Disruption of origin chromatin structure by helicase activation in the absence of DNA replication.

Prior to initiation of DNA replication, the eukaryotic helicase, Mcm2-7, must be activated to unwind DNA at replication start sites in early S phase. To study helicase activation within origin chromatin, we constructed a conditional mutant of the polymerase α subunit Cdc17 (or Pol1) to prevent priming and block replication. Recovery of these cells at permissive conditions resulted in the generation of unreplicated gaps at origins, likely due to helicase activation prior to replication initiation. We used micrococcal nuclease (MNase)-based chromatin occupancy profiling under restrictive conditions to study chromatin dynamics associated with helicase activation. Helicase activation in the absence of DNA replication resulted in the disruption and disorganization of chromatin, which extends up to 1 kb from early, efficient replication origins. The CMG holohelicase complex also moves the same distance out from the origin, producing single-stranded DNA that activates the intra-S-phase checkpoint. Loss of the checkpoint did not regulate the progression and stalling of the CMG complex but rather resulted in the disruption of chromatin at both early and late origins. Finally, we found that the local sequence context regulates helicase progression in the absence of DNA replication, suggesting that the helicase is intrinsically less processive when uncoupled from replication.

Intraoperative consultation of vascular surgeons is increasing at a major American trauma center.

OBJECTIVE: Vascular surgeons are often called to aid other surgical specialties for complex exposure, hemorrhage control, or revascularization. The evolving role of the vascular surgeon in the management of intraoperative emergencies involving trauma patients remains undefined. The primary aims of this study included determining the prevalence of intraoperative vascular consultation in trauma, describing how these interactions have changed over time, and characterizing the outcomes achieved by vascular surgeons in these settings. We hypothesized that growing endovascular capabilities of vascular surgeons have resulted in an increased involvement of vascular surgery faculty in the management of the trauma patient over time. METHODS: A retrospective review of all operative cases at a single level I trauma center where a vascular surgeon was involved, but not listed as the primary surgeon, between 2002 and 2017 was performed. Cases were abstracted using Horizon Surgical Manager, a documentation system used in our operating room to track staff present, the type of case, and use. All elective cases were excluded. RESULTS: Of the 256 patients initially identified, 22 were excluded owing to the elective or joint nature of the procedure, leaving 234 emergent operative vascular consultations. Over the 15-year study period, a 529% increase in the number of vascular surgery consultations was seen, with 65% (n = 152) being intraoperative consultations requiring an immediate response. Trauma surgery (n = 103 [44%]) and orthopedic surgery (n = 94 [40%]) were the most common consulting specialties, with both demonstrating a trend of increasing consultations over time (general surgery, 1400%; orthopedic surgery, 220%). Indications for consultation were extremity malperfusion, hemorrhage, and concern for arterial injury. The average operative time for the vascular component of the procedures was 2.4 hours. Of patients presenting with ischemia, revascularization was successful in 94% (n = 116). Hemorrhage was controlled in 99% (n = 122). In-hospital mortality was relatively low at 7% (n = 17). Overall, despite the increase in intraoperative vascular consultations over time, a concomitant increase in the proportion of procedures done using endovascular techniques was not seen. CONCLUSIONS: Vascular surgeons are essential team members at a level I trauma center. Vascular consultation in this setting is often unplanned and often requires immediate intervention. The number of intraoperative vascular consultations is increasing and cannot be attributed solely to an increase in endovascular hemorrhage control, and instead may reflect the declining experience of trauma surgeons with vascular trauma. When consulted, vascular surgeons are effective in quickly gaining control of the situation to provide exposure, hemorrhage control, or revascularization.

The Impact of a Limb Preservation Service on the Incidence of Major Amputations for All Indications at a Level I Trauma Center.

BACKGROUND: Multidisciplinary limb preservation services (LPS) have improved the care of patients with limb-threatening vascular disease. However, the impact of an LPS on major amputations for nonvascular etiologies is unknown. We sought to characterize the trends in major amputations performed at a level I trauma center following the institution of an LPS. METHODS: A retrospective review of all patients undergoing amputation at a level I trauma center from January 2009 to December 2018 was performed. Patients were divided into 2 cohorts: those undergoing amputation pre-LPS (2009-2013) and post-LPS (2014-2018). Major amputations were defined as any amputation at or proximal to the below-knee level. Indications for amputation included chronic limb-threatening ischemia (CLTI), acute limb ischemia (ALI), trauma, infection, and revision amputations. RESULTS: During the study period, 609 major amputations were performed, 490 pre-LPS and 119 post-LPS, representing a 76% reduction. Reductions were seen for every indication, including trauma (95%), ALI (90%), chronic infection (83%), revision (79%), CLTI (68%), and acute infection (62%). CONCLUSIONS: Although previous work has validated the role of an LPS in advanced vascular disease, its value extends beyond vascular disease alone. The drastic reductions seen in the number of amputations performed for a variety of indications, including trauma and diabetic foot infections, further validate the use of a multidisciplinary LPS.

