Using Affinity Pulldown Assays to Study Protein-Protein Interactions of Human NEIL1 Glycosylase and the Checkpoint Protein RAD9-RAD1-HUS1 (9-1-1) Complex.

Affinity pulldown is a powerful technique to discover novel interaction partners and verify a predicted physical association between two or more proteins. Pulldown assays capture a target protein fused with an affinity tag and analyze the complexed proteins. Here, we detail methods of pulldown assays for two high-affinity peptide fusion tags, Flag tag (DYKDDDDK) and hexahistidine tag (6xHis), to study protein-protein interactions of human NEIL1 glycosylase and the checkpoint protein complex RAD9-RAD1-HUS1 (9-1-1). We uncover unique interactions between 9-1-1 and NEIL1, which suggest a possible inhibitory role of the disordered, phosphorylated C-terminal region of RAD9 in regulating NEIL1 activity in base excision repair through lack of physical association of 9-1-1 and NEIL1.

Real-World Implementation of Best-Evidence Cancer Distress Management: Truly Comprehensive Cancer Care.

BACKGROUND: Cancer distress management is an evidence-based component of comprehensive cancer care. Group-delivered cognitive behavioral therapy for cancer distress (CBT-C) is the first distress treatment associated with replicated survival advantages in randomized clinical trials. Despite research supporting patient satisfaction, improved outcomes, and reduced costs, CBT-C has not been tested sufficiently within billable clinical settings, profoundly reducing patient access to best-evidence care. This study aimed to adapt and implement manualized CBT-C as a billable clinical service. PATIENTS AND METHODS: A stakeholder-engaged, mixed-methods, hybrid implementation study design was used, and the study was conducted in 3 phases: (1) stakeholder engagement and adaptation of CBT-C delivery, (2) patient and therapist user testing and adaptation of CBT-C content, and (3) implementation of practice-adapted CBT-C as a billable clinical service focused on evaluation of reach, acceptability, and feasibility across stakeholder perspectives. RESULTS: A total of 40 individuals and 7 interdisciplinary group stakeholders collectively identified 7 primary barriers (eg, number of sessions, workflow concerns, patient geographic distance from center) and 9 facilitators (eg, favorable financial model, emergence of oncology champions). CBT-C adaptations made before implementation included expanding eligibility criteria beyond breast cancer, reducing number of sessions to 5 (10 total hours), eliminating and adding content, and revising language and images. During implementation, 252 patients were eligible; 100 (40%) enrolled in CBT-C (99% covered by insurance). The primary reason for declining enrollment was geographic distance. Of enrollees, 60 (60%) consented to research participation (75% women; 92% white). All research participants completed at least 60% of content (6 of 10 hours), with 98% reporting they would recommend CBT-C to family and friends. CONCLUSIONS: CBT-C implementation as a billable clinical service was acceptable and feasible across cancer care stakeholder measures. Future research is needed to replicate acceptability and feasibility results in more diverse patient groups, test effectiveness in clinical settings, and reduce barriers to access via remote delivery platforms.

Virtual Reality Improves Patient Experience and Anxiety During In-office Carpal Tunnel Release.

Background: This study examined how wide- awake local anesthesia no tourniquet (WALANT) surgery in the office versus the standard operating room (OR) impacts patient experience, and the effect wide awake virtual reality (WAVR) has in conjunction with WALANT on patient experience. Methods: This is a patient-reported outcome study of patients undergoing carpal tunnel release by a single surgeon between August 2017 and March 2021. Patients were classified by location; traditional OR versus WALANT in-office. In-office patients were further classified by whether they chose to use WAVR or not. Patients rated overall experience, enjoyability, and anxiety using a Likert scale (1-7). Results: The online survey had a 44.8% response rate. OR patients were twice as likely to report a neutral or negative experience (23% versus 11%, P = 0.03), significantly lower enjoyment scores (44% versus 20%, P = 0.0007)' and higher anxiety (42% versus 26%, P = 0.04) compared with office-based WALANT patients. With the addition of WAVR, office patients reported higher enjoyment than those who did not use WAVR (85% versus 73%, P = 0.05). Patients reporting an anxiety disorder were more likely to choose WAVR when compared with patients without anxiety disorder (73.8% versus 56.4%). When they chose WAVR, they had greater anxiolysis (79% versus 47%, P = 0.01)' and increased enjoyment (90% versus 59%, P = 0.005). Conclusions: This study demonstrates improved patient experience in the office setting, further amplified by WAVR. Preexisting anxiety disorder is a positive predictive variable toward the patients' choice to use WAVR.

