA‐specific antibodies.
Hypermethylated promoter region of DR3, the death receptor 3 gene, in rheumatoid arthritis synovial cells
Authors: Takami Nozomi , Osawa Kayo , Miura Yasushi , Komai Koichiro , Taniguchi Mariko , Shiraishi Masahiko , Sato Keizo , Iguchi Tetsuhiro , Shiozawa Kazuko , Hashiramoto Akira , Shiozawa Shunichi
Published: February 2006

Objective

To examine the promoter activity and protein expression of the death receptor 3 gene DR3, a member of the apoptosis‐inducing Fas gene family, with particular reference to the methylation status of its promoter region in rheumatoid arthritis (RA).

Methods

Genomic DNA was prepared from peripheral blood mononuclear cells obtained from healthy individuals and from patients with RA and synovial cells obtained from patients with RA and osteoarthritis. The methylation status of the DR3 promoter was analyzed by bisulfite genomic sequencing and methylation‐specific polymerase chain reaction techniques. Gene promoter activity and protein expression were examined using the luciferase reporter and Western blotting techniques.

Results

The promoter region of the DR3 gene contained many CpG motifs, including one CpG island that was specifically hypermethylated in synovial cells from patients with RA. Promoter assays showed that the promoter CpG island was essential for the transactivation of the DR3 gene and that forced hypermethylation of the CpG island with the bacterial methylase Sss I in vitro resulted in inhibition of the DR3 gene expression. Furthermore, the expression of DR‐3 protein was down‐modulated in association with methylation of the promoter CpG island in RA synovial cells.

Conclusion

The CpG island in the DR3 gene promoter was specifically methylated to down‐modulate the expression of DR‐3 protein in rheumatoid synovial cells, which may provide resistance to apoptosis in RA synovial cells.
Comparative analysis of gene expression profiles in intact and damaged regions of human osteoarthritic cartilage
Authors: Sato Tomoo , Konomi Koji , Yamasaki Satoshi , Aratani Satoko , Tsuchimochi Kaneyuki , Yokouchi Masahiro , Masuko‐Hongo Kayo , Yagishita Naoko , Nakamura Hiroshi , Komiya Setsuro , Beppu Moroe , Aoki Haruhito , Nishioka Kusuki , Nakajima Toshihiro
Published: February 2006

Objective

To analyze the differences in gene expression profiles of chondrocytes in intact and damaged regions of cartilage from the same knee joint of patients with osteoarthritis (OA) of the knee.

Methods

We compared messenger RNA expression profiles in regions of intact and damaged cartilage (classified according to the Mankin scale) obtained from patients with knee OA. Five pairs of intact and damaged regions of OA cartilage were evaluated by oligonucleotide array analysis using a double in vitro transcription amplification technique. The microarray data were confirmed by real‐time quantitative polymerase chain reaction (PCR) amplification and were compared with previously published data.

Results

About 1,500 transcripts, which corresponded to 8% of the expressed transcripts, showed ≥2‐fold differences in expression between the cartilage tissue pairs. Approximately 10% of these transcripts (n = 151) were commonly expressed in the 5 patient samples. Accordingly, 114 genes (35 genes expressed in intact > damaged; 79 genes expressed in intact < damaged) were selected. The expression of some genes related to the wound‐healing process, including cell proliferation and interstitial collagen synthesis, was higher in damaged regions than in intact regions, similar to the findings for genes that inhibit matrix degradation. Comparisons of the real‐time quantitative PCR data with the previously reported data support the validity of our microarray data.

Conclusion

Differences between intact and damaged regions of OA cartilage exhibited a similar pattern among the 5 patients examined, indicating the presence of common mechanisms that contribute to cartilage destruction. Elucidation of this mechanism is important for the development of effective treatments for OA.
Catabolic stress induces features of chondrocyte senescence through overexpression of caveolin 1: Possible involvement of caveolin 1–induced down‐regulation of articular chondrocytes in the pathogenesis of osteoarthritis
Authors: Dai Sheng‐Ming , Shan Zheng‐Zheng , Nakamura Hiroshi , Masuko‐Hongo Kayo , Kato Tomohiro , Nishioka Kusuki , Yudoh Kazuo
Published: February 2006

Objective

Articular chondrocyte senescence is responsible, at least in part, for the increased incidence of osteoarthritis (OA) with increased age. Recently, it was suggested that caveolin 1, a 21–24‐kd membrane protein, participates in premature cellular senescence. Caveolin 1 is the principal structural component of caveolae, vesicular invaginations of the plasma membrane. This study was undertaken to investigate whether the catabolic factors oxidative stress and interleukin‐1β (IL‐1β) induce features of premature senescence of articular chondrocytes through up‐regulation of caveolin 1 expression.