Prospective, multicenter, randomized, controlled trial evaluating the performance of a novel combination powder vs hemostatic matrix in cardiothoracic operations.

AIM: This trial compared the hemostatic performance of a novel combination powder (CP) to a control hemostatic matrix (HM) in cardiothoracic operations. METHODS: Patients meeting eligibility criteria were enrolled after providing informed consent. Subjects were randomized intraoperatively to receive CP (HEMOBLAST Bellows; Biom'up, France) or HM (FLOSEAL Hemostatic Matrix; Baxter Healthcare Corporation, Hayward, CA). Bleeding was assessed using a clinically validated, quantitative bleeding severity scale. The primary endpoint was total time to hemostasis (TTTH), from the start of device preparation, as an indicator of when a surgeon asks for a surgical hemostat until hemostasis was achieved. TTTH at 3 minutes was utilized for the primary analysis, while TTTH at 5 minutes was considered as a secondary endpoint. RESULTS: A total of 105 subjects were enrolled across four institutions. The primary efficacy endpoint for the superiority of CP relative to HM for success at achieving hemostasis within 3 minutes was met, with 64.2% of the CP group achieving hemostasis compared with 9.6% of the HM group, a difference of 54.54% (37.4%-71.6%; P < .001 for superiority). The secondary efficacy endpoint was also met, with 92.5% of the CP group achieving hemostasis at 5 minutes versus 44.2% in the HM group, a difference of 48.2% (31.1%-65.4%; P < .001 for noninferiority). There were no device-related adverse events. CONCLUSIONS: In this multicenter, randomized, controlled trial, comparison of CP to HM revealed CP superiority and noninferiority for TTTH at 3 and 5 minutes, respectively.

Evaluation of the safety and efficacy of a new hemostatic powder using a quantitative surface bleeding severity scale.

AIMS OF THE STUDY: The safety and efficacy of a hemostatic powder (HP) versus a control agent, absorbable gelatin sponge and thrombin (G + T), were assessed, using a validated, quantitative bleeding severity scale. METHODS: Subjects were randomized to receive HP (256 subjects) or G + T (132 subjects) for treatment of minimal, mild, or moderate bleeding at 20 investigational sites. The primary efficacy endpoint was non-inferiority of HP relative to G + T for success at achieving hemostasis within 6 minutes. Secondary endpoints in rank order included: superiority of HP relative to G + T in mean preparation time; non-inferiority of HP relative to G + T for achieving hemostasis within 3 min; superiority of HP relative to G + T for achieving hemostasis within 6 min; and superiority of HP relative to G + T for success for achieving hemostasis within 3 min. RESULTS: A total of 388 subjects were included in the primary efficacy analysis. At 6 min, hemostasis was achieved in 93.0% (238/256) of the HP group compared to 77.3% (102/132) of the G + T group (non-inferiority P < 0.0001, superiority P < 0.0001). All secondary endpoints were met. Complications were comparable between treatment groups. CONCLUSIONS: HP had superior rates of hemostasis, shorter preparation time, and a similar safety profile compared to G + T in this prospective, randomized trial using quantitative bleeding severity criteria.

Wound healing outcomes: Using big data and a modified intent-to-treat method as a metric for reporting healing rates.

Chronic wounds are increasing in prevalence and are a costly problem for the US healthcare system and throughout the world. Typically outcomes studies in the field of wound care have been limited to small clinical trials, comparative effectiveness cohorts and attempts to extrapolate results from claims databases. As a result, outcomes in real world clinical settings may differ from these published studies. This study presents a modified intent-to-treat framework for measuring wound outcomes and measures the consistency of population based outcomes across two distinct settings. In this retrospective observational analysis, we describe the largest to date, cohort of patient wound outcomes derived from 626 hospital based clinics and one academic tertiary care clinic. We present the results of a modified intent-to-treat analysis of wound outcomes as well as demographic and descriptive data. After applying the exclusion criteria, the final analytic sample includes the outcomes from 667,291 wounds in the national sample and 1,788 wounds in the academic sample. We found a consistent modified intent to treat healing rate of 74.6% from the 626 clinics and 77.6% in the academic center. We recommend that a standard modified intent to treat healing rate be used to report wound outcomes to allow for consistency and comparability in measurement across providers, payers and healthcare systems.

Exploratory Analysis of Clinical Predictors of Outcomes of Nonsurgical Treatment in Patients With Lumbar Spinal Stenosis.