A subset of gut leukocytes has telomerase-dependent "hyper-long" telomeres and require telomerase for function in zebrafish.

BACKGROUND: Telomerase, the enzyme capable of elongating telomeres, is usually restricted in human somatic cells, which contributes to progressive telomere shortening with cell-division and ageing. T and B-cells cells are somatic cells that can break this rule and can modulate telomerase expression in a homeostatic manner. Whereas it seems intuitive that an immune cell type that depends on regular proliferation outbursts for function may have evolved to modulate telomerase expression it is less obvious why others may also do so, as has been suggested for macrophages and neutrophils in some chronic inflammation disease settings. The gut has been highlighted as a key modulator of systemic ageing and is a key tissue where inflammation must be carefully controlled to prevent dysfunction. How telomerase may play a role in innate immune subtypes in the context of natural ageing in the gut, however, remains to be determined. RESULTS: Using the zebrafish model, we show that subsets of gut immune cells have telomerase-dependent"hyper-long" telomeres, which we identified as being predominantly macrophages and dendritics (mpeg1.1+ and cd45+mhcII+). Notably, mpeg1.1+ macrophages have much longer telomeres in the gut than in their haematopoietic tissue of origin, suggesting that there is modulation of telomerase in these cells, in the gut. Moreover, we show that a subset of gut mpeg1.1+ cells express telomerase (tert) in young WT zebrafish, but that the relative proportion of these cells decreases with ageing. Importantly, this is accompanied by telomere shortening and DNA damage responses with ageing and a telomerase-dependent decrease in expression of autophagy and immune activation markers. Finally, these telomerase-dependent molecular alterations are accompanied by impaired phagocytosis of E. coli and increased gut permeability in vivo. CONCLUSIONS: Our data show that limiting levels of telomerase lead to alterations in gut immunity, impacting on the ability to clear pathogens in vivo. These are accompanied by increased gut permeability, which, together, are likely contributors to local and systemic tissue degeneration and increased susceptibility to infection with ageing.

Vulnerability of Inexperience: A Qualitative Exploration of Physician Grief and Coping after Impactful Pediatric Patient Deaths.

Background: Caring for dying patients can result in burnout, stress, and emotional trauma for some physicians,1,2 particularly among trainees. Research is lacking that focuses on the emotional impact and coping techniques utilized by novice and experienced pediatricians after impactful pediatric patient deaths. Objectives: To define the salient features of an impactful pediatric patient death and physicians' grief and coping responses. As a secondary aim, we explored the cognitive and emotional training physicians described as helpful or would be helpful when coping after impactful patient deaths. Design: We conducted a prospective qualitative study using semistructured interviews and applied descriptive thematic content analysis to the transcribed interviews. Setting/Subjects: We enrolled pediatric intensive care unit trainees and attendings in a single United States institution over a six-month period from January 2021 to June 2021. Results: Both trainee and attending physicians were most impacted by acute or unexpected patient deaths. Trainees were particularly impacted by their first or early career patient deaths. Both groups found talking about the death of a patient the most helpful coping mechanism. Attending physicians coped with positive reframing, whereas novices more frequently utilized avoidance, numbing, and rumination. The importance of experienced physician's role modeling vulnerability and supporting trainee growth rather than "getting it right" were highlighted as trainee coping gaps. Conclusions: Novice physicians are particularly vulnerable to acute stress after the death of a patient and require additional coping resources and supports. Future projects should explore the impact of teaching emotion-focused coping techniques on trainee resiliency and coping after early career patient deaths.

Prognostic value of coronary computed tomography angiographic derived fractional flow reserve: a systematic review and meta-analysis.