Methods

Caveolin 1 expression was investigated in human OA cartilage by real‐time polymerase chain reaction and in rat OA cartilage by immunohistologic analysis. We studied whether IL‐1β and H2O2 induce caveolin 1 expression in OA chondrocytes and analyzed the relationship between cellular senescent phenotypes and caveolin 1 expression in human chondrocytes.

Results

In human and rat OA articular cartilage, caveolin 1 positivity was associated with cartilage degeneration. Both IL‐1β and H2O2 up‐regulated caveolin 1 messenger RNA and protein levels, and both treatments induced marked expression of senescent phenotypes: altered cellular morphology, cell growth arrest, telomere erosion, and specific senescence‐associated β‐galactosidase activity. Caveolin 1 overexpression induced p38 MAPK activation and impaired the ability of chondrocytes to produce type II collagen and aggrecan. In contrast, down‐regulation of caveolin 1 with antisense oligonucleotide significantly inhibited the features of chondrocyte senescence induced by catabolic factors. Caveolin 1 induction and stresses with both IL‐1β and H2O2 up‐regulated p53 and p21 and down‐regulated phosphorylated retinoblastoma (Rb), suggesting that the p53/p21/Rb phosphorylation pathway, as well as prolonged p38 MAPK activation, may mediate the features of chondrocyte senescence induced by stress.

Conclusion

Our findings suggest that IL‐1β and oxidative stress induce features of premature senescence in OA chondrocytes, mediated, at least in part, by stress‐induced caveolin 1 expression. This indicates that caveolin 1 plays a role in the pathogenesis of OA via promotion of chondrocyte down‐regulation.
Vasoactive intestinal peptide induces CD4+,CD25+ T regulatory cells with therapeutic effect in collagen‐induced arthritis
Authors: Gonzalez‐Rey Elena , Fernandez‐Martin Amelia , Chorny Alejo , Delgado Mario
Published: February 2006

Objective

CD4+,CD25+ T regulatory cells (Treg) control the immune response to a variety of antigens, including self antigens, and may offer opportunities to intervene in the course of autoimmune diseases. Several models support the idea of the peripheral generation of CD4+,CD25+ Treg from CD4+,CD25− T cells, but little is known about the endogenous factors and mechanisms controlling the peripheral expansion of CD4+,CD25+ Treg. We undertook this study to investigate the capacity of the vasoactive intestinal peptide (VIP), an immunosuppressive antiarthritic neuropeptide, to induce functional Treg in vivo during the development of collagen‐induced arthritis (CIA).

Methods

We measured the number of CD4+,CD25+ Treg following VIP administration to CIA mice, and we characterized their phenotype and their ability to suppress activation of autoreactive T cells. We determined the capacity of VIP to induce Treg in vitro as well as the use of Treg in the treatment of CIA, measuring the clinical evolution and the inflammatory and autoimmune components of the disease.

Results

The administration of VIP to arthritic mice resulted in the expansion of CD4+,CD25+,Foxp3+ Treg in the periphery and joints, which inhibited autoreactive T cell activation/expansion. VIP induced more efficient suppressors on a per‐cell basis. The VIP‐generated CD4+,CD25+ Treg transfer suppressed and significantly ameliorated the progression of the disease.

Conclusion

These results demonstrate the involvement of the generation of Treg in the therapeutic effect of VIP on CIA. The generation of highly efficient Treg by VIP ex vivo could be used as an attractive therapeutic tool in the future, avoiding the administration of the peptide to patients with rheumatoid arthritis.
Involvement of phosphatidylserine, αvβ3, CD14, CD36, and complement C1q in the phagocytosis of primary necrotic lymphocytes by macrophages
Authors: Böttcher Andrea , Gaipl Udo S. , Fürnrohr Barbara G. , Herrmann Martin , Girkontaite Irute , Kalden Joachim R. , Voll Reinhard E.
Published: February 2006

Objective

Uningested dead cells may be an important source of autoantigens and may trigger autoimmune diseases such as systemic lupus erythematosus (SLE). Multiple receptors involved in the clearance of apoptotic cells have been described; however, little is known about the receptors and ligands involved in uptake of necrotic cells that release autoantigens as well.