OBJECTIVE: The purpose of this study was to explore potential baseline physical examination and demographic predictors of clinical outcomes in patients with lumbar spinal stenosis. METHODS: This was a secondary analysis of data obtained from a pilot randomized controlled trial. Primary and secondary outcome measures were the Swiss Spinal Stenosis (SSS) Questionnaire and visual analog scale (VAS) for leg pain. Multiple regression models were used to assess 2 different outcomes: SSS at completion of care and VAS at completion of care. Separate regression models were built for each of the 2 outcomes to identify the best subset of variables that predicted improvement. Predictors with a significant contribution were retained in a final "best" model. RESULTS: Three variables were identified as having an association with SSS score at completion of care: baseline SSS score, qualitative description of leg pain, and age (adjusted R(2) = 33.2). Four variables were identified as having an association with VAS score at completion of care: baseline VAS score, qualitative description of leg pain, body mass index, and age (adjusted R(2) = 38.3). CONCLUSION: This study provides preliminary evidence supporting an association between certain baseline characteristics and nonsurgical clinical outcomes in patients with lumbar spinal stenosis.

Device Prototype for Vaginal Delivery of Extremely Preterm Fetuses in the Breech Presentation.

Vaginal delivery is typically avoided in the extremely preterm breech population due to the concern of entrapment by the cervix of the aftercoming head. A mechanical device concept is presented to enable vaginal delivery by preventing retraction of the cervix against the fetus during delivery. The two-part device was designed to dilate the cervix, prevent prolapse of small fetal parts and maintain sufficient dilation during delivery. The two-part device was designed and manufactured with the following modules: an inflatable saline-filled cervical balloon for dilation and a cervical retractor composed of semirigid beams to stabilize the cervix and maintain adequate dilation. The device was tested using a cervical phantom designed to simulate the compressive force the cervix exerts. The cervical balloon reached a maximum dilation of 8.5 cm, after which there was leakage of saline from the balloon. While this dilation was less than the target goal of 10 cm, the leaking was attributed to prototype manufacturing defects, which could be resolved with further development. The cervical retractor was able to withstand between 1-3 kPa. Although estimates of cervical pressure values can be upward of 30 kPa, there are no in vivo measurements to formally identify the pressure values for patients in preterm labor. This device serves as a viable proof-of-concept for utilizing an inflatable balloon device to prevent cervical retraction in the setting of extremely preterm vaginal breech delivery. Further manufacturing improvements and design changes could improve the device for continued development and testing.

SPOT GRADE II: Clinical Validation of a New Method for Reproducibly Quantifying Surgical Wound Bleeding: Prospective, Multicenter, Multispecialty, Single-Arm Study.

The SPOT GRADE (SG), a Surface Bleeding Severity Scale, is a unique visual method for assessing bleeding severity based on quantitative determinations of blood flow. This study assessed the reliability of the SG scale in a clinical setting and collected initial data on the safety and efficacy of HEMOBLAST Bellows (HB), a hemostatic agent, in abdominal and orthopedic operations. Twenty-seven patients were enrolled across 3 centers and received the investigational device. Bleeding severity and hemostasis were independently assessed by 2 surgical investigators at baseline and at 3, 6, and 10 minutes after application of HB and compared for agreement. The mean paired κ statistic for assignment of SG scores was .7754. The mean paired κ statistics for determining eligibility for participation in the trial based on bleeding severity and the mean paired κ statistics determining the presence of hemostasis were .9301 and .9301, respectively. The proportion of patients achieving hemostasis within 3, 6, and 10 minutes of HB application were 50.0%, 79.2%, and 91.7%, respectively. There were no unanticipated adverse device effects and one possible serious adverse device effect, as determined by the Independent Data Monitoring Committee (IDMC). The reliability of the SG scale was validated in a clinical setting. Initial data on the safety and efficacy of HB in abdominal and orthopedic operations were collected, and there were no concerns raised by the investigators or the IDMC.

The SPOT GRADE: A New Method for Reproducibly Quantifying Surgical Wound Bleeding.