OBJECTIVES: To obtain more powerful assessment of the prognostic value of fractional flow reserveCT testing we performed a systematic literature review and collaborative meta-analysis of studies that assessed clinical outcomes of CT-derived calculation of FFR (FFRCT) (HeartFlow) analysis in patients with stable coronary artery disease (CAD). METHODS: We searched PubMed and Web of Science electronic databases for published studies that evaluated clinical outcomes following fractional flow reserveCT testing between 1 January 2010 and 31 December 2020. The primary endpoint was defined as 'all-cause mortality (ACM) or myocardial infarction (MI)' at 12-month follow-up. Exploratory analyses were performed using major adverse cardiovascular events (MACEs, ACM+MI+unplanned revascularisation), ACM, MI, spontaneous MI or unplanned (>3 months) revascularisation as the endpoint. RESULTS: Five studies were identified including a total of 5460 patients eligible for meta-analyses. The primary endpoint occurred in 60 (1.1%) patients, 0.6% (13/2126) with FFRCT>0.80% and 1.4% (47/3334) with FFRCT ≤0.80 (relative risk (RR) 2.31 (95% CI 1.29 to 4.13), p=0.005). Likewise, MACE, MI, spontaneous MI or unplanned revascularisation occurred more frequently in patients with FFRCT ≤0.80 versus patients with FFRCT >0.80. Each 0.10-unit FFRCT reduction was associated with a greater risk of the primary endpoint (RR 1.67 (95% CI 1.47 to 1.87), p<0.001). CONCLUSIONS: The 12-month outcomes in patients with stable CAD show low rates of events in those with a negative FFRCT result, and lower risk of an unfavourable outcome in patients with a negative test result compared with patients with a positive test result. Moreover, the FFRCT numerical value was inversely associated with outcomes.

Virtual Reality Improves the Patient Experience during Wide-Awake Local Anesthesia No Tourniquet Hand Surgery: A Single-Blind, Randomized, Prospective Study.

Wide-awake local anesthesia no tourniquet surgery has been shown to decrease cost and hospital length of stay. The authors studied the use of virtual reality during wide-awake local anesthesia no tourniquet outpatient upper extremity surgery to assess its effect on patient pain, anxiety and fun. Patients undergoing wide-awake local anesthesia no tourniquet surgery were randomized to use (virtual reality) or not use (non-virtual reality) virtual reality during their procedures. Pain, fun, and anxiety were measured with a Likert scale at several time points, as were blood pressure and heart rate. A postoperative questionnaire was used to assess overall satisfaction. Virtual reality patients exhibited lower anxiety scores during injection, during the procedure, and at the end of the procedure. There were no differences in blood pressure, heart rate, or pain scores. Compared with non-virtual reality patients, virtual reality patients' fun scores were higher. Virtual reality patients felt the experience helped them to relax, and they would recommend virtual reality-assisted wide-awake local anesthesia no tourniquet surgery. Among patients with self-reported preexisting anxiety, virtual reality patients had lower pain and anxiety scores during injection of local anesthesia compared with non-virtual reality patients. This study demonstrates that readily available virtual reality hardware and software can provide a virtual reality experience that reduces patient anxiety both during the injection of local anesthesia and during the surgical procedure. (Plast. Reconstr. Surg. 144: 408, 2019.) CLINICAL QUESTION/LEVEL OF EVIDENCE:: Therapeutic, II.

Validation of CyTOF Against Flow Cytometry for Immunological Studies and Monitoring of Human Cancer Clinical Trials.

Flow cytometry is a widely applied approach for exploratory immune profiling and biomarker discovery in cancer and other diseases. However, flow cytometry is limited by the number of parameters that can be simultaneously analyzed, severely restricting its utility. Recently, the advent of mass cytometry (CyTOF) has enabled high dimensional and unbiased examination of the immune system, allowing simultaneous interrogation of a large number of parameters. This is important for deep interrogation of immune responses and particularly when sample sizes are limited (such as in tumors). Our goal was to compare the accuracy and reproducibility of CyTOF against flow cytometry as a reliable analytic tool for human PBMC and tumor tissues for cancer clinical trials. We developed a 40+ parameter CyTOF panel and demonstrate that compared to flow cytometry, CyTOF yields analogous quantification of cell lineages in conjunction with markers of cell differentiation, function, activation, and exhaustion for use with fresh and viably frozen PBMC or tumor tissues. Further, we provide a protocol that enables reliable quantification by CyTOF down to low numbers of input human cells, an approach that is particularly important when cell numbers are limiting. Thus, we validate CyTOF as an accurate approach to perform high dimensional analysis in human tumor tissue and to utilize low cell numbers for subsequent immunologic studies and cancer clinical trials.

