bluuebunny/crossref_metadata_2025_split huggingface dataset:
Multigene expression of protein complexes by iterative modification of genomic Bacmid DNA
Authors: Noad Rob J , Stewart Meredith , Boyce Mark , Celma Cristina C , Willison Keith R , Roy Polly 
Published: September 2009

Abstract

Background
Many cellular multi-protein complexes are naturally present in cells at low abundance. Baculovirus expression offers one approach to produce milligram quantities of correctly folded and processed eukaryotic protein complexes. However, current strategies suffer from the need to produce large transfer vectors, and the use of repeated promoter sequences in baculovirus, which itself produces proteins that promote homologous recombination. One possible solution to these problems is to construct baculovirus genomes that express each protein in a complex from a separate locus within the viral DNA. However current methods for selecting such recombinant genomes are too inefficient to routinely modify the virus in this way.

Results
This paper reports a method which combines the lambda red and bacteriophage P1 Cre-recombinase systems to efficiently generate baculoviruses in which protein complexes are expressed from multiple, single-locus insertions of foreign genes. This method is based on an 88 fold improvement in the selection of recombinant viruses generated by red recombination techniques through use of a bipartite selection cassette. Using this system, seven new genetic loci were identified in the AcMNPV genome suitable for the high level expression of recombinant proteins. These loci were used to allow the recovery two recombinant virus-like particles with potential biotechnological applications (influenza A virus HA/M1 particles and bluetongue virus VP2/VP3/VP5/VP7 particles) and the mammalian chaperone and cancer drug target CCT (16 subunits formed from 8 proteins).

Conclusion

1. Use of bipartite selections can significantly improve selection of modified bacterial artificial chromosomes carrying baculovirus DNA. Furthermore this approach is sufficiently robust to allow routine modification of the virus genome. 2. In addition to the commonly used p10 and polyhedrin loci, the ctx, egt, 39k, orf51, gp37, iap2 and odv-e56 loci in AcMNPV are all suitable for the high level expression of heterologous genes. 3. Two protein, four protein and eight protein complexes including virus-like particles and cellular chaperone complexes can be produced using the new approach.
Protective effect of transgenic expression of porcine heat shock protein 70 on hypothalamic ischemic and oxidative damage in a mouse model of heatstroke
Authors: Chen Zhih-Cherng , Wu Wen-Shian , Lin Mao-Tsun , Hsu Chuan-Chih 
Published: September 2009

Abstract

Background
Transgenic mice have been used to examine the role of heat shock protein (HSP)72 in experimental heatstroke. Transgenic mice that were heterozygous for a porcine HSP70β gene ([+] HSP72) and transgene-negative littermate controls ([-] HSP72), under pentobarbital sodium anesthesia, were subjected to heat stress to induce heatstroke. It was found that the overexpression of HSP72 in multiple organs improved survival during heatstroke by reducing hypotension and cerebral ischemia and damage in mice. Herein we attempted to further assess the effect of heat exposure on thermoregulatory function, hypothalamic integration, and survival in unrestrained, unanesthetized [+]HSP72 and compare with those of [-]HSP72. In this research with the transgenic mice, we first conducted several biochemical, physiologic and histological determinations and then investigated the beneficial effects of HSP72 overexpression on the identified hypothalamic deficits, thermoregulatory dysfunction, and mortality during heatstroke.

Results
We report here that when [-]HSP72 mice underwent heat stress (ambient temperature 42.4°C for 1 h), the fraction survival and core temperature at 4 h after heat stress were found to be 0 of 12 and 34.2°C ± 0.4°C, respectively. Mice that survived to day 4 after heat stress were considered as survivors. In [+]HSP72 mice, when exposed to the same heat treatment, both fraction survival and core temperature values were significantly increased to new values of 12/12 and 37.4°C ± 0.3°C, respectively. Compared to [-]HSP mice, [+]HSP72 mice displayed lower hypothalamic values of cellular ischemia (e.g., glutamate and lactate-to-pyruvate ratio) and damage (e.g., glycerol) markers, pro-oxidant enzymes (e.g., lipid peroxidation and glutathione oxidation), pro-inflammatory cytokines (e.g., interleukin-1beta and tumor necrosis factor-alpha), and neuronal damage score evaluated 4 h after heat stress. In contrast, [+]HSP72 mice had higher hypothalamic values of antioxidant defences (e.g., glutathione peroxidase and glutathione reductase), ATP, and HSP72 expression.

