Mitochondrial fission induces immunoescape in solid tumors through decreasing MHC-I surface expression.

Mitochondrial dynamics can regulate Major Histocompatibility Complex (MHC)-I antigen expression by cancer cells and their immunogenicity in mice and in patients with malignancies. A crucial role in the mitochondrial fragmentation connection with immunogenicity is played by the IRE1α-XBP-1s axis. XBP-1s is a transcription factor for aminopeptidase TPP2, which inhibits MHC-I complex cell surface expression likely by degrading tumor antigen peptides. Mitochondrial fission inhibition with Mdivi-1 upregulates MHC-I expression on cancer cells and enhances the efficacy of adoptive T cell therapy in patient-derived tumor models. Therefore mitochondrial fission inhibition might provide an approach to enhance the efficacy of T cell-based immunotherapy.

Bibliometric Analysis of Chimeric Antigen Receptor-Based Immunotherapy in Cancers From 2001 to 2021.

Background: Chimeric antigen receptor (CAR)-based immunotherapy has shown great potential for the treatment of both hematopoietic malignancies and solid tumors. Nevertheless, multiple obstacles still block the development of CAR-based immunotherapy in the clinical setting. In this study, we aimed to summarize the research landscape and highlight the front lines and trends of this field. Methods: Literature published from 2001 to 2021 was searched in the Web of Science Core Collection database. Full records and cited references of all the documents were extracted and screened. Bibliometric analysis and visualization were conducted using CiteSpace, Microsoft Excel 2019, VOSviewer and R software. Results: A total of 5981 articles and reviews were included. The publication and citation results exhibited increasing trends in the last 20 years. Frontiers in Immunology and Blood were the most productive and most co-cited journals, respectively. The United States was the country with the most productive organizations and publications in the comprehensive worldwide cooperation network, followed by China and Germany. June, C.H. published the most papers with the most citations, while Maude, S.L. ranked first among the co-cited authors. The hotspots in CAR-based therapy research were multiple myeloma, safety and toxicity, solid tumors, CAR-engineered immune cells beyond T cells, and gene editing. Conclusion: CAR-based immunotherapy is a promising treatment for cancer patients, and there is an emerging movement toward using advanced gene modification technologies to overcome therapeutic challenges, especially in solid tumors, and to generate safer and more effective universal CAR-engineered cell products.

Correlation Between TCF7+ T Cells and Prognosis of Patients With Oral Squamous Cell Carcinoma.

Objective: To investigate whether TCF7+ T cells constitute an important factor to improve the existing postoperative prediction model for patients with oral squamous cell carcinoma. Method: TCF7+ T cells were detected in the tissues of 167 OSCC patients by multiplex immunofluorescence. The percentage of TCF7+ T cells was transformed into a dichotomous variable, combined with the clinicopathological data for the OSCC patients, and then subjected to univariate and multivariate analyses. The derived independent predictors were then incorporated into risk models to analyze their relationship with the prognosis of patients. Results: The high TCF7+ group had a better prognosis than the low TCF7+ group (OS: p<0.001; RFS: p<0.001). Univariate and multivariate analyses showed that TCF7+ T cells serve as an independent predictor of OSCC (univariate/multivariate analysis: p<0.001). In Cox risk progression models, inclusion of the TCF7+ T cell percentage improved the predictive accuracy of Grade and TNM stage (Grade-OS/RFS: p<0.001; TNM-OS/RFS: p<0.001; TNM+Grade-OS: p<0.001, TNM+Grade-RFS: p=0.004). Inclusion of the TCF7+ T cell percentage improved the clinical utility. Conclusions: TCF7+ T cells can act as an independent predictor for postoperative OSCC patients. The inclusion of TCF7+ T cells improved the predictive accuracy and clinical utility of the nomograms to different degrees.

The validation of oxygen uptake efficiency slope in patients with stroke.