Resident Immunity in Tissue Repair and Maintenance: The Zebrafish Model Coming of Age.

The zebrafish has emerged as an exciting vertebrate model to study different aspects of immune system development, particularly due to its transparent embryonic development, the availability of multiple fluorescent reporter lines, efficient genetic tools and live imaging capabilities. However, the study of immunity in zebrafish has largely been limited to early larval stages due to an incomplete knowledge of the full repertoire of immune cells and their specific markers, in particular, a lack of cell surface antibodies to detect and isolate such cells in living tissues. Here we focus on tissue resident or associated immunity beyond development, in the adult zebrafish. It is our view that, with our increasing knowledge and the development of improved tools and protocols, the adult zebrafish will be increasingly appreciated for offering valuable insights into the role of immunity in tissue repair and maintenance, in both health and disease throughout the lifecourse.

Parathyroid Hormone Senses Extracellular Calcium To Modulate Endocrine Signaling upon Binding to the Family B GPCR Parathyroid Hormone 1 Receptor.

Parathyroid hormone (PTH) binds to a family B G protein coupled receptor, parathyroid hormone 1 receptor (PTH1R). One of its functions is to regulate Ca2+ homeostasis in bone remodeling, during which Ca2+ can reach up to 40 mM. A truncated version of PTH, PTH(1-34), can fully activate PTH1R and has been used for osteoporosis treatments. Here, we used fluorescence anisotropy to examine the binding of PTH(1-34) to PTH1R purified in nanodiscs (PTH1R-ND) and found that the affinity increases 5-fold in the presence of 15 mM Ca2+. However, PTHrP(1-36), another truncated endogenous agonist for PTH1R, does not show this Ca2+ effect. Mutations of Glu19 and Glu22 in PTH(1-34) that are not conserved in PTHrP(1-36) largely abolished the Ca2+ effect. The results support that PTH(1-34) not only activates PTH1R but also uniquely senses Ca2+. This dual function of a peptide hormone is a novel observation that couples changes in extracellular environment with endocrine signaling. Understanding this can potentially reveal the complex role of PTH signaling in bone remodeling and improve the PTH(1-34) treatment for osteoporosis.

Comprehensive Multicenter Graduate Surgical Education Initiative Incorporating Entrustable Professional Activities, Continuous Quality Improvement Cycles, and a Web-Based Platform to Enhance Teaching and Learning.

BACKGROUND: It is increasingly important for faculty to teach deliberately and provide timely, detailed, and formative feedback on surgical trainee performance. We initiated a multicenter study to improve resident evaluative processes and enhance teaching and learning behaviors while engaging residents in their education. STUDY DESIGN: Faculty from 7 US postgraduate training programs rated resident operative performances using the perioperative briefing, intraoperative teaching, debriefing model, and rated patient visits/academic performances using the entrustable professional activities model via a web-based platform. Data were centrally analyzed and iterative changes made based on participant feedback, individual preferences, and database refinements, with trends addressed using the Plan, Do, Check, Act improvement methodology. RESULTS: Participants (92 surgeons, 150 residents) submitted 3,880 assessments during July 2014 through September 2017. Evidence of preoperative briefings improved from 33.9% ± 2.5% to 95.5% ± 1.5% between April and September 2014 compared with April and September 2017 (p < 0.001). Postoperative debriefings improved from 10.6% ± 2.7% to 90.2% ± 2.5% (p < 0.001) for the same period. Meaningful self-reflection by residents improved from 28.6% to 67.4% (p < 0.001). The number of assessments received per resident during a 6-month period increased from 6.4 ± 6.2 to 13.4 ± 10.1 (p < 0.003). Surgeon-entered assessments increased from 364 initially to 685 in the final period, and the number of resident assessments increased from 308 to 445. We showed a 4-fold increase in resident observed activities being rated. CONCLUSIONS: By adopting recognized educational models with repeated Plan, Do, Check, Act cycles, we increased the quality of preoperative learning objectives, showed more frequent, detailed, and timely assessments of resident performance, and demonstrated more effective self-reflection by residents. We monitored trends, identified opportunities for improvement and successfully sustained those improvements over time, applying a team-based approach.