Conclusion
This study indicates that HSP72 overexpression appears to be critical to the development of thermotolerance and protection from heat-induced hypothalamic ischemic and oxidative damage.
Otitis media: a genome-wide linkage scan with evidence of susceptibility loci within the 17q12 and 10q22.3 regions
Authors: Casselbrant Margaretha L , Mandel Ellen M , Jung Jeesun , Ferrell Robert E , Tekely Kathleen , Szatkiewicz Jin P , Ray Amrita , Weeks Daniel E 
Published: September 2009

Abstract

Background
Otitis media (OM) is a common worldwide pediatric health care problem that is known to be influenced by genetics. The objective of our study was to use linkage analysis to map possible OM susceptibility genes.

Methods
Using a stringent diagnostic model in which only those who underwent tympanostomy tube insertion at least once for recurrent/persistent OM are considered affected, we have carried out a genome-wide linkage scan using the 10K Affymetrix SNP panel. We genotyped 403 Caucasian families containing 1,431 genotyped individuals and 377 genotyped affected sib pairs, and 26 African American families containing 75 genotyped individuals and 27 genotyped affected sib pairs. After careful quality control, non-parametric linkage analysis was carried out using 8,802 SNPs.

Results
In the Caucasian-only data set, our most significant linkage peak is on chromosome 17q12 at rs226088 with a p-value of 0.00007. Other peaks of potential interest are on 10q22.3 (0.00181 at rs1878001), 7q33 (0.00105 at rs958408), 6p25.1 (0.00261 at rs554653), and 4p15.2 (0.00301 at rs2133507). In the combined Caucasian and African American dataset, the 10q22.3 peak becomes more significant, with a minimal p-value of 0.00026 at rs719871. Family-based association testing reveals signals near previously implicated genes: 513 kb from SFTPA2 (10q22.3), 48 kb from IFNG (12q14), and 870 kb from TNF (6p21.3).

Conclusion
Our scan does not provide evidence for linkage in the previously reported regions of 10q26.3 and 19q13.43. Our best-supported linkage regions may contain susceptibility genes that influence the risk for recurrent/persistent OM. Plausible candidates in 17q12 include AP2B1, CCL5, and a cluster of other CCL genes, and in 10q22.3, SFTPA2.
Biomarkers of apoptosis and survival in esophageal squamous cell carcinoma
Authors: Takikita Mikiko , Hu Nan , Shou Jian-zhong , Wang Quan-Hong , Giffen Carol , Taylor Philip R , Hewitt Stephen M 
Published: September 2009

Abstract

Background
Cancer of the esophagus is a deadly malignancy, and development of biomarkers that predict survival is an urgent need. The apoptotic pathways have been hypothesized as important in progression of esophageal squamous cell carcinoma (ESCC). We investigated a panel of proteins that regulate apoptosis as candidate of biomarkers of prognosis in ESCC.

Methods
Tissue microarray (TMA) including 313 surgically-resected cases of ESCC specimens was built for immunohistochemical interrogation. We evaluated seven genes in the FasL-Fas apoptotic pathway - FasL, Fas, FAS-associated death domain protein (FADD), phosphorylated-FADD, and caspase 8 and 10, and the antiapoptotic protein bcl-2. We studied pathway integrity and relations to risk and clinical factors, and determined the prognostic significance of each marker.

Results
Five markers showed strong inter-marker correlations (r ≥ 0.28, p < 0.001), including FasL, Fas, FADD, and caspases 8 and 10. FasL and FADD also showed modest correlations with one or more cancer risk factors, but none of the markers was significantly associated with either tumor stage or lymph node metastasis, the only two clinical factors that predicted survival in these ESCC cases. Multivariate-adjusted proportional hazard regression models showed no association between protein expression and risk of death for any of the seven markers examined.