ABSTRACT: To evaluate the real aerobic capacity is difficult due to impaired limbs function in stroke patients. Oxygen uptake efficiency slope (OUES) could represent the aerobic capacity in submaximal exercise test. Hence, we designed this observational study to investigate the application of the OUES for evaluating aerobic capacity in these patients.Thirty-seven stroke patients were classified into 2 groups according to their Brunnstrom stage of affected lower limbs. Patients underwent cardiopulmonary exercise testing to assess cardiorespiratory fitness. Minute ventilation and oxygen consumption were measured, and OUES was calculated, compared with healthy reference values, and correlated with the peak oxygen consumption. The predictive validity of submaximal OUES was derived.Study participants' OUES (median 566.2 [IQR, 470.0-711.6]) was 60% of healthy reference values and correlated positively with the peak oxygen consumption (r = 0.835) (P < .01). The predictive validity of oxygen uptake efficiency slope at 50% of maximal exercise duration (OUES50) and oxygen uptake efficiency slope at 75% of maximal exercise duration (OUES75) for oxygen uptake efficiency slope at 100% of maximal exercise duration (OUES100) was 0.877 and 0.973, respectively (P < .01). The OUES50, OUES75, and OUES100 groups were not significantly different; agreement of submaximal and maximal OUES values was strong.OUES is a valuable submaximal index for evaluating cardiorespiratory fitness in stroke patients. Moderate-to-high concurrent validity of this parameter with peak oxygen consumption and the high predictive validity of OUES50 and OUES75 for OUES100 suggest maximal exercise testing in stroke patients who cannot reach maximal exercise is unnecessary.

Differential Patterns of Gyral and Sulcal Morphological Changes During Normal Aging Process.

The cerebral cortex is a highly convoluted structure with distinct morphologic features, namely the gyri and sulci, which are associated with the functional segregation or integration in the human brain. During the lifespan, the brain atrophy that is accompanied by cognitive decline is a well-accepted aging phenotype. However, the detailed patterns of cortical folding change during aging, especially the changing age-dependencies of gyri and sulci, which is essential to brain functioning, remain unclear. In this study, we investigated the morphology of the gyral and sulcal regions from pial and white matter surfaces using MR imaging data of 417 healthy participants across adulthood to old age (21-92 years). To elucidate the age-related changes in the cortical pattern, we fitted cortical thickness and intrinsic curvature of gyri and sulci using the quadratic model to evaluate their age-dependencies during normal aging. Our findings show that comparing to gyri, the sulcal thinning is the most prominent pattern during the aging process, and the gyrification of pial and white matter surfaces were also affected differently, which implies the vulnerability of functional segregation during aging. Taken together, we propose a morphological model of aging that may provide a framework for understanding the mechanisms underlying gray matter degeneration.

Easy Synthesis of BiVO4 for Photocatalytic Overall Water Splitting.

Developing a photocatalyst system to generate hydrogen from water is a topic of great interest for fundamental and practical importance. In this study, we develop a new Z-scheme photocatalytic system for overall water splitting that consists of Rh/K4Nb6O8 for H2 evolution, Pt/BiVO4 for O2 evolution, and I-/IO3 - for an electron mediator under UV light irradiation. The oxygen evolution photocatalyst BiVO4 was prepared by the microwave-assisted hydrothermal method. The method is fast and simple, as compared to conventional hydrothermal synthesis. The catalysts were characterized by powder X-ray diffraction, scanning electron microscopy, transmission electron microscopy, and UV-visible spectroscopy. The photocatalytic water splitting is investigated in (i) aqueous AgNO3 as sacrificial electron scavengers and (ii) a Z-scheme photocatalytic water splitting system. The BiVO4 photocatalysts prepared by the microwave-assisted hydrothermal method not only showed a very high oxygen evolution rate (2622 µmol g-1 h-1) of water splitting reaction in an aqueous AgNO3 solution but also achieved a high H2 evolution rate (340 µmol g-1 h-1) and O2 evolution rate (172 µmol g-1 h-1) in a Z-scheme overall water splitting system.

Sulfur-doped g-C3N4 nanosheets for photocatalysis: Z-scheme water splitting and decreased biofouling.