Methods

The uptake of autologous necrotic peripheral blood lymphocytes into human monocyte‐derived macrophages was qualitatively and quantitatively monitored by confocal microscopy and 2‐color flow cytometry, respectively. Blocking experiments were performed to examine the receptors and molecules involved in the phagocytosis of necrotic cells. Cytokine secretion by lipopolysaccharide‐activated monocytes and macrophages was determined by enzyme‐linked immunosorbent assay.

Results

Phosphatidylserine, which was exposed on necrotic as well as apoptotic cells, promoted the recognition and removal of primary necrotic lymphocytes. Several macrophage receptor systems, including the thrombospondin–CD36–αvβ3 complex, CD14, and the complement component C1q, contributed to the engulfment of necrotic cells. Necrotic peripheral blood lymphocytes slightly increased the lipopolysaccharide‐induced secretion of interleukin‐10 and reduced the secretion of tumor necrosis factor α in monocytes and macrophages.

Conclusion

Our results indicate that at least some of the receptors and adaptors mediating the uptake of apoptotic cells are also involved in the clearance of necrotic cells. Hence, necrotic cells engage phagocyte receptors such as CD36, which mediate antiinflammatory signals from apoptotic cells. Necrotic cells consequently also have the potency to provide antiinflammatory signals to phagocytes; however, these signals may be overridden by proinflammatory factors released during necrosis. These findings have implications regarding the etiopathogenesis of autoimmune diseases such as SLE, in which impaired clearance of dead cells may foster autoimmunity by the release of potential autoantigens.
Obstetric hospitalizations in the United States for women with systemic lupus erythematosus and rheumatoid arthritis
Authors: Chakravarty Eliza F. , Nelson Lorene , Krishnan Eswar
Published: February 2006

Objective

To estimate the national occurrence of pregnancies in women with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) and to compare pregnancy outcomes in these patients with those in women with pregestational diabetes mellitus (DM) and with the general obstetric population.

Methods

We studied the 2002 Nationwide Inpatient Sample of the Healthcare Cost and Utilization Project to estimate the number of obstetric hospitalizations, deliveries, and cesarean deliveries in women with SLE, RA, pregestational DM, and the general obstetric population. Pregnancy outcomes included length of hospital stay, hypertensive disorders including preeclampsia, premature rupture of membranes, and intrauterine growth restriction.

Results

Of an estimated 4.04 million deliveries, 3,264 occurred in women with SLE, 1,425 in women with RA, and 13,574 in women with pregestational DM. Women with SLE, RA, and pregestational DM had significantly increased rates of hypertensive disorders compared with the general obstetric population (23.2%, 11.1%, 27.4%, and 7.8%, respectively), longer hospital stays, and significantly higher risk of cesarean delivery. Although women with SLE, RA, and pregestational DM were significantly older than women in the general obstetric population, disparities in the risk of adverse outcomes of pregnancy remained statistically significant after adjustment for maternal age.

Conclusion

To our knowledge, this is the first study to examine national data on pregnancy outcomes in women with common rheumatic diseases. As with underlying pregestational DM, women with SLE and RA appear to have a higher age‐adjusted risk of adverse outcomes of pregnancy and longer hospital stays than do pregnant women in the general population, and careful antenatal monitoring should be performed.
The association of meniscal pathologic changes with cartilage loss in symptomatic knee osteoarthritis
Authors: Hunter D. J. , Zhang Y. Q. , Niu J. B. , Tu X. , Amin S. , Clancy M. , Guermazi A. , Grigorian M. , Gale D. , Felson D. T.
Published: February 2006

Objective

To explore the role of meniscal tears and meniscal malposition as risk factors for subsequent cartilage loss in subjects with symptomatic osteoarthritis (OA).

Methods

Study subjects were patients with symptomatic knee OA from the Boston Osteoarthritis of the Knee Study. Baseline assessments included knee magnetic resonance imaging (MRI) with followup MRI at 15 and 30 months. Cartilage and meniscal damage were scored on MRI in the medial and lateral tibiofemoral joints using the semiquantitative whole‐organ magnetic resonance imaging score. Tibiofemoral cartilage was scored on MR images of all 5 plates of each tibiofemoral joint, and the meniscal position was measured using eFilm Workstation software. A proportional odds logistic regression model with generalized estimating equations was used to assess the effect of each predictor (meniscal position factor and meniscal damage as dichotomous predictors in each model) on cartilage loss in each of the 5 plates within a compartment. Models were adjusted for age, body mass index (BMI), tibial width, and sex.