Conclusion
Individual biomarkers in the apoptosis pathway do not appear to predict survival of patients with ESCC.
Adaptation and evaluation of the bottle assay for monitoring insecticide resistance in disease vector mosquitoes in the Peruvian Amazon
Authors: Zamora Perea Elvira , Balta León Rosario , Palomino Salcedo Miriam , Brogdon William G , Devine Gregor J 
Published: September 2009

Abstract

Background
The purpose of this study was to establish whether the "bottle assay", a tool for monitoring insecticide resistance in mosquitoes, can complement and augment the capabilities of the established WHO assay, particularly in resource-poor, logistically challenging environments.

Methods
Laboratory reared Aedes aegypti and field collected Anopheles darlingi and Anopheles albimanus were used to assess the suitability of locally sourced solvents and formulated insecticides for use with the bottle assay. Using these adapted protocols, the ability of the bottle assay and the WHO assay to discriminate between deltamethrin-resistant Anopheles albimanus populations was compared. The diagnostic dose of deltamethrin that would identify resistance in currently susceptible populations of An. darlingi and Ae. aegypti was defined. The robustness of the bottle assay during a surveillance exercise in the Amazon was assessed.

Results
The bottle assay (using technical or formulated material) and the WHO assay were equally able to differentiate deltamethrin-resistant and susceptible An. albimanus populations. A diagnostic dose of 10 μg a.i./bottle was identified as the most sensitive discriminating dose for characterizing resistance in An. darlingi and Ae. aegypti. Treated bottles, prepared using locally sourced solvents and insecticide formulations, can be stored for > 14 days and used three times. Bottles can be stored and transported under local conditions and field-assays can be completed in a single evening.

Conclusion
The flexible and portable nature of the bottle assay and the ready availability of its components make it a potentially robust and useful tool for monitoring insecticide resistance and efficacy in remote areas that require minimal cost tools.
Modelling the progression of pandemic influenza A (H1N1) in Vietnam and the opportunities for reassortment with other influenza viruses
Authors: Boni Maciej F , Manh Bui Huu , Thai Pham Quang , Farrar Jeremy , Hien Tran Tinh , Hien Nguyen Tran , Van Kinh Nguyen , Horby Peter 
Published: September 2009

Abstract

Background
A novel variant of influenza A (H1N1) is causing a pandemic and, although the illness is usually mild, there are concerns that its virulence could change through reassortment with other influenza viruses. This is of greater concern in parts of Southeast Asia, where the population density is high, influenza is less seasonal, human-animal contact is common and avian influenza is still endemic.

Methods
We developed an age- and spatially-structured mathematical model in order to estimate the potential impact of pandemic H1N1 in Vietnam and the opportunities for reassortment with animal influenza viruses. The model tracks human infection among domestic animal owners and non-owners and also estimates the numbers of animals may be exposed to infected humans.

Results
In the absence of effective interventions, the model predicts that the introduction of pandemic H1N1 will result in an epidemic that spreads to half of Vietnam's provinces within 57 days (interquartile range (IQR): 45-86.5) and peaks 81 days after introduction (IQR: 62.5-121 days). For the current published range of the 2009 H1N1 influenza's basic reproductive number (1.2-3.1), we estimate a median of 410,000 cases among swine owners (IQR: 220,000-670,000) with 460,000 exposed swine (IQR: 260,000-740,000), 350,000 cases among chicken owners (IQR: 170,000-630,000) with 3.7 million exposed chickens (IQR: 1.9 M-6.4 M), and 51,000 cases among duck owners (IQR: 24,000 - 96,000), with 1.2 million exposed ducks (IQR: 0.6 M-2.1 M). The median number of overall human infections in Vietnam for this range of the basic reproductive number is 6.4 million (IQR: 4.4 M-8.0 M).