In this study, an S-doped g-C3N4 nanosheet was prepared as a photocatalyst for effective oxygen evolution reaction. Sulfur plays a crucial role in S-doped g-C3N4 not only in increasing the charge density but also in reducing the energy band gap of S-doped g-C3N4 via substitution of nitrogen sites. S-doped g-C3N4 can serve as an oxygen-evolved photocatalyst, when combined with Ru/SrTiO3:Rh in the presence of [Co(bpy)3]3+/2+ as an electron mediator, enables photocatalytic overall water splitting under visible light irradiation with hydrogen and oxygen production rates of 24.6 and 14.5 µmol-h-1, respectively. Moreover, the photocatalytic overall water splitting to produce H2 and O2 using this Z-scheme system could use for five runs to at least 94.5 h under visible light irradiation. On the other hand, S-doped g-C3N4 can reduce biofouling by bacteria such as Escherichia coli by more than 70%, by simply incubating the S-doped g-C3N4 sample with bacterial solution under light irradiation. Our results suggest that S-doped g-C3N4 is a potentially effective, green, and promising material for a variety of photocatalytic applications.

Effects of a comprehensive physical therapy on moderate and severe obstructive sleep apnea- a preliminary randomized controlled trial.

BACKGROUND: Critically compromised by upper airway anatomical impaired properties, obstructive sleep apnea (OSA) can be categorized into different phenotypic traits, mainly including oropharyngeal muscle dysfunction. The upper airway muscle strength training was targeted on oropharyngeal muscle dysfunction by re-educating the oropharyngeal muscles to maintain the upper airway patency. OSA was characterized with multilevel collapsibility of the upper airway; however, the programs are still inconsistent and the effects are unknown. Therefore, the purpose of this study was to investigate the effects of a comprehensive physical therapy on OSA. METHODS: Fifteen subjects with newly diagnosed moderate or severe OSA (AHI ≥ 15) were randomized into intervention and control groups. The intervention group underwent a 12-week-intervention of hospital based physical therapy, while the control group was kept on waiting for 12 weeks. Polysomnography (PSG) data, oropharyngeal and respiratory muscle performance were measured before and after intervention. RESULTS: In intervention group (n = 8), AHI was significantly improved (from 46.96 ± 19.45 to 32.78 ± 10.78 events/h, p = 0.017); in control group (n = 7), AHI was significantly increased (from 35.77 ± 17.49 to 42.96 ± 17.32 events/h, p = 0.043). While the control group remained no change between pre- and post- intervention, the intervention group demonstrated that other PSG outcomes significantly improved, including arousal index (46.04 ± 18.9 versus 32.98 ± 8.35/h), mean SpO2 (92.88 ± 2.1 versus 94.13 ± 1.46%), and oxygen desaturation index (ODI) (31.13 ± 19.48 versus 20.57 ± 7.83/h). CONCLUSION: This comprehensive physical therapy can be prescribed for the significant clinical improvement on sleep apnea for the patients with moderate and severe OSA.

Photocatalytic Water Splitting on Rh/K4Nb6O17 Nanosheets.

The layered potassium niobate, K4Nb6O17, is known as a photocatalyst for hydrogen production from water splitting under UV light. Here we show that potassium niobate nanosheets can be obtained by exfoliation of K4Nb6O17 followed by proton exchange. Photocatalytic water splitting has been improved in this work by loading Rh nanoparticles as cocatalyst. The catalysts were characterized by powder X-ray diffraction (XRD), X-ray photoelectron spectroscopy analysis (XPS), and ultraviolet-visible spectroscopy (UV-vis), scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Their photocatalytic activities were in (i) an aqueous MeOH, used as sacrificial hole scavengers for hydrogen production and (ii) pure water for hydrogen and oxygen production under UV light irradiation. The potassium niobate nanosheets with uniform dispersion of Rh nanoparticles exhibited a very high photocatalytic overall water splitting activity, where the H2 production and O2 production rate were 142 µmol h-1 g-1 and 68 µmol h-1 g-1, which was about 10 times higher than that exhibited by the origin K4Nb6O17 photocatalyst.

Hash Transformation and Machine Learning-Based Decision-Making Classifier Improved the Accuracy Rate of Automated Parkinson's Disease Screening.

