bluuebunny/crossref_metadata_2025_split huggingface dataset:
Synthesis and purification of RNA substrates
Authors: Chabot Benoit 
Published: November 2023

Abstract
In recent years, the purification of RNA molecules from different organisms and cell types has been crucial to the analysis of gene expression and RNA processing events. Likewise, the rapid development of in vitro RNA maturation systems has relied extensively on the ability to synthesize and purify RNA substrates. Generating pure, undegraded RNA is thus critical to in vivo and in vitro analyses of RNA processing. For the most part, procedures used to obtain intact and pure RNA are relatively simple and have been described in detail in a variety of publications and technical books. This chapter does not attempt to provide a comprehensive coverage of all these procedures. Instead, it describes a few of the most rapid, convenient, and economical ways to prepare RNA for direct analysis or for use as substrates in in vitro systems of RNA processing. Researchers who work with RNA are always very much concerned about RNase contamination and thus take great pains to establish an RNase-free microenvironment. However, in most cases problems with RNase contamination can be avoided by taking the following simple precautions:
Overview of the steroid receptor superfamily of gene regulatory proteins
Authors: O’Malley Bert W , Tsai Ming-Jer 
Published: November 2023

Abstract
Over the past two decades, a great deal of evidence has accumulated in favour of the hypothesis that steroid hormones act at the level of nuclear DNA to regulate gene expression (1-4). The earliest studies were qualitative and involved experiments which showed that steroid hormones caused: (1) accumulation of new species of hybridizable RNAs that did not exist before stimulation; (2) stimulation of synthesis of new specific proteins; (3) a corresponding increase in the cellular levels of specific messenger RNAs (mRNAs); and (4) stimulation of the rate of transcription of select endogenous nuclear genes (5). At this point, the early 1970s, the primary pathway for steroid hormone action was defined as follows: steroid→ (steroid-receptor) → (steroid-receptor-DNA) → mRNA → protein → functional response (6). Steroid was thought to enter cells by passive diffusion and to activate receptors in either the cytoplasm or nucleus. The activated receptor was predicted to bind at the regulatory region of target genes and to stimulate transcription, and subsequently protein synthesis. Although initially controversial, this pathway has been generally accepted for the past 15 years (5-13).
In Vitro Sv40 Dna Replication
Authors: Kenny Mark K 
Published: November 2023

Abstract
Simian virus 40 (SV40) has served as an excellent model for various cellular processes including transformation, transcription, RNA processing, and DNA replication (1). Replication of the genome of this small DNA tumour virus very closely resembles the replication of cellular DNA. SV40 has a 5 kb circular duplex genome which exists in a chromatin structure indistinguishable from that of the cell. Replication initiates at a single 64 bp core origin of replication and proceeds bidirectionally in a semi-conservative manner. SV40 uses only a single viral protein for its replication, the large T antigen, albeit it serves several functions. All of the other replication proteins are provided by the host cell. There is a species specificity to SV40 DNA replication. It will replicate in primate cells but not in rodent cells.
Purification and assay of PLC-δ
Authors: Homma Yoshimi , Emori Yasufumi 
Published: November 2023

Abstract
In contrast to what is known about the PLC-β and PLC-γ subfamilies, the activation mechanism of PLC-δ remains unclear. Neither receptors nor transducers that couple to PLC-δ members have been identified. The PLC-δ subfamily contains so far four members: PLC-δ1, -δ2, -δ3, and -δ4. PLC-δ1 and PLC-δ2 can be purified from either rat or bovine brains (4—7). PLC-δ3 is known only at the cDNA level (8). PLC-δ4 cDNA and protein have been reported very recently (9). Structural analyses demonstrate that the molecular structure of PLC-δ contains three characteristic regions: the X and Y regions, which both show sequence similarity to the other PLC isoforms and are essential for enzymatic activity; a region which contains two EF hand-like motifs; and the pleckstrin homology (PH) region (Figure 1).
Analysis ofRNA:protein and RNA:RNA contacts between ribosomes and functional ligands using site-directed cross-linking techniques
Authors: Rinke-Appel Jutta , Brimacombe Richard 
Published: November 2023

Abstract
Cross-linking methods have been widely applied for the investigation of contacts or neighbourhoods of the individual components of a variety of biological systems. The ribosome is particularly well-suited to this type of approach; while on the one hand the ribosomal particles are too large and complex to allow a structural determination by X-ray crystallographic techniques, on the other hand it is this very complexity which makes the ribosome-with its numerous different components and functional ligands-an ideal substrate for cross-linking studies.
Cloning transcription factors by sequence similarity
Authors: Ashworth Alan 
Published: November 2023

