It has been proposed that guanine-rich DNA forms four-stranded structures in vivo called G-quadruplexes or G4 DNA. G4 DNA has been implicated in several biological processes, but tools to study G4 ...DNA structures in cells are limited. Here we report the development of novel murine monoclonal antibodies specific for different G4 DNA structures. We show that one of these antibodies designated 1H6 exhibits strong nuclear staining in most human and murine cells. Staining intensity increased on treatment of cells with agents that stabilize G4 DNA and, strikingly, cells deficient in FANCJ, a G4 DNA-specific helicase, showed stronger nuclear staining than controls. Our data strongly support the existence of G4 DNA structures in mammalian cells and indicate that the abundance of such structures is increased in the absence of FANCJ. We conclude that monoclonal antibody 1H6 is a valuable tool for further studies on the role of G4 DNA in cell and molecular biology.
We report the use of DNA origami nanostructures, functionalized with aptamers, as a vehicle for delivering the antibacterial enzyme lysozyme in a specific and efficient manner. We test the system ...against Gram‐positive (Bacillus subtilis) and Gram‐negative (Escherichia coli) targets. We use direct stochastic optical reconstruction microscopy (dSTORM) and atomic force microscopy (AFM) to characterize the DNA origami nanostructures and structured illumination microscopy (SIM) to assess the binding of the origami to the bacteria. We show that treatment with lysozyme‐functionalized origami slows bacterial growth more effectively than treatment with free lysozyme. Our study introduces DNA origami as a tool in the fight against antibiotic resistance, and our results demonstrate the specificity and efficiency of the nanostructure as a drug delivery vehicle.
Antibiotic resistance is a growing health issue that is now rendering humans vulnerable once again to infections that have been treatable for decades. Various approaches have been proposed to overcome this threat and effectively treat bacterial infections. DNA nanostructures, functionalized with aptamers, were used as a vehicle for delivering the antibacterial enzyme lysozyme in a specific and efficient manner, to destroy bacterial targets.
Background The purpose of this article is to describe the background, design, and patient populations of the Patient Safety in Surgery Study, as a preliminary to the articles in this journal that ...will report the results of the Study. Study Design The Patient Safety in Surgery Study was a prospective cohort study. Trained nurses collected preoperative risk factors, operative variables, and 30-day postoperative mortality and morbidity outcomes in patients undergoing major general and vascular operations at 128 Veterans Affairs (VA) medical centers and 14 selected university medical centers between October 1, 2001 and September 30, 2004. An Internet-based data collection system was used to input data from the different private medical centers. Semiannual feedback of observed to expected mortality and morbidity ratios was provided to the participating medical centers. Results During the 3-year study, total accrual in general surgery was 145,618 patients, including 68.5% from the VA and 31.5% from the private sector. Accrual in vascular surgery totaled 39,225 patients, including 77.8% from the VA and 22.2% from the private sector. VA patients were older and included a larger proportion of male patients and African Americans and Hispanics. The VA population included more inguinal, umbilical, and ventral hernia repairs, although the private-sector population included more thyroid and parathyroid, appendectomy, and operations for breast cancer. Preoperative comorbidities were similar in the two populations, but the rates of comorbidities were higher in the VA. American Society of Anesthesiologists classification tended to be higher in the VA. Conclusions The National Surgical Quality Improvement Program methodology was successfully implemented in the 14 university medical centers. The data from the study provided the basis for the articles in this issue of the Journal of the American College of Surgeons.
The Veterans Affairs' (VA) National Surgical Quality Improvement Program (NSQIP) has been associated with significant reductions in postoperative morbidity and mortality. We sought to determine if ...NSQIP methods and risk models were applicable to private sector (PS) hospitals and if implementation of the NSQIP in the PS would be associated with reductions in adverse postoperative outcomes.
Data from patients (n = 184,843) undergoing major general or vascular surgery between October 1, 2001, and September 30, 2004, in 128 VA hospitals and 14 academic PS hospitals were used to develop prediction models based on VA patients only, PS patients only, and VA plus PS patients using logistic regression modeling, with measures of patient-related risk as the independent variables and 30-day postoperative morbidity or mortality as the dependent variable.