Digitalized hand-drawn pattern is a noninvasive and reproducible assistive manner to obtain hand actions and motions for evaluating functional tremors and upper-limb movement disorders. In this study, spirals and straight lines in polar coordinates are used to extract polar expression features such as the key parameters deviation (cm) and accumulation angle (rad). These parameters are quantitative manner to scale the variations of functional tremors in normal control subjects and patients with Parkinson's disease (PD) and essential tremor (ET). However, difficulty arises in using nonlinear polar expression features in the two-dimensional feature space to separate normal control subjects from those with PD and ET. To solve the nonlinear separable classification problem, hash transformation is used to map polar expression features to a high-dimensional space using hash weighing function and modulo operation. Then, a machine learning method, such as the generalized regression neural network (GRNN), is implemented to train a decision-making classifier using the particle swarm optimization (PSO) algorithm for possible class assessment. With the enrolled data from 50 subjects, the fivefold cross validation, mean true positive, mean true negative, and mean hit rates of 98.93%, 98.96%, and 98.93%, respectively, are obtained to quantify the performance of the proposed decision-making classifier to identify normal controls and subjects with PD or ET. The experimental results indicate that the proposed screening model can improve the accuracy rate compared with the conventional machine learning classifier.

Models of anatomically based oropharyngeal rehabilitation with a multilevel approach for patients with obstructive sleep apnea: a meta-synthesis and meta-analysis.

OBJECTIVES: Obstructive sleep apnea (OSA) is a sleep-related breathing disorder associated with dysfunction of oropharyngeal muscles to maintain upper airway patency during sleep. Oropharyngeal rehabilitation (OPR) was developed to restore, reconstruct, and reeducate oropharyngeal muscle function, but current protocols and effectiveness of OPR have been inconsistent. The purpose of this study was to review (1) indications of OPR, (2) protocols of OPR, and (3) effectiveness of OPR. METHODS: We searched MEDLINE, EMBASE, and the Cochrane Library and then conducted both meta-synthesis and meta-analysis according to the statement of Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA). RESULTS: A total of eight studies with 203 patients were included. By means of meta-synthesis, the patients with middle age, BMI < 40 kg/m2, mild-to-moderate OSA, and non-severe upper airway anatomical abnormality were found to benefit from OPR. The protocol of OPR was summarized to be an anatomically based, multilevel approach, including the retropalatal, retroglossal, hypopharyngeal, TMJ, and facial levels. By using meta-analysis, overall outcomes were presented as apnea hypopnea index (AHI) with significant improvement from 25.2 ± 7.8/h to 16.1 ± 6.6/h (mean difference [MD] - 9.8 [95% CI - 11.0 to - 8.6], p < 0.0001); the lowest oxygen saturation (LSAT) improved from 80.2 ± 4.7 to 83.8 ± 2.9% (MD 3.0% [95% CI 2.0 to 4.0], p < 0.0001); Epworth sleepiness scale (ESS) improved from 11.8 ± 1.9 to 6.3 ± 1.6 (MD - 5.9 [95% CI - 7.5 to - 4.2], p < 0.001), neck circumference (NC) from 35.2 ± 1.1 to 34.7 ± 0.9 cm (MD - 0.6 [95% CI - 0.9 to - 0.2], p = 0.002), BMI from 24.8 ± 3.7 to 24.8 ± 4.1 kg/m2 (MD - 0.0; 95% CI - 0.5 to 0.5, p = 0.95). All outcomes except BMI demonstrated significant improvement from OPR. CONCLUSIONS: Meta-analysis of previous OPR reports shows an improvement in AHI of 39%, compared with the usual surgical definition of success at 50%. Only mild and moderate cases of OSA were referred for OPR in the prior studies. In order to improve outcomes with OPR, a comprehensive approach to rehabilitation should be emphasized.

Worsen depression after viscosupplementation treatment for geriatric people with knee osteoarthritis?