Abstract
Knowledge of the primary structure of different classes of transcription factors has revealed the modular nature of these proteins. Functional domains, such as DNA binding motifs, are often highly conserved between different transcription factors. The sequence conservation of these domains can be taken advantage of to isolate clones related to a particular transcription factor. In some cases, these gene families may have more than 500 members (1).
Identification of target genes for a transcription factor by genomic binding-site cloning
Authors: Inoue Satoshi , Kondo, Shigeru , Muramatsu Masami 
Published: November 2023

Abstract
Whereas a great number of genes or cDNAs are isolated by molecular cloning, relatively few genes are known with respect to their function. In order to gain more insight into the life processes of higher eukaryotes, including humans, a greater understanding of gene function is of prime importance.
Analysis of cloned factors
Authors: White Roger , Parker Malcolm 
Published: November 2023

Abstract
The analysis of a cloned transcription factor and the characterization of its functional properties is carried out primarily by the identification and map ping of domains within the protein using deletion and point mutagenesis. This is accomplished by expression of either the factor itself, or altered forms of the protein, in vitro and in vivo in different cell types. The aim of this chapter is to describe some of the approaches and methods used in this analysis. Methods will also be described by which the cloned factor itself, or fragments of the factor, may be overexpressed with the aim of producing large amounts of the protein for biochemical and structural studies. Finally a number of assay systems will be described which may be used to measure the transcriptional activity of the cloned factor and its ability to interact with different components of the cell, in particular other proteins and DNA.
Surface plasmon resonance
Authors: Van Der Merwe P Anton 
Published: November 2023

Abstract
Since the development almost a decade ago (1, 2) of the first biosensor based on surface plasmon resonance (SPR), the use of this technique has increased steadily. Although there are several SPR-based systems (3-5), by far the most widely used one is the BlAcore (1, 2), produced by BlAcore AB, which has developed into a range of instruments (Table 1). By December 1998 over 1200 publications had reported results obtained using the BlAcore. It is likely that it would be even more widely used were it not for its high cost and the pitfalls associated with obtaining accurate quantitative data (5-8). The latter has discouraged many investigators and led to the perception that the technique may be flawed. This is unjustified because the pitfalls are common to many binding techniques and, once understood, they are easily avoided (4, 8, 9). Furthermore, the BlAcore offers particular advantages for analysing weak macromolecular interactions, allowing measurements that are not possible using any other technique (4, 10). This chapter aims to provide guidance to users of SPR, with an emphasis on avoiding pitfalls. No attempt is made to describe the routine operation and maintenance of the BlAcore, as this is comprehensively described in the BlAcore instrument manual (see Section 9.3). Although written for BlAcore users, the general principles will be applicable to experiments on any SPR instrument.
Introduction to macromolecular visualization
Authors: Martz Eric , Driscoll Timothy 
Published: November 2023

Abstract
The ability to view a macromolecule in three dimensions is a great asset to science students and researchers alike. A decade ago, the primary method for examining 3D molecular structures was stereoscopic viewing, which often required training or special glasses. Now a number of excellent computer tools, many of them free, are available for rendering molecular structures in 3D, interacting with them, and customizing their appearance. Some are stand-alone programs like WebLab ViewerPro and DeepView Swiss-Pdb Viewer. Others are browser plug-ins like Chime. Still others, like Protein Explorer, a browser-based package using the Chime plug-in, can work entirely over the Web. This Chapter explains which of these programs to use for your particular objectives. However, the scope of this Chapter is limited-it does not cover some other freeware visualization packages with very strong features, including Cn3D, DINO, MAGE, PyMOL, and VMD (find links in the World Index, see the WebLinks box below).
Invasive Hemodynamic and Vasoreactivity Testing with Inhaled Iloprost in Children with Pulmonary Arterial Hypertension Associated with Congenital Heart Defects
Authors: Li Qiangqiang , He Yuan , Zhang Chen , Gu Hong 
Published: November 2023

Objective:
Invasive hemodynamic evaluation and acute vasoreactivity testing are recommended in the diagnosis of pulmonary arterial hypertension (PAH), but their clinical role in children with PAH associated with congenital heart defects (CHD) is unclear. This study aims to investigate acute hemodynamic responses to inhaled iloprost, and its role in prognosis in children with PAH-CHD.