Perioperative self-reflection among surgical residents.

BACKGROUND: We studied prevalence and predictors of meaningful self-reflection among surgical residents and with prompting/structured interventions, sought to improve/sustain resident skills. METHODS: Residents from six programs recorded 1032 narrative self-reflective comments (120 residents), using a web-based platform. If residents identified something learned or to be improved, self-reflection was deemed meaningful. Independent variables PGY level, resident/surgeon gender, study site/Phase1: July2014-August2015 vs. Phase2: September2015-September2016) were analyzed. RESULTS: Meaningful self-reflection was documented in 40.6% (419/1032) of entries. PGY5's meaningfully self-reflected less than PGY1-4's, 26.1% vs. 49.6% (p = 0.002). In multivariate analysis, resident narratives during Phase 2 were 4.7 times more likely to engage in meaningful self-reflection compared to Phase1 entries (p < 0.001). Iterative changes during Phase2 showed a 236% increase in meaningful self-reflection, compared to Phase1. CONCLUSIONS: Surgical residents uncommonly practice meaningful self-reflection, even when prompted, and PGY5/chief residents reflect less than more junior residents. Substantial/sustained improvements in resident self-reflection can occur with both training and interventions.

Do female surgeons learn or teach differently?

BACKGROUND: Gender and/or gender-stereotypes might influence surgical education. We hypothesized that female surgeons might focus their learning and teaching differently from male surgeons. METHODS: Residents and surgeons (multi-institutional) individually recorded preoperatively discussed learning objectives (LO) for matching cases. Narratives were classified as knowledge-based, skill-based, or attitude-based. Multinomial logistic regression analyses, LO = dependent variable; independent variables = resident/surgeon gender, PGY level, timing of entry-to-procedure date, and quarters-of-year. RESULTS: 727 LOs from 125 residents (41% female) and 49 surgeons (20% female) were classified. Female residents were 1.4 times more likely to select knowledge over skill. With female surgeons, residents were 1.6 and 2.1 times more likely to select knowledge over skill and attitude over skill than if the surgeon was male. PGY 4/5 residents chose attitude-based LOs over junior residents. CONCLUSION: Resident, surgeon gender and year-of-training influence learning objectives. Whether this reflects gender stereotyping by residents or differences in attending teaching styles awaits further exploration.

A taxonomy of perioperative surgical learning: Trending resident skill acquisition.

BACKGROUND: Resident and curriculum evaluation require tracking surgical resident operative performance, yet what and when to measure remains unclear. METHODS: From a multi-institutional database, we reviewed 611 resident/surgeon-paired assessments of ACGME Milestones and modified OPRS ratings for different cases and postgraduate years. RESULTS: Faculty Milestone ratings increased with each PGY (p=<0.001) and correlated with resident self-ratings (ICC = 0.83). Mean OPRS scores increased in small increments with substantial intra-year variability. Progression among individual OPRS subcategories was not apparent from more global analyses. Interestingly, male faculty offered lower ratings than female faculty. CONCLUSIONS: Milestones and modified mean OPRS ratings suggest residents are learning, yet lack sufficient discrimination for promotion or curricular analysis. Differential progression through OPRS subcategories suggests a taxonomy of surgical learning that can be tailored to focus on different skills at each point in the training continuum. The effect of faculty gender on resident ratings awaits further study.

ß-Peptide bundles: Design. Build. Analyze. Biosynthesize.

Peptides containing ß-amino acids are unique non-natural polymers known to assemble into protein-like tertiary and quaternary structures. When composed solely of ß-amino acids, the structures formed, defined assemblies of 14-helices called ß-peptide bundles, fold cooperatively in water solvent into unique and discrete quaternary assemblies that are highly thermostable, bind complex substrates and metal ion cofactors, and, in certain cases, catalyze chemical reactions. In this Perspective, we recount the design and elaboration of ß-peptide bundles and provide an outlook on recent, unexpected discoveries that could influence research on ß-peptides and ß-peptide bundles (and ß-amino acid-containing proteins) for decades to come.

The effect of the national shortage of vitamin A on death or chronic lung disease in extremely low-birth-weight infants.