Nine of the top 10 predictors of postoperative mortality and 7 of the top 10 for postoperative morbidity were the same in the VA and PS models. The ratios of observed to expected mortality and morbidity in the PS hospitals based on a model using PS data only versus VA + PS data were nearly identical (correlation coefficient = 0.98). Outlier status of PS hospitals was concordant in 26 of 28 comparisons. Implementation of the NSQIP in PS hospitals was associated with statistically significant reductions in overall postoperative morbidity (8.7%, P = 0.002), surgical site infections (9.1%, P = 0.02), and renal complications (23.7%, P = 0.004).
The VA NSQIP methods and risk models in general and vascular surgery were fully applicable to PS hospitals. Thirty-day postoperative morbidity in PS hospitals was reduced with the implementation of the NSQIP.
Prior small studies have shown multiple benefits of frequent nocturnal hemodialysis compared to conventional three times per week treatments. To study this further, we randomized 87 patients to three ...times per week conventional hemodialysis or to nocturnal hemodialysis six times per week, all with single-use high-flux dialyzers. The 45 patients in the frequent nocturnal arm had a 1.82-fold higher mean weekly stdKt/Vurea, a 1.74-fold higher average number of treatments per week, and a 2.45-fold higher average weekly treatment time than the 42 patients in the conventional arm. We did not find a significant effect of nocturnal hemodialysis for either of the two coprimary outcomes (death or left ventricular mass (measured by MRI) with a hazard ratio of 0.68, or of death or RAND Physical Health Composite with a hazard ratio of 0.91). Possible explanations for the left ventricular mass result include limited sample size and patient characteristics. Secondary outcomes included cognitive performance, self-reported depression, laboratory markers of nutrition, mineral metabolism and anemia, blood pressure and rates of hospitalization, and vascular access interventions. Patients in the nocturnal arm had improved control of hyperphosphatemia and hypertension, but no significant benefit among the other main secondary outcomes. There was a trend for increased vascular access events in the nocturnal arm. Thus, we were unable to demonstrate a definitive benefit of more frequent nocturnal hemodialysis for either coprimary outcome.
The bacterial SbcC/SbcD DNA repair proteins were identified over a quarter of a century ago. Following the subsequent identification of the homologous Mre11/Rad50 complex in the eukaryotes and ...archaea, it has become clear that this conserved chromosomal processing machinery is central to DNA repair pathways and the maintenance of genomic stability in all forms of life. A number of experimental studies have explored this intriguing genome surveillance machinery, yielding significant insights and providing conceptual advances towards our understanding of how this complex operates to mediate DNA repair. However, the inherent complexity and dynamic nature of this chromosome-manipulating machinery continue to obfuscate experimental interrogations, and details regarding the precise mechanisms that underpin the critical repair events remain unanswered. This review will summarize our current understanding of the dramatic structural changes that occur in Mre11/Rad50 complex to mediate chromosomal tethering and accomplish the associated DNA processing events. In addition, undetermined mechanistic aspects of the DNA enzymatic pathways driven by this vital yet enigmatic chromosomal surveillance and repair apparatus will be discussed. In particular, novel and putative models of DNA damage recognition will be considered and comparisons will be made between the modes of action of the Rad50 protein and other related ATPases of the overarching SMC superfamily.