Methods:
A retrospective analysis was conducted on 83 pediatric patients with PAH-CHD in whom invasive hemodynamics were evaluated before and after a single inhaled dose of iloprost at a single center between 2010 and 2022. Details of the CHD corrective operation, medical treatment, and outcome for each patient were obtained via medical records or telephone contact. A composite endpoint of all-cause death, admission for worsening heart failure during follow-up, and event-free survival was defined, and risk factors associated with this composite endpoint were analyzed.

Results:
The patient age was (11.3 ± 4.6) years, and 60 (72.3%) were female. Fifty-nine (71.1%) patients were diagnosed with Eisenmenger syndrome. After iloprost inhalation, mean pulmonary artery pressure decreased from (78.2 ± 11.5) to (72.3 ± 13.2) mmHg (P < 0.001), and pulmonary vascular resistance index (PVRI) decreased from (18.0 ± 7.9) to (14.5 ± 8.1) WU·m2 (P < 0.001). A total of 38 (45.8%) patients had a positive response, defined as a PVRI decrease >25% with stable systemic pressure. Thirty-seven (44.6%) patients underwent a corrective CHD operation at a median of 24 d after hemodynamic evaluation. Nine patients died, and 15 met the composite endpoint during a follow-up period of 5.4 (3.8, 8.8) years. Five-year event-free survival estimates were 96.7% (95% confidence interval: 90.3%–100%) in patients with a positive response, and 82.8% (95% confidence interval: 71.7%–94.5%) in patients with a non-positive response (log-rank P = 0.012). A positive PVRI response and higher pulmonary arterial oxygen saturation after iloprost inhalation, lower baseline brain natriuretic peptide, and PAH-targeted therapy at follow-up were significantly associated with a favorable clinical outcome. A positive acute vasoreactivity testing response and PAH-targeted therapy at follow-up were independent predictors of outcome in multivariate Cox analysis.

Conclusions:
Acute inhalation of iloprost can lead to a significant decrease in hemodynamic parameters. Responsiveness to inhaled iloprost is associated with better outcomes and can be a valuable predictor of outcomes.
Treatment of Internal Mammary Nodes is Associated With Improved Overall Survival in Breast Cancer
Authors: Shaikh Parvez Memet , Mulherkar Ria , Khasawneh Mohammad T. , Clump David , Hazard-Jenkins Hannah , Hafez Maria , Vargo John A. 
Published: November 2023

Introduction:
The role of internal mammary nodal irradiation (IMNI) as a component of regional nodal radiotherapy is a controversial issue in breast radiation oncology with conflicting results presented in recent landmark trials. We thus created a meta-analysis of available data to better ascertain the potential benefit of IMNI. We hypothesize that with the increased power available within a meta-analysis, IMNI will prove to improve overall survival (OS) in breast cancer.

Methods:
Literature search was conducted for prospective studies comparing IMNI to no IMNI. Primary endpoint was OS and secondary endpoints included local recurrence, regional recurrence, disease-free survival (DFS), breast cancer mortality (BCM), distant metastasis-free survival (DMFS), grade 2+ skin toxicity, cardiac events, and pneumonitis events. Subgroup analyses were performed for tumor location (medial/central vs. lateral), and nodal status (pN+ vs. pN0). Fixed-effect model was used if there was no heterogeneity, random-effects model otherwise.

Results:
Four studies with a total of 5258 patients (IMNI: n=2592; control: n=2666) were included in the study. Pooled results showed IMNI significantly improved OS for all-comers (hazard ratio [HR]=0.89; 95% CI 0.81-0.97; P=0.008), as well as subgroups of pN+ with medial/central tumor location (HR=0.84; 95% CI 0.73-0.96; P=0.01) and pN+ with lateral tumor location (HR=0.87; 95% CI 0.77-0.99; P=0.04). There was no significant difference in OS for subgroups of pN0 and medial/central tumor location. There was no difference in local recurrence, but regional recurrence was significantly improved (P=0.04). Endpoints of DFS (HR 0.91, 95% CI 0.84-0.99 P=0.03), BCM (HR 0.87, 95% CI 0.77-0.98, P=0.03), and DMFS (HR=0.87; 95% CI, 0.78-0.98; P=0.02) were all improved with IMNI. Grade 2+ skin toxicity, cardiac events and pneumonitis events were not significantly different between patient in the IMNI and no IMNI groups.