Background/objective: Knee osteoarthritis (OA) in older people may result in psychological impairment, including anxiety and depression. This study investigates the effect of intraarticular hyaluronic acid injection (IAHA) on geriatric patients with OA. Method: A total of 102 geriatric patients with knee OA undergoing IAHA were prospectively enrolled in this study. Geriatric Depression Scale (GDS), State-Trait Anxiety Inventory (STAI), Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and the International Knee Documentation Committee Subjective Knee Evaluation Form (IKDC), and Visual Analogue Scale (VAS) for pain were recorded. All outcomes were measured at baseline before injection and during two, four, and six month follow-ups. Results: IAHA had a significant short-term effect, relieving pain at the two month follow-up, but the effect was weaker at the four month follow-up. Both IKDC and WOMAC scores were significantly improved at the two month follow-up. Viscosupplementation did not improve STAI. GDS exhibited significant deterioration at the four month follow-up. Conclusions: Although IAHA for the treatment of OA provided short-term efficiency, it had no effects on anxiety and increased depression of geriatric people. Health education should be provided with caution before viscosupplementation treatment to manage expectations of the efficacy of treatment for geriatric OA patients.

Antecedentes/objetivo: La artrosis de rodilla (AR) en personas mayores puede causar ansiedad y depresión. Se investiga el efecto de la inyección intra-articular de ácido hialurónico (IAAH) en pacientes mayores con AR. Método: Ciento dos pacientes mayores tratados con IAAH fueron inscritos de forma prospectiva. Contestaron la Geriatric Depression Scale (GDS), State-Trait Anxiety Inventory (STAI), Western Ontario and McMaster University Osteoarthritis Index (WOMAC), International Knee Documentation Committee Subjective Knee Evaluation Form (IKDC) y Visual Analogue Scale (VAS). Se evaluó antes de la inyección y durante el seguimiento a dos, cuatro y seis meses. Resultados: La IAAH produjo un efecto significativo a corto plazo, aliviando el dolor a los dos meses de seguimiento, pero el efecto fue más débil a los cuatro meses. Las puntuaciones del IKDC y WOMAC mejoraron significativamente a los dos meses de seguimiento. La visco-suplementación no mejoró la ansiedad. La GDS mostró un deterioro significativo a los cuatro meses de seguimiento. Conclusiones: Aunque la IAAH fue efectiva a corto plazo, no tuvo efectos sobre la ansiedad, y aumentó la depresión. Debe prestarse atención a la educación para la salud antes del tratamiento con viscosuplementación para controlar las expectativas de la eficacia del tratamiento en pacientes mayores con AR.

Worsen depression after viscosupplementation treatment for geriatric people with knee osteoarthritis?

Background/objective: Knee osteoarthritis (OA) in older people may result in psychological impairment, including anxiety and depression. This study investigates the effect of intraarticular hyaluronic acid injection (IAHA) on geriatric patients with OA. Method: A total of 102 geriatric patients with knee OA undergoing IAHA were prospectively enrolled in this study. Geriatric Depression Scale (GDS), State-Trait Anxiety Inventory (STAI), Western Ontario and McMaster University Osteoarthritis Index (WOMAC) and the International Knee Documentation Committee Subjective Knee Evaluation Form (IKDC), and Visual Analogue Scale (VAS) for pain were recorded. All outcomes were measured at baseline before injection and during two, four, and six month follow-ups. Results: IAHA had a significant short-term effect, relieving pain at the two month follow-up, but the effect was weaker at the four month follow-up. Both IKDC and WOMAC scores were significantly improved at the two month follow-up. Viscosupplementation did not improve STAI. GDS exhibited significant deterioration at the four month follow-up. Conclusions: Although IAHA for the treatment of OA provided short-term efficiency, it had no effects on anxiety and increased depression of geriatric people. Health education should be provided with caution before viscosupplementation treatment to manage expectations of the efficacy of treatment for geriatric OA patients