Conclusion
It is likely that, in the absence of effective interventions, the introduction of a novel H1N1 into a densely populated country such as Vietnam will result in a widespread epidemic. A large epidemic in a country with intense human-animal interaction and continued co-circulation of other seasonal and avian viruses would provide substantial opportunities for H1N1 to acquire new genes.
Identification of a high incidence region for retroviral vector integration near exon 1 of the LMO2locus
Authors: Yamada Koichiro , Tsukahara Tomonori , Yoshino Kazuhisa , Kojima Katsuhiko , Agawa Hideyuki , Yamashita Yuki , Amano Yuji , Hatta Mariko , Matsuzaki Yasunori , Kurotori Naoki , Wakui Keiko , Fukushima Yoshimitsu , Osada Ryosuke , Shiozawa Tanri , Sakashita Kazuo , Koike Kenichi , Kumaki Satoru , Tanaka Nobuyuki , Takeshita Toshikazu 
Published: September 2009

Abstract
Therapeutic retroviral vector integration near the oncogene LMO2 is thought to be a cause of leukemia in X-SCID gene therapy trials. However, no published studies have evaluated the frequency of vector integrations near exon 1 of the LMO2 locus. We identified a high incidence region (HIR) of vector integration using PCR techniques in the upstream region close to the LMO2 transcription start site in the TPA-Mat T cell line. The integration frequency of the HIR was one per 4.46 × 104 cells. This HIR was also found in Jurkat T cells but was absent from HeLa cells. Furthermore, using human cord blood-derived CD34+ cells we identified a HIR in a similar region as the TPA-Mat T cell line. One of the X-linked severe combined immunodeficiency (X-SCID) patients that developed leukemia after gene therapy had a vector integration site in this HIR. Therefore, the descriptions of the location and the integration frequency of the HIR presented here may help us to better understand vector-induced leukemogenesis.
Chondrogenic differentiation potential of osteoarthritic chondrocytes and their possible use in matrix-associated autologous chondrocyte transplantation
Authors: Dehne Tilo , Karlsson Camilla , Ringe Jochen , Sittinger Michael , Lindahl Anders 
Published: September 2009

Abstract
Introduction
Autologous chondrocyte transplantation (ACT) is a routine technique to regenerate focal cartilage lesions. However, patients with osteoarthritis (OA) are lacking an appropriate long-lasting treatment alternative, partly since it is not known if chondrocytes from OA patients have the same chondrogenic differentiation potential as chondrocytes from donors not affected by OA.

Methods
Articular chondrocytes from patients with OA undergoing total knee replacement (Mankin Score > 3, Ahlbäck Score > 2) and from patients undergoing ACT, here referred to as normal donors (ND), were isolated applying protocols used for ACT. Their chondrogenic differentiation potential was evaluated both in high-density pellet and scaffold (Hyaff-11) cultures by histological proteoglycan assessment (Bern Score) and immunohistochemistry for collagen types I and II. Chondrocytes cultured in monolayer and scaffolds were subjected to gene expression profiling using genome-wide oligonucleotide microarrays. Expression data were verified by using real-time PCR.

Results
Chondrocytes from ND and OA donors demonstrated accumulation of comparable amounts of cartilage matrix components, including sulphated proteoglycans and collagen types I and II. The mRNA expression of cartilage markers (ACAN, COL2A1, COMP, CRTL1, SOX9) and genes involved in matrix synthesis (BGN, CILP2, COL9A2, COL11A1, TIMP4) was highly induced in 3D cultures of chondrocytes from both donor groups. Genes associated with hypertrophic or OA cartilage (ALPL, COL1A1, COL3A1, COL10A1, MMP13, POSTN, PTH1R, RUNX2) were not significantly regulated between the two groups of donors. The expression of 661 genes, including COMP, FN1, and SOX9, was differentially regulated between OA and ND chondrocytes cultured in monolayer. During scaffold culture, the differences diminished between the OA and ND chondrocytes, and only 184 genes were differentially regulated.

Conclusions
Only few genes were differentially expressed between OA and ND chondrocytes in Hyaff-11 culture. The risk of differentiation into hypertrophic cartilage does not seem to be increased for OA chondrocytes. Our findings suggest that the chondrogenic capacity is not significantly affected by OA, and OA chondrocytes fulfill the requirements for matrix-associated ACT.
Mechanical effects of surgical procedures on osteochondral grafts elucidated by osmotic loading and real-time ultrasound
Authors: Hattori Koji , Uematsu Kota , Matsumoto Tomohiro , Ohgushi Hajime 
Published: September 2009

Abstract
Introduction
Osteochondral grafts have become popular for treating small, isolated and full-thickness cartilage lesions. It is recommended that a slightly oversized, rather than an exact-sized, osteochondral plug is transplanted to achieve a tight fit. Consequently, impacting forces are required to insert the osteochondral plug into the recipient site. However, it remains controversial whether these impacting forces affect the biomechanical condition of the grafted articular cartilage. The present study aimed to investigate the mechanical effects of osteochondral plug implantation using osmotic loading and real-time ultrasound.