Results

We assessed 257 subjects whose mean ± SD age was 66.6 ± 9.2 years and BMI was 31.5 ± 5.7 kg/m2; 42% of subjects were female, and 77% of knees had a Kellgren/Lawrence radiographic severity grade ≥2. In the medial tibiofemoral joint, each measure of meniscal malposition was associated with an increased risk of cartilage loss. There was also a strong association between meniscal damage and cartilage loss. Since meniscal coverage and meniscal height diminished with subluxation, less coverage and reduced height also increased the risk of cartilage loss.

Conclusion

This study highlights the importance of an intact and functioning meniscus in patients with symptomatic knee OA, since the findings demonstrate that loss of this function has important consequences for cartilage loss.
2‐(N‐Fmoc)‐3‐(N‐Boc‐N‐methoxy)‐diaminopropanoic acid, an amino acid for the synthesis of mimics of O‐linked glycopeptides
Authors: Carrasco Michael R. , Brown Ryan T. , Doan Vu H. , Kandel Sean M. , Lee Franklin C.
Published: February 2006

Amino acids with N‐alkylaminooxy side chains have proven effective for the rapid synthesis of neoglycopeptides. Chemoselective reaction of reducing sugars with peptides containing these amino acids provides glycoconjugates that are structurally similar to their natural counterparts. 2‐(N‐Fmoc)‐3‐(N‐Boc‐N‐methoxy)‐diaminopropanoic acid (Fmoc: 9‐fluorenylmethoxycarbonyl; Boc: t‐butyloxycarbonyl) was synthesized from Boc–Ser–OH in >40% overall yield and incorporated into peptides by standard Fmoc chemistry based solid phase peptide synthesis. The resulting peptides are efficiently glycosylated and serve as mimics of O‐linked glycopeptides. The synthesis of this derivative greatly expands the availability of the N‐alkylaminooxy strategy for neoglycopeptides. © 2006 Wiley Periodicals, Inc. Biopolymers(Pept Sci) 84: 414–420, 2006

This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
Restrained molecular dynamics simulations of HIV‐1 protease: The first step in validating a new target for drug design
Authors: Perryman Alexander L. , Lin Jung‐Hsin , McCammon J. Andrew
Published: February 2006

To test the anticorrelated relationship that was recently displayed in conventional molecular dynamics (MD) simulations, several different restrained MD simulations on a wild type and on the V82F/I84V drug‐resistant mutant of HIV‐1 protease were performed. This anticorrelated relationship refers to the observation that compression of the peripheral ear‐to‐cheek region of HIV protease (i.e., the elbow of the flap to the fulcrum and the cantilever) occurred as the active site flaps were opening, and, conversely, expansion of that ear‐to‐cheek region occurred as both flaps were closing. An additional examination of this anticorrelated relationship was necessary to determine whether it can be harnessed in a useful manner. Consequently, six different MD experiments were performed that incorporated pairwise distance restraints in that ear‐to‐cheek region (i.e., the distance between the α‐carbons of Gly40 and Gln61 was restrained to either 7.7 or 10.5 Å, in both monomers). Pushing the backbones of the ear and the cheek regions away from each other slightly did force the flaps that guard the active site to remain closed in both the wild type and the mutant systems—even though there were no ligands in the active sites. Thus, these restrained MD simulations provided evidence that the anticorrelated relationship can be exploited to affect the dynamic behavior of the flaps that guard the active site of HIV‐1 protease. These simulations supported our hypothesis of the mechanism governing flap motion, and they are the first step towards validating that peripheral surface as a new target for drug design. © 2006 Wiley Periodicals, Inc. Biopolymers 82: 272–284, 2006

This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
Spatial patterns of protein expression in focal infections of human cytomegalovirus
Authors: Lam Vy , Boehme Karl W. , Compton Teresa , Yin John
Published: February 2006