Antecedentes/objetivo: La artrosis de rodilla (AR) en personas mayores puede causar ansiedad y depresión. Se investiga el efecto de la inyección intra-articular de ácido hialurónico (IAAH) en pacientes mayores con AR. Método: Ciento dos pacientes mayores tratados con IAAH fueron inscritos de forma prospectiva. Contestaron la Geriatric Depression Scale (GDS), State-Trait Anxiety Inventory (STAI), Western Ontario and McMaster University Osteoarthritis Index (WOMAC), International Knee Documentation Committee Subjective Knee Evaluation Form (IKDC) y Visual Analogue Scale (VAS). Se evaluó antes de la inyección y durante el seguimiento a dos, cuatro y seis meses. Resultados: La IAAH produjo un efecto significativo a corto plazo, aliviando el dolor a los dos meses de seguimiento, pero el efecto fue más débil a los cuatro meses. Las puntuaciones del IKDC y WOMAC mejoraron significativamente a los dos meses de seguimiento. La visco-suplementación no mejoró la ansiedad. La GDS mostró un deterioro significativo a los cuatro meses de seguimiento. Conclusiones: Aunque la IAAH fue efectiva a corto plazo, no tuvo efectos sobre la ansiedad, y aumentó la depresión. Debe prestarse atención a la educación para la salud antes del tratamiento con viscosuplementación para controlar las expectativas de la eficacia del tratamiento en pacientes mayores con AR

Reduction of lipid contamination in MR spectroscopy imaging using signal space projection.

PURPOSE: Lipid contamination can complicate the metabolite quantification in MR spectroscopic imaging (MRSI). In addition to various experimental methods demonstrated to be feasible for lipid suppression, the postprocessing method is beneficial in the flexibility of applications. In this study, the signal space projection (SSP) algorithm is proposed to suppress the lipid signal in the MRSI. METHODS: The performance of lipid suppression using SSP and SSP combined with the Papoulis-Gerchberg (PG) algorithm (PG+SSP) is examined in 2D MRSI data and the results were compared with outer volume saturation (OVS) methods. Up to 10 lipid spatial components were extracted by SSP from lipid signals in the range of 0.8~1.5 ppm. RESULTS: Our results show that most lipid signals were found in the first 4 to 5 components and that lipid signals on the spectra can be suppressed using 4 to 5 components. Metabolites concentrations were quantified using LCModel. Two regions of interest (ROIs) were manually selected on the peripheral and inner brain regions. The quantification of metabolites in terms of fitting reliability (CRLB) and spatial variations within ROIs (SpaVar) is improved using SSP. When 5 to 6 components were used in SSP and PG+SSP, the metabolite concentrations and the associated SpaVar and CRLB are at the same level as those from the OVS. CONCLUSION: We have demonstrated that the SSP method can be used to suppress the lipid signals of MRSI and SSP with 5 to 6 components is suggested to have a similar suppression performance as the OVS method.

Fluorescent fusions of the N protein of phage Mu label DNA damage in living cells.

The N protein of phage Mu was indicated from studies in Escherichia coli to hold linear Mu chromosomes in a circular conformation by non-covalent association, and thus suggested potentially to bind DNA double-stranded ends. Because of its role in association with linear Mu DNA, we tested whether fluorescent-protein fusions to N might provide a useful tool for labeling DNA damage including double-strand break (DSB) ends in single cells. We compared N-GFP with a biochemically well documented DSB-end binding protein, the Gam protein of phage Mu, also fused to GFP. We find that N-GFP produced in live E. coli forms foci in response to DNA damage induced by radiomimetic drug phleomycin, indicating that it labels damaged DNA. N-GFP also labels specific DSBs created enzymatically by I-SceI double-strand endonuclease, and by X-rays, with the numbers of foci corresponding with the numbers of DSBs generated, indicating DSB labeling. However, whereas N-GFP forms about half as many foci as GamGFP with phleomycin, its labeling of I-SceI- and X-ray-induced DSBs is far less efficient than that of GamGFP. The data imply that N-GFP binds and labels DNA damage including DSBs, but may additionally label phleomycin-induced non-DSB damage, with which DSB-specific GamGFP does not interact. The data indicate that N-GFP labels DNA damage, and may be useful for general, not DSB-specific, DNA-damage detection.

Optimal Fungal Space Searching Algorithms.

Previous experiments have shown that fungi use an efficient natural algorithm for searching the space available for their growth in micro-confined networks, e.g., mazes. This natural "master" algorithm, which comprises two "slave" sub-algorithms, i.e., collision-induced branching and directional memory, has been shown to be more efficient than alternatives, with one, or the other, or both sub-algorithms turned off. In contrast, the present contribution compares the performance of the fungal natural algorithm against several standard artificial homologues. It was found that the space-searching fungal algorithm consistently outperforms uninformed algorithms, such as Depth-First-Search (DFS). Furthermore, while the natural algorithm is inferior to informed ones, such as A*, this under-performance does not importantly increase with the increase of the size of the maze. These findings suggest that a systematic effort of harvesting the natural space searching algorithms used by microorganisms is warranted and possibly overdue. These natural algorithms, if efficient, can be reverse-engineered for graph and tree search strategies.