Background Since the Institute of Medicine patient safety reports, a number of survey-based measures of organizational climate safety factors (OCSFs) have been developed. The goal of this study was ...to measure the impact of OCSFs on risk-adjusted surgical morbidity and mortality. Study Design Surveys were administered to staff on general/vascular surgery services during a year. Surveys included multiitem scales measuring OCSFs. Additionally, perceived levels of communication and collaboration with coworkers were assessed. The National Surgical Quality Improvement Program was used to assess risk-adjusted morbidity and mortality. Correlations between outcomes and OCSFs were calculated and between outcomes and communication/collaboration with attending and resident doctors, nurses, and other providers. Results Fifty-two sites participated in the survey: 44 Veterans Affairs and 8 academic medical centers. A total of 6,083 surveys were returned, for a response rate of 52%. The OCSF measures of teamwork climate, safety climate, working conditions, recognition of stress effects, job satisfaction, and burnout demonstrated internal validity but did not correlate with risk-adjusted outcomes. Reported levels of communication/collaboration with attending and resident doctors correlated with risk-adjusted morbidity. Conclusions Survey-based teamwork, safety climate, and working conditions scales are not confirmed to measure organizational factors that influence risk-adjusted surgical outcomes. Reported communication/collaboration with attending and resident doctors on surgical services influenced patient morbidity. This suggests the importance of doctors’ coordination and decision-making roles on surgical teams in providing high-quality and safe care. We propose risk-adjusted morbidity as an effective measure of surgical patient safety.
The spinal cord contains neural circuits that can produce the rhythm and pattern of locomotor activity. It has previously been postulated that a population of glutamatergic neurons, termed Hb9 ...interneurons, contributes to locomotor rhythmogenesis. These neurons were identified by their expression of the homeobox gene, Hb9, which is also expressed in motor neurons. We developed a mouse line in which Cre recombinase activity is inducible in neurons expressing Hb9. We then used this line to eliminate vesicular glutamate transporter 2 from Hb9 interneurons, and found that there were no deficits in treadmill locomotion. We conclude that glutamatergic neurotransmission by Hb9 interneurons is not required for locomotor behaviour. The role of these neurons in neural circuits remains elusive.
Isotropic–nematic (I–N) transitions in cellulose nanocrystal (CNC) suspension and self-assembled structures in the isotropic and nematic phases were investigated using scattering and microscopy ...methods. A CNC suspension with a mass fraction of 7.4% spontaneously phase separated into an isotropic phase of 6.9% in the top layer and a nematic phase of 7.9% in the bottom layer. In both the phases, the CNC particles formed stacks with an interparticle distance being of ≈37 nm. One-dimensional small-angle neutron scattering (SANS) profiles due to both phases could be fitted using a stacking model considering finite particle sizes. SANS and atomic force microscopy studies indicate that the nematic phase in the bottom layer contains more populations of larger particles. A weak magnetic field of ≈0.5 T was able to induce a preferred orientation of CNC stacks in the nematic phase, with the stack normals being aligned with the field (perpendicular to the long axis of CNC particles). The Hermans orientation parameter, ⟨P 2⟩, was ≈0.5 for the nematic phase; it remained unchanged during the relaxation process of ≈10 h. The fraction of oriented CNC populations decreased during the relaxation; dramatic decrease occurred in the first 3 h. The top layer remained isotropic in the weak field. Polarized microscopy studies revealed that the nematic phase was chiral. Adjacent particles in a stack form a twisting angle of ≈0.6 °, resulting in a helix pitch distance of ≈22 μm.
Diblock copolymer thermoplastic elastomers (TPEs) of poly(methyl methacrylate)-block-poly(n-butyl acrylate-random-vinylimidazole) PMMA-b-P(BA-r-VI) have been designed and synthesized as a model ...system for demonstrating simultaneously tailoring photoluminescence (PL) and mechanical properties of TPEs via lanthanide coordination. Europium ions with red PL and terbium ions with green PL have been incorporated in TPEs to coordinate imidazole groups on soft blocks. PL spectra can be continuously tuned by varying both the overall ion composition and their relative portions. The rubber elasticity of TPEs arises from entropic restoring of entangled coil chains in the soft matrix filled with glassy PMMA microdomains as physical cross-links. Upon incorporating lanthanide ions, a second network forms due to the lanthanide coordination cross-linkages among soft P(BA-r-VI) chains. Synergistic interactions and reinforcement of local and global networks greatly enhance both the tensile strength and toughness of TPEs without compromising the stretchability and elasticity. In situ small-angle X-ray scattering reveals deformation and relaxation of microstructures upon cyclic uniaxial stretching and recovery. The novel strategy in the design and synthesis of TPEs with tunable optical and mechanical properties enables a wide range of smart materials and technologies.
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