Human cytomegalovirus (HCMV) is a medically significant human pathogen that infects a wide range of cell and tissue types. During infection, HCMV activates a variety of signal transduction pathways that induce profound changes in cellular processes and dramatically affect cellular gene expression patterns. To better define how these virus‐host interactions affect the local microenvironment and influence the spatial and temporal spread of HCMV, we initiated HCMV focal infections on normal human dermal fibroblast monolayers and monitored viral gene expression patterns and infection spread over 45 days. To establish baseline temporal measurements of HCMV infection and spread in cell monolayers, we characterized the influence of three experimental variables on viral gene expression: cell plating density, the presence of serum, and neutralization of cellular antiviral responses with an antibody against interferon‐β. We found that high cell plating density or the inclusion of serum correlated with enhanced HCMV infection spread. Dramatic differences in the expression pattern of the viral immediate early 2 (IE2) gene were observed under these conditions as compared to low plating density or the absence of serum. In the latter case round, uniform foci were observed with a clear wave of IE2 expression visible in advance of a late stage viral protein, envelope glycoprotein B. By contrast, larger irregular foci with arms of IE2 expression were observed in the presence of serum. Addition of the antibody had little effect on the rate of spread, which is consistent with the knowledge that HCMV represses antiviral responses during infection. This experimental system provides a useful means to visualize and quantify complex virus‐host interactions. © 2006 Wiley Periodicals, Inc.
Tyrosine‐based “Activatable Pro‐Tag”: Enzyme‐catalyzed protein capture and release
Authors: Lewandowski Angela T. , Small David A. , Chen Tianhong , Payne Gregory F. , Bentley William E.
Published: February 2006

Protein recovery is often achieved by a series of capture and release steps that often involve chromatographic binding and elution. We report an alternative, non‐chromatographic, capture and release approach that employs enzymes and the stimuli‐responsive polysaccharide chitosan. We capture our protein using the enzyme tyrosinase that oxidizes accessible tyrosine residues of the protein and “activates” these residues for covalent capture (i.e., conjugation) onto chitosan. Using fusions of green fluorescent protein (GFP) we observed that: (i) enzymatic activation is required for protein capture to chitosan; and (ii) capture is enhanced (approximately five‐fold) by engineering the protein to have a penta‐tyrosine fusion tag that provides additional accessible tyrosine residues for enzymatic activation. Because the fusion tag appears to be the primary site for capture, and capture requires activation, we designate penta‐tyrosine as a “pro‐tag.” The captured GFP‐chitosan conjugate possesses the pH‐responsive solubility that is characteristic of chitosan. We exploit this pH‐responsive solubility to facilitate purification of the captured protein. Two enzymatic methods were explored to release the captured GFP from the chitosan conjugate. The first method employs enterokinase (EK) to cleave the protein at an engineered EK‐cleavage site. The second method employs chitosanase to hydrolyze the chitosan backbone. Using GFP as a model protein, we demonstrated that enzymatic capture and release provides a simple, non‐chromatographic means to recover proteins directly from cell lysates. © 2006 Wiley Periodicals, Inc.
A population-based study of the incidence, management and prognosis of hepatic metastases from colorectal cancer
Authors: Leporrier J , Maurel J , Chiche L , Bara S , Segol P , Launoy G
Published: March 2006

Background

The aim of this population-based study was to evaluate the incidence, management and prognosis of patients with hepatic metastases related to colorectal cancer using data from the Digestive Cancer Registry of Calvados, France.

Methods

Of 1325 patients with colorectal cancer registered between January 1994 and December 1999, 358 developed hepatic metastases. Logistic regression was used to analyse prognostic factors. Survival analysis was carried out with Cox's proportional hazards model.

Results

Some 18·8 per cent of patients had synchronous metastases, while 29·3 per cent developed metastases at 3 years. Of patients with hepatic metastases, 17·3 per cent had a surgical resection, 40·2 per cent were treated with palliative chemotherapy and 42·5 per cent had symptomatic treatment. Factors associated with receiving symptomatic treatment only were age over 75 years and more than one metastasis, but not place of treatment. Median survival after a diagnosis of hepatic metastases was 10·7 (range 4·6–23·1) months. Significant adverse prognostic factors were: age over 75 years (P = 0·001), lymph node invasion of primary tumour (P = 0·024), bilateral distribution of metastases (P = 0·001), other metastases (P = 0·004) and symptomatic treatment only (P = 0·041).

Conclusion

Despite improvement in treatment for hepatic metastases, age and extent of disease remain limiting factors for surgical resection and palliative chemotherapy.
Prognosis of small thyroid cancer in patients with Graves' disease
Authors: Kikuchi S , Noguchi S , Yamashita H , Uchino S , Kawamoto H
Published: March 2006

Background

To find the best ways to follow up patients with small thyroid cancer (STC; 1 cm or less in diameter) and concomitant Graves' disease, this study examined whether such patients had the same excellent prognosis as those with STC without Graves' disease.