Conclusion:
Inclusion of IMN irradiation improves OS, DFS, BCM, and DMFS in breast cancer. Largest effect on OS was noted in the subgroup of patients with pN+ and medial/central tumor location.
Requirements for the successful set-up of an international, multicentre, multilanguage, noninterventional patient-reported outcomes pain study
Authors: Weinmann Claudia , Komann Marcus , Liedgens Hiltrud , Pogatzki-Zahn Esther M. , Fletcher Dominique , Meissner Winfried 
Published: November 2023

BACKGROUND
For setting up and implementing an international follow-up survey study in different countries, many prerequisites are necessary: obtaining permission or licenses to use the tools/questionnaires selected, translating the questionnaires according to a standardised translation process, implementing the survey technically, recruiting, training and monitoring the data collecting study sites, obtaining ethics approvals and data protection clearance, developing the study and training materials, and registering the study.

OBJECTIVE
Within the IMI-PainCare project (Improving the care of patients suffering from acute or chronic pain), we needed to conduct a large noninterventional patient survey – PROMPT NIT-1 – aiming to find out which PROMs (patient-reported outcome measures) work best for postoperative pain assessment. Planning and implementing this international multicentre, multilanguage, follow-up study was complex and time consuming. Consequently, experiences from its successful implementation may help future study coordinators.

DESIGN
The NIT-1 surveys consisted of several established tools. We planned to include 4000 patients, and to collect data at seven time points: baseline, postoperative days (POD) 1, 3 and 7 and follow-up on postoperative months (POM) 1, 3 and 6.

SETTING
Eighteen hospitals, mostly primary care, across Europe collected data for our study. First patient in was in August 2019, last patient out in June 2022.

PATIENTS
Adult patients of both sexes who had undergone one of four defined surgical procedures answered our surveys; total knee arthroplasty (TKA), breast surgery, endometriosis surgery and sternotomy.

MAIN OUTCOMES
Results from the study related to the primary and secondary outcomes are not part of this publication and will be presented separately. This publication describes only the technical, administrative and coordinating implementation.

RESULTS
Although the COVID-19 pandemic affected patient inclusion, and some preparations took longer than foreseen, 18 hospitals across Europe included 3303 patients. We were able to attain sufficient case numbers, and follow-up rates were very good.

CONCLUSION
The setting up and implementation of a large, international, multilanguage data collection is challenging and ambitious but feasible if it is well planned, if sufficient resources (project management and technical/IT expertise, qualified staff, funding, commitment) are available and if the timeframe for preparing the study is generously calculated.

TRIAL REGISTRATION
clinicaltrials.gov (ID NCT03834922).
Taxonomic descriptions of Aeromicrobium duanguangcaii sp. nov., Aeromicrobium wangtongii sp. nov. and Aeromicrobium senzhongii sp. nov.
Authors: Ye Lin , Zhang Gui , Yang Jing , Yang Caixin , Liu Yue , Pu Ji , Huang Yuyuan , Zhang Sihui , Jin Dong , Lu Shan , Liu Liyun , Xu Jianguo 
Published: November 2023

Six Gram-stain-positive, facultative anaerobic, nonmotile and rod-shaped strains, designated zg-Y50T, zg-Y1362, zg-Y1379T, zg-Y869, zg-629T and zg-Y636, were isolated from the intestinal contents of Marmota himalayana in Qinghai Province, PR China. Strains zg-Y50T, zg-Y1379T and zg-629T exhibited the highest 16S rRNA gene sequence similarities of 99.2, 98.9 and 98.8 % to

Aeromicrobium choanae

9 H-4T,

Aeromicrobium ginsengisoli

JCM 14732T and

Aeromicrobium flavum

TYLN1T, respectively. Phylogenetic and phylogenomic analyses based on the 16S rRNA gene and genomic sequences, respectively, revealed that the six strains formed three distinct clades within the genus

Aeromicrobium

. The genome sizes of strains zg-Y50T, zg-Y1379T and zg-629T were 3.1–3.7 Mb, with DNA G+C contents of 69.6–70.4 mol%. Average nucleotide identity and digital DNA–DNA hybridization values between each novel strain and available members of the genus

Aeromicrobium

were all below species thresholds. All novel strains contained MK-9 (H4) as the major menaquinone and diphosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol as the polar lipids. The predominant fatty acid of the six isolates was C18 : 1
ω9c. The cell-wall peptidoglycan contained ʟʟ-diaminopimelic acid as the diagnostic diamino acid. Based on the results from this polyphasic taxonomic study, three novel species in the genus