Holliday junction trap shows how cells use recombination and a junction-guardian role of RecQ helicase.

DNA repair by homologous recombination (HR) underpins cell survival and fuels genome instability, cancer, and evolution. However, the main kinds and sources of DNA damage repaired by HR in somatic cells and the roles of important HR proteins remain elusive. We present engineered proteins that trap, map, and quantify Holliday junctions (HJs), a central DNA intermediate in HR, based on catalytically deficient mutant RuvC protein of Escherichia coli. We use RuvCDefGFP (RDG) to map genomic footprints of HR at defined DNA breaks in E. coli and demonstrate genome-scale directionality of double-strand break (DSB) repair along the chromosome. Unexpectedly, most spontaneous HR-HJ foci are instigated, not by DSBs, but rather by single-stranded DNA damage generated by replication. We show that RecQ, the E. coli ortholog of five human cancer proteins, nonredundantly promotes HR-HJ formation in single cells and, in a novel junction-guardian role, also prevents apparent non-HR-HJs promoted by RecA overproduction. We propose that one or more human RecQ orthologs may act similarly in human cancers overexpressing the RecA ortholog RAD51 and find that cancer genome expression data implicate the orthologs BLM and RECQL4 in conjunction with EME1 and GEN1 as probable HJ reducers in such cancers. Our results support RecA-overproducing E. coli as a model of the many human tumors with up-regulated RAD51 and provide the first glimpses of important, previously elusive reaction intermediates in DNA replication and repair in single living cells.

An ultra-dense library resource for rapid deconvolution of mutations that cause phenotypes in Escherichia coli.

With the wide availability of whole-genome sequencing (WGS), genetic mapping has become the rate-limiting step, inhibiting unbiased forward genetics in even the most tractable model organisms. We introduce a rapid deconvolution resource and method for untagged causative mutations after mutagenesis, screens, and WGS in Escherichia coli. We created Deconvoluter-ordered libraries with selectable insertions every 50 kb in the E. coli genome. The Deconvoluter method uses these for replacement of untagged mutations in the genome using a phage-P1-based gene-replacement strategy. We validate the Deconvoluter resource by deconvolution of 17 of 17 phenotype-altering mutations from a screen of N-ethyl-N-nitrosourea-induced mutants. The Deconvoluter resource permits rapid unbiased screens and gene/function identification and will enable exploration of functions of essential genes and undiscovered genes/sites/alleles not represented in existing deletion collections. This resource for unbiased forward-genetic screens with mapping-by-sequencing ('forward genomics') demonstrates a strategy that could similarly enable rapid screens in many other microbes.

HDR Brachytherapy Dose Distribution is Influenced by the Metal Material of the Applicator.

Applicators containing metal have been widely used in recent years when applying brachytherapy to patients with cervical cancer. However, the high dose rate (HDR) treatment-planning system (TPS) that is currently used in brachytherapy still assumes that the treatment environment constitutes a homogeneous water medium and does not include a dose correction for the metal material of the applicator. The primary purpose of this study was to evaluate the HDR (192)Ir dose distribution in cervical cancer patients when performing brachytherapy using a metal-containing applicator. Thermoluminescent dosimeter (TLD) measurements and Monte Carlo N-Particle eXtended (MCNPX) code were used to explore the doses to the rectum and bladder when using a Henschke applicator containing metal during brachytherapy. When the applicator was assumed to be present, the absolute dose difference between the TLD measurement and MCNPX simulation values was within approximately 5%. A comparison of the MCNPX simulation and TPS calculation values revealed that the TPS overestimated the International Commission of Radiation Units and Measurement (ICRU) rectum and bladder reference doses by 57.78% and 49.59%, respectively. We therefore suggest that the TPS should be modified to account for the shielding effects of the applicator to ensure the accuracy of the delivered doses.