Methods

Between 1970 and 1996, 2199 patients were diagnosed as having STC by pathology after thyroidectomy. Of those, 509 patients (33 males and 476 females, mean age 43·5 years) underwent thyroidectomy for Graves' disease. Control patients with STC without Graves' disease were matched for age, sex, treatment year and tumour size (33 males and 476 females, mean age 44·0 years).

Results

Patients with STC and Graves' disease had a longer disease-free survival than those with STC alone (99 and 93 per cent at 20 years' follow-up, respectively; P < 0·001). The Cox's proportional hazard analysis showed that concomitant Graves' disease and age at surgery are more significant factors for predicting disease-free survival than surgical procedures.

Conclusion

Patients who undergo thyroidectomy for Graves' disease and are found to have STC have an excellent prognosis and longer disease-free survival than patients with STC alone.
Randomized clinical trial to determine the effect of nasogastric drainage on tracheal acid aspiration following oesophagectomy
Authors: Shackcloth M J , McCarron E , Kendall J , Russell G N , Pennefather S H , Tran J , Page R D
Published: March 2006

Background

The aim of this study was to investigate tracheal acid aspiration after oesophagectomy and to determine whether it is influenced by nasogastric (NG) drainage.

Methods

Thirty-four patients undergoing oesophagectomy were randomized to one of three methods of NG drainage: a single-lumen tube with free drainage and 4-hourly aspiration, a sump-type tube on continuous suction drainage, or no NG tube. A tracheal pH probe was used to collect information on acid aspiration for 48 h after surgery. A pH < 5·5 was considered abnormal (normal pH 6·8–7·2). Total time with tracheal pH < 5·5, number of reflux episodes and longest reflux time were compared between groups.

Results

There was significant and persistent tracheal acid aspiration in all patients. Patients with a sump-type tube had a significantly shorter total time with tracheal pH < 5·5 than those in the other groups (sump-type tube versus single-lumen tube, P = 0·0069; sump-type tube versus no tube, P = 0·0071). Patients randomized to no NG tube experienced more respiratory complications after surgery than those who had either single-lumen or sump-type tubes (seven of 12 versus four of 22 patients; P = 0·023). Insertion of a NG tube was necessary in the first week after surgery in seven of 12 patients in this group.

Conclusion

Routine NG drainage after oesophagectomy is necessary. A sump-type NG tube is better at preventing tracheal acid aspiration and may reduce the incidence of respiratory complications.
Selective cyclo-oxygenase 2 inhibition affects ileal but not colonic anastomotic healing in the early postoperative period
Authors: de Hingh I H J T , van Goor H , de Man B M , Lomme R M L M , Bleichrodt R P , Hendriks T
Published: March 2006

Background

Selective cyclo-oxygenase 2 (COX-2) inhibitors are increasingly prescribed in the perioperative period. Recent recognition of a possible role for COX-2 in wound healing has raised concerns about the safety of their use in surgical practice. Therefore, the influence of celecoxib, a selective COX-2 inhibitor, on early anastomotic healing was investigated.

Methods

Celecoxib, in doses of 15, 50 or 200 mg per kg per day, was given daily from the day before operation onwards to male Wistar rats that received both ileal and colonic anastomoses. Anastomotic strength was assessed by measuring the breaking strength and bursting pressure on the third day after operation. A second group received a dose of 50 mg per kg per day and a colonic anastomosis only, and healing was assessed on the third and fifth day after surgery.

Results

Expression of COX-2 protein was upregulated in the anastomotic area. Administration of celecoxib, at all doses tested, resulted in a significantly higher ileal dehiscence rate than in control rats (P = 0·002). In contrast, colonic anastomoses healed normally within the same animals. The latter was confirmed in rats with colonic anastomoses only.

Conclusion

In this model, administration of the COX-2 inhibitor celecoxib affected ileal but not colonic anastomotic healing in the early postoperative period.
Decontamination of organochlorine pesticides in Radix Codonopsis by supercritical fluid extractions and determination by gas chromatography
Authors: Zhao Chunjie , Hao Guiming , Li Huanxin , Luo Xu , Chen Yingjie
Published: February 2006