STUDY DESIGN: Benchtop model with prospective surgeon video testing. OBJECTIVE: To create a surface bleeding severity scale, the SPOT GRADE (SG), for quantitative assessment of target bleeding site (TBS) blood loss. This is of particular interest for spinal surgery due to epidural bleeding and an inability to use diathermy and radiofrequency cautery close to nerve roots. SUMMARY OF BACKGROUND DATA: A novel apparatus perfusable at known flow rates and simulating different sized wounds was used to create movies to educate surgeons on specific degrees of bleeding. METHODS: Training (36) and testing (108) videos were created using a benchtop apparatus employing different bleeding severities based on the six-level SG (none, minimal, mild, moderate, severe, and extreme) and TBS sizes (1, 10, and 50 cm). Fourteen surgeons in four specialties (cardiothoracic, abdominal, spine, and orthopedic lower extremity) were trained and tested to evaluate SG characteristics including inter-rater and intrarater reliability. RESULTS: The interclass correlation coefficient was estimated to be 0.89840 (95% confidence interval [CI]: 0.85771, 1), whereas the intraclass correlation coefficient was estimated to be 0.93673 (95% CI: 0.89603, 1). In 98% of cases (95% CI: 0.9736, 0.9927), surgeons correctly identified eligible bleeds for a future clinical trial (scores = 1, 2, or 3) and in 91% of cases (95% CI: 0.8895, 0.9344), surgeons correctly identified noneligible bleeds (scores = 4 or 5). In 98.6% of cases (95% CI: 0.9777, 0.9945), physicians correctly identified true hemostasis (score = 0). Based upon these data the probability of a physician rating a bleed incorrectly as hemostasis (score = 0) is estimated to be 1.51% (95% CI: 0.0061, 0.0363). CONCLUSION: This SG is reproducible and reliable providing a basis for educating surgeons on TBS blood loss. It appears to be a new standard for evaluating wound blood loss. LEVEL OF EVIDENCE: 2.

Music and the brain - design of an MEG compatible piano.

Magnetoencephalography (MEG) neuroimaging has been used to study subjects' responses when listening to music, but research into the effects of playing music has been limited by the lack of MEG compatible instruments that can operate in a magnetically shielded environment without creating electromagnetic interference. This paper describes the design and preliminary testing of an MEG compatible piano keyboard with 25 full size keys that employs a novel 3-state optical encoder design and electronics to provide realistic velocity-controlled volume modulation. This instrument will allow researchers to study musical performance on a finer timescale than fMRI and enable a range of MEG studies.

A Novel Histone Crosstalk Pathway Important for Regulation of UV-Induced DNA Damage Repair in Saccharomyces cerevisiae.

Histone post-translational modifications play vital roles in a variety of nuclear processes, including DNA repair. It has been previously shown that histone H3K79 methylation is important for the cellular response to DNA damage caused by ultraviolet (UV) radiation, with evidence that specific methylation states play distinct roles in UV repair. Here, we report that H3K79 methylation is reduced in response to UV exposure in Saccharomyces cerevisiae This reduction is specific to the dimethylated state, as trimethylation levels are minimally altered by UV exposure. Inhibition of this reduction has a deleterious effect on UV-induced sister chromatid exchange, suggesting that H3K79 dimethylation levels play a regulatory role in UV repair. Further evidence implicates an additional role for H3K79 dimethylation levels in error-free translesion synthesis, but not in UV-induced G1/S checkpoint activation or double-stranded break repair. Additionally, we find that H3K79 dimethylation levels are influenced by acetylatable lysines on the histone H4 N-terminal tail, which are hyperacetylated in response to UV exposure. Preclusion of H4 acetylation prevents UV-induced reduction of H3K79 dimethylation, and similarly has a negative effect on UV-induced sister chromatid exchange. These results point to the existence of a novel histone crosstalk pathway that is important for the regulation of UV-induced DNA damage repair.

Monodisperse, air-stable PbS nanocrystals via precursor stoichiometry control.

Despite their technological importance, lead sulfide (PbS) nanocrystals have lagged behind nanocrystals of cadmium selenide (CdSe) and lead selenide (PbSe) in terms of size and energy homogeneity. Here, we show that the ratio of lead to sulfur precursor available during nucleation is a critical parameter affecting subsequent growth and monodispersity of PbS nanocrystal ensembles. Applying this knowledge, we synthesize highly monodisperse (size dispersity <5%) PbS nanocrystals over a wide range of sizes (exciton energies from 0.70 to 1.25 eV, or 1000-1800 nm) without the use of size-selective precipitations. This degree of monodispersity results in absorption peak half width at half max (HWHM) values as small as 20 meV, indicating an ensemble that is close to the homogeneous limit. Photoluminescence emission is correspondingly narrow and exhibits small Stokes shifts and quantum efficiencies of 30-60%. The nanocrystals readily self-assemble into ordered superlattices and exhibit exceptional air stability over several months.

Training adults with an autism spectrum disorder to conduct discrete-trial training for young children with autism: a pilot study.

We evaluated a behavioral skills training program for adults with autism spectrum disorder and mild or no intellectual disabilities who were interested in learning the skills used by behavior therapists to work with young children with autism and other developmental disabilities. Four adults, aged 21 to 30 years, participated. We trained each individually using verbal and written instructions, modeling, and role play with feedback to teach 2 basic skills to an adult who played the role of a young child with autism. We evaluated generalization of the discrete-trial training skills by having the participant (a) teach the adult confederate 2 additional targets that we had not included in training and (b) teach a new skill to a young child with autism. Results indicated that 3 of the 4 participants rapidly acquired discrete-trial training skills and that these skills generalized to new targets with the confederate adults and to teaching an actual child with autism.