Aeromicrobium

are proposed, namely, Aeromicrobium duanguangcaii sp. nov. (zg-Y50T=GDMCC 1.2981T=KCTC 49764T), Aeromicrobium wangtongii sp. nov. (zg-Y1379T=GDMCC 1.2982T=KCTC 49765T) and Aeromicrobium senzhongii sp. nov. (zg-629T=CGMCC 1.17414T=JCM 33888T).
Trinickia violacea sp. nov. and Trinickia terrae sp. nov., isolated from forest soil
Authors: Gao Zeng-hong , Guo Xiu-yin , Liu Yi-zhi , Zhang Qiu-mei , Tong Xin-jie , Qiu Li-hong 
Published: November 2023

Two Gram-stain negative, aerobic and rod-shaped bacterial strains, DHOD12T and 7GSK02T, were isolated from forest soil of Dinghushan Biosphere Reserve, Guangdong Province, PR China. Strain DHOD12T grew at 4–42 °C (optimum, 28–33 °C), pH 4.0–8.5 (optimum, pH 5.5–6.5) and in the presence of 0–1.5 % (w/v; optimum, 0–0.5 %)NaCl; while strain 7GSK02T grew at 12–42 °C (optimum, 28–33 °C), pH 4.0–8.5 (optimum, pH 5.0–6.0) and in the presence of 0–0.5 % (w/v; optimum, 0 %) NaCl. Strains DHOD12T and 7GSK02T had the highest 16S rRNA sequence similarities of 98.0 and 98.3 % with the same species Trinickia mobilis DHG64T, respectively, and 98.4 % between themselves. In the 16S rRNA phylogeny, they formed a clade that was sister to a major cluster consisting of all described

Trinickia

species. Phylogenomic analyses with the UBCG and PhyloPhlAn methods consistently showed that strains DHOD12T and 7GSK02T formed a clade with T. mobilis DHG64T that was a sister of a cluster containing the remainder of the

Trinickia

species. The DNA G+C contents of strains DHOD12T and 7GSK02T were 63.1 and 64.6 mol%, respectively. Digital DNA–DNA hybridization and average nucleotide identity values of strains DHOD12T, 7GSK02T and their closely related strains were in the ranges of 21.6–31.4 % and 77.1–86.9 %, respectively. These two strains had the same major respiratory quinone, ubiquinone-8, and both had C16 : 0, C17 : 0 cyclo and summed feature 8 (C18 : 1
ω7c/C18 : 1
ω6c) as their major fatty acids. Their major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. Genomic analysis indicated that the two strains could have the potential to degrade aromatic compounds like other

Trinickia

species. On the basis of phenotypic and phylogenetic results, strains DHOD12T and 7GSK02T represent two novel species of the genus

Trinickia

, for which the names Trinickia violacea sp. nov. (type strain DHOD12T=LMG 30258T=CGMCC 1.15436T) and Trinickia terrae sp. nov. (type strain 7GSK02T=CGMCC 1.15432T=KCTC 62468T) are proposed.
Fatty acid metabolism in neutrophils promotes lung damage and bacterial replication during tuberculosis
Authors: Sankar Poornima , Ramos Ramon Bossardi , Corro Jamie , Saqib Mohd , Nafiz Tanvir Noor , Bhargavi Gunapati , Ojha Anil K , Cai Yi , Subbian Selvakumar , Mishra Bibhuti B. 
Published: November 2023

ABSTRACTMycobacterium tuberculosis(Mtb) infection triggers a significant influx of neutrophils to the lungs, which is linked to tuberculosis (TB) severity. The mechanism by which Mtb infection induces neutrophillic inflammation remains unclear. Using a clinically relevant and hypervirulent Mtb strain from the W-Beijing family, HN878, we found that genes related to both glycolysis and fatty acid metabolism are upregulated in the lung neutrophils of susceptible mice. Similar effects in gene expression were observed in rabbits, and humans with pulmonary TB compared to healthy controls. Inhibiting glycolysis with 2-deoxy D-glucose (2-DG) exacerbated disease pathology, while fatty acid oxidation (FAO) inhibitor Etomoxir (ETO) improved outcomes by reducing weight loss, immunopathology, and bacterial replication within neutrophils in genetically susceptible mice. Notably, ETO reduced neutrophil production in the bone marrow and their recruitment to the lungs. ETO specifically restrained the recruitment of Ly6Glow/dimimmature neutrophil population, which is elevated during disease progression and harbors the bulk of bacilli. In a transwell setup, we demonstrated that ETO dose-dependently inhibited neutrophil chemotaxis towards infected macrophages. In summary, our research highlights the crucial role of fatty acid metabolism in regulating neutrophilic inflammation during TB and provides a rationale for targeting immunometabolism of neutrophils for potential TB treatment.