A method involving depuration of 12 organochlorine pesticides (OCPs) from Radix codonopsis was developed using supercritical fluid extraction (SFE). The pesticides investigated in the study included α‐, β‐, γ‐ and δ‐benzene hexachloride, PCNB (pentachloro‐nitrobenzene), PCA (pentachloroaniline), HEPT (heptachlor), MPCPS (methyl‐pentachlorophenyl sulfide), pp′‐DDE [1,1‐dichloro‐2, 2‐bis(p‐chlorophenyl) ethylene], op′‐DDT [1,1,1‐trichloro‐2‐(o‐chlorophenyl)‐2‐(p‐chlorophenyl)ethane], pp′‐DDD [1,1‐dichloro‐2‐2‐bis(p‐chlorophenyl)ethane] and pp′‐DDT [1,1,1‐trichloro‐2,2‐bis(p‐chlorophenyl)ethane]. A series of experiments was conducted to optimize the final extraction conditions as follows: pure CO2, extraction pressure 15 MPa, extraction temperature 60°C, extraction time 20 min and flow rate 55 mL/h. A GC method with electron capture detection was employed for the determination of the OCPs in Radix codonopsis. An HPLC method was developed for the quantitative determination of active constituents. SFE was used to remove the organochlorine pesticide from Radix codonopsis. The results showed that at least 93.5% of the organochlorine pesticide residues in the herb sample were removed by SPE, while 95.0% of the active constituent marker (atractylenoide III) remained. Copyright © 2006 John Wiley & Sons, Ltd.
Determination of erythromycin A in rat plasma and urine by high‐performance liquid chromatography with chemiluminescence detection using Tris(2,2′‐bipyridine)ruthenium(II)
Authors: Hori Toshihiko , Hashimoto Hiroshi , Konishi Masaharu
Published: February 2006

A simple and sensitive method was developed for the determination of erythromycin A (EA), decladinosyl erythromycin A (dClEA) and erythromycin B (EB) in rat plasma and urine by high‐performance liquid chromatography with electrogenerated chemiluminescence detection using Tris(2,2′‐bipyridine)ruthenium(II). The recovery rates of EA, dClEA and EB were 97, 94 and 85% from rat plasma and 89, 83 and 93% from rat urine, respectively. The calibration curves were linear over the concentration ranges 0.05–5 µg/mL for plasma and 0.5–50 µg/mL for urine. The precision and accuracy for all analytes in rat plasma were ≤9.0 and −6.3–7.2%, and those in urine were ≤9.4% and −6.1–7.6%, respectively. This method proved to be a powerful tool for determination of EA, dClEA and EB concentrations in samples from rats. Copyright © 2006 John Wiley & Sons, Ltd.
A structure–activity relationship study of compounds with antihistamine activity
Authors: Brzezińska Elżbieta , Kośka Grażyna
Published: February 2006

A structure–activity relationship (SAR) analysis of H1‐, H2‐ and H3‐antihistamine activity was carried out and chromatographic data of 2‐[2‐(phenylamino)thiazol‐4‐yl]ethanamine, 2‐(2‐benzyl‐4‐thiazolyl)ethanamine, 2‐(2‐benzhydrylthiazol4‐yl)ethanamine, 2‐(1‐piperazinyl‐ and 2‐(hexahydro‐1H‐1,4‐diazepin‐1‐yl)benzothiazole, 2‐(1‐piperazinyl)benzothiazole, 2‐[4‐(1‐alkyl)piperidinyl]benzothiazole, 2‐(N,N′,N′‐dimethylalkyl‐1,2‐ethanediamino)benzothiazole, 2[1‐(4‐aminopiperidinyl)]benzothiazole, 2‐[2‐phenyl‐4‐thiazolyl]ethanamine derivatives and selected H1‐ and H2‐antihistamine drugs were obtained. NP TLC and RP2 TLC plates (silica gel NP 60F254 and silica gel RP2 60F254 silanized precoated), impregnated with a solution of aspartic acid (l‐Asp) and a solution of an analogue of aspartic acid (propionic acid), were used in two developing solvents as H1‐, H2‐ and H3‐antihistaminic interaction models. The lipophilicity data of the examined compounds were obtained and used in the SAR assay. Biochromatographic tests using TLC plates impregnated with solutions of asparic acid or propionic acid were found to be a source of useful data for the qualitative analysis of compounds with different structures, demonstrating activity to histamine H1‐, H2‐ and H3‐receptors. The four presented discriminant models based on biochromatographic studies are an efficient tool in the SAR analysis for initial prediction of compound activity direction within histamine receptors. Copyright © 2006 John Wiley & Sons, Ltd.
Pharmacokinetic study of p‐coumaric acid in mouse after oral administration of extract of Ananas comosus L. leaves
Authors: Meng Zhen , Wang Wei , Xing Dong‐Ming , Lei Fan , Lan Jia‐Qi , Du Li‐Jun
Published: February 2006