Methods
A full-thickness cylindrical osteochondral defect (diameter, 3.5 mm; depth, 5 mm) was created in the lateral lower quarter of the patella. Using graft-harvesting instruments, an osteochondral plug (diameter, 3.5 mm as exact-size or 4.5 mm as oversize; depth, 5 mm) was harvested from the lateral upper quarter of the patella and transplanted into the defect. Intact patella was used as a control. The samples were monitored by real-time ultrasound during sequential changes of the bathing solution from 0.15 M to 2 M saline (shrinkage phase) and back to 0.15 M saline (swelling phase). For cartilage sample assessment, three indices were selected, namely the change in amplitude from the cartilage surface (amplitude recovery rate: ARR) and the maximum echo shifts from the cartilage surface and the cartilage-bone interface.

Results
The ARR is closely related to the cartilage surface integrity, while the echo shifts from the cartilage surface and the cartilage-bone interface are closely related to tissue deformation and NaCl diffusion, respectively. The ARR values of the oversized plugs were significantly lower than those of the control and exact-sized plugs. Regarding the maximum echo shifts from the cartilage surface and the cartilage-bone interface, no significant differences were observed among the three groups.

Conclusions
These findings demonstrated that osmotic loading and real-time ultrasound were able to assess the mechanical condition of cartilage plugs after osteochondral grafting. In particular, the ARR was able to detect damage to the superficial collagen network in a non-destructive manner. Therefore, osmotic loading and real-time ultrasound are promising as minimally invasive methods for evaluating cartilage damage in the superficial zone after trauma or impact loading for osteochondral grafting.
Brachial artery peak velocity variation to predict fluid responsiveness in mechanically ventilated patients
Authors: Monge García Manuel Ignacio , Gil Cano Anselmo , Díaz Monrové Juan Carlos 
Published: September 2009

Abstract
Introduction
Although several parameters have been proposed to predict the hemodynamic response to fluid expansion in critically ill patients, most of them are invasive or require the use of special monitoring devices. The aim of this study is to determine whether noninvasive evaluation of respiratory variation of brachial artery peak velocity flow measured using Doppler ultrasound could predict fluid responsiveness in mechanically ventilated patients.

Methods
We conducted a prospective clinical research in a 17-bed multidisciplinary ICU and included 38 mechanically ventilated patients for whom fluid administration was planned due to the presence of acute circulatory failure. Volume expansion (VE) was performed with 500 mL of a synthetic colloid. Patients were classified as responders if stroke volume index (SVi) increased ≥ 15% after VE. The respiratory variation in Vpeakbrach (ΔVpeakbrach) was calculated as the difference between maximum and minimum values of Vpeakbrach over a single respiratory cycle, divided by the mean of the two values and expressed as a percentage. Radial arterial pressure variation (ΔPPrad) and stroke volume variation measured using the FloTrac/Vigileo system (ΔSVVigileo), were also calculated.

Results
VE increased SVi by ≥ 15% in 19 patients (responders). At baseline, ΔVpeakbrach, ΔPPrad and ΔSVVigileo were significantly higher in responder than nonresponder patients [14 vs 8%; 18 vs. 5%; 13 vs 8%; P < 0.0001, respectively). A ΔVpeakbrach value >10% predicted fluid responsiveness with a sensitivity of 74% and a specificity of 95%. A ΔPPrad value >10% and a ΔSVVigileo >11% predicted volume responsiveness with a sensitivity of 95% and 79%, and a specificity of 95% and 89%, respectively.

Conclusions
Respiratory variations in brachial artery peak velocity could be a feasible tool for the noninvasive assessment of fluid responsiveness in patients with mechanical ventilatory support and acute circulatory failure.

Trial Registration
ClinicalTrials.gov ID: NCT00890071