Quantification of p‐coumaric acid in mouse plasma following oral administration of Ananas comosus L. leaves was achieved by reversed‐phase high‐performance liquid chromatography using a mobile phase of water–acetonitrile (82:18, v/v) and UV detection at 310 nm. The method was linear (determination coefficient, r2 = 0.9997) within the tested range (0.04–1.28 µg/mL). Intra‐ and inter‐day precision coefficients of variation and accuracy bias were acceptable (maximal CV value was 4.06% for intra‐day and 4.19% for inter‐day) over the entire range. The recoveries were 90.63, 97.98 and 100.01% for concentrations of 0.04, 0.32 and 1.28 µg/mL, respectively. This is a very rapid, sensitive and economical way to determine p‐coumaric acid concentration in mouse plasma after oral administration of A. comosus leaves. The concentration–time curve was fitted to the one‐compartment model. This is the first time that p‐coumaric acid extracted from A. comosus leaves was detected by HPLC‐UV method and its pharmacokinetic characteristic was comprehensively studied. Copyright © 2006 John Wiley & Sons, Ltd.
Sensitive determination of 4‐(4‐chlorophenyl)‐4‐hydroxypiperidine, a metabolite of haloperidol, in a rat biological sample by HPLC with fluorescence detection after pre‐column derivatization using 4‐fluoro‐7‐nitro‐2,1,3‐benzoxadiazole
Authors: Higashi Yasuhiko , Nakamura Shota , Fujii Youichi
Published: February 2006

4‐(4‐Chlorophenyl)‐4‐hydroxypiperidine (CPHP), one of the metabolites of haloperidol, is considered to exhibit brain toxicity. CPHP concentrations in plasma and tissue homogenates (each 200 µL) from rats were analyzed by HPLC fluorescence detection after pre‐column derivatization with 4‐fluoro‐7‐nitro‐2,1,3‐benzoxadiazole (NBD‐F). After basic extraction of the samples with benzene, the derivatization with NBD‐F was conducted in borate buffer (pH 8.0) at 60°C for 3 min. Mexiletine was carried through the procedure as an internal standard. The regression equation for CPHP showed a good linearity in the range of 0.03–1 µg/mL with a detection limit of 0.008 µg/mL. The coefficient of variation was less than 11.6%. Plasma concentration–time courses of CPHP after intraperitoneal or per oral administration of CPHP, haloperidol or reduced haloperidol were examined, and the pharmacokinetic parameters were estimated. Additionally, CPHP levels in various tissues at 8 h after intraperitoneal administration of these compounds were compared. The method was simple and sensitive, useful for determination of CPHP in rat biological samples using as little as 200 µL of sample volume and could be applied for pharmacokinetic study. Copyright © 2006 John Wiley & Sons, Ltd.
Polyurethane foam chips combined with liquid chromatography in the determination of unmetabolized polycyclic aromatic hydrocarbons excreted in human urine
Authors: Buratti Marina , Pellegrino Oronzo , Valla Carla , Rubino Federico Maria , Verduci Cinzia , Colombi Antonio
Published: February 2006

A method suitable for the determination of unmetabolized polycyclic aromatic hydrocarbons (PAHs) excreted at trace levels (ng/L) in human urine for the monitoring of exposure of the general population to PAH contamination was developed. PAHs were determined, after enrichment by solid‐phase extraction on polyurethane foam (PUF) chips, by HPLC with fluorescence detection. Different parameters affecting analyte extraction to the PUF, including urine salting‐out and organic additives, and optimization of conditions for clean‐up and desorption have been investigated. Optimized conditions were 40 mL acidified urine sample, added with magnesium sulfate, tetrahydrofuran and a 2 cm3 PUF chip, and extracted by shaking at 30 rpm for 1 h at ambient temperature. Desorption was performed, after a clean‐up step with diluted sodium hydroxide, using a small amount of diethyl ether. The recovery of PAH congeners from spiked urines was >90% in the 2–100 ng/L range; the detection limit was 0.1–0.5 ng/L, depending on the considered PAH congener; day‐to‐day precision, at 50 ng/L native PAH content, was CV = 10–20%. The proposed technique provides a simple, economical and effective procedure for the determination of trace amounts of unmetabolized PAHs excreted in human urine spot samples. Copyright © 2006 John Wiley & Sons, Ltd.