IMPORTANCE: Prophylactic vitamin A supplementation has been shown to reduce the incidence of chronic lung disease or death in extremely low-birth-weight infants. Beginning in 2010, a national shortage reduced the supply of vitamin A available. OBJECTIVE: To estimate the association between vitamin A supplementation and death or chronic lung disease in the context of the recent drug shortage. Intercenter variability in vitamin A use was assessed secondarily. DESIGN, SETTING, AND PARTICIPANTS: Retrospective cohort study of 7925 infants with birth weights between 401 and 1000 g who were cared for in US neonatal intensive care units managed by the Pediatrix Medical Group. Infants were discharged between January 1, 2010, and June 30, 2012, and data were collected from the Pediatrix Clinical Data Warehouse. Infants who had major congenital anomalies, died during the first 3 days of life, or had missing data were excluded from the analysis. EXPOSURES: Vitamin A supplementation. MAIN OUTCOMES AND MEASURES: The primary outcome was either death before hospital discharge or chronic lung disease, defined as receiving any respiratory support at 36 weeks' corrected gestational age. RESULTS: Of the 6210 eligible infants, 3011 (48.5%) experienced the primary outcome. Those who received vitamin A were more immature and more likely to receive mechanical ventilation during the first 3 days of life. During the study period, vitamin A supplementation significantly decreased (27.2% to 2.1%); however, the primary outcome was similar (48.4% to 49.5%; P = .40). Vitamin A was unrelated to death or chronic lung disease in unadjusted or multivariable analyses (relative risk [RR], 0.97; 95% CI, 0.91-1.03; P = .32) when demographic and clinical information were considered. After classifying centers by vitamin A use, the center of birth was significantly associated with the outcome, with birth in low- and medium-use centers related to a reduced likelihood of death or chronic lung disease. CONCLUSIONS AND RELEVANCE: The occurrence of death or chronic lung disease appears unaffected by the recent shortage of vitamin A. However, the center of birth appears to be an important risk factor for these infants' outcomes.

Design and high-resolution structure of a ß³-peptide bundle catalyst.

Despite the widespread exploration of α-peptides as catalysts, there are few examples of ß-peptides that alter the course of a chemical transformation. Our previous work demonstrated that a special class of ß(3)-peptides spontaneously self-assembles in water into discrete protein-like bundles possessing unique quaternary structures and exceptional thermodynamic stability. Here we describe a series of ß(3)-peptide bundles capable of both substrate binding and chemical catalysis--ester hydrolysis. A combination of kinetic and high-resolution structural analysis suggests an active site triad composed of residues from at least two strands of the octameric bundle structure.

Peer involvement in adolescent dating violence.

This study investigated the ways in which peers are involved in adolescent dating violence. Eighty-eight young adults aged 18-21 were interviewed and asked to reflect on aggressive dating relationships they experienced as teens. The researchers used grounded theory to analyze the data. Findings showed that male and female peers were involved in adolescent dating violence in unique ways. Male peers were involved in dating violence by participating in the aggression, agitating the aggression, being the competition, trivializing the aggression, and keeping tabs on the recipient. Female peers were involved in dating violence by deserting the recipient, cheating with the boyfriend, being the audience, needling the male dating partner, and helping the recipient. Male and female peers were involved similarly in adolescent dating violence by confronting the partner. School nurses working with adolescents are uniquely positioned to approach adolescents about dating violence. Interventions aimed at promoting discussions with adolescents are discussed.

Outbreak of carbapenem-resistant enterobacteriaceae at a long-term acute care hospital: sustained reductions in transmission through active surveillance and targeted interventions.

OBJECTIVE: To describe a Klebsiella pneumoniae carbapenemase (KPC)-producing carbapenem-resistant Enterobacteriaceae (CRE) outbreak and interventions to prevent transmission. DESIGN, SETTING, AND PATIENTS: Epidemiologic investigation of a CRE outbreak among patients at a long-term acute care hospital (LTACH). METHODS: Microbiology records at LTACH A from March 2009 through February 2011 were reviewed to identify CRE transmission cases and cases admitted with CRE. CRE bacteremia episodes were identified during March 2009-July 2011. Biweekly CRE prevalence surveys were conducted during July 2010-July 2011, and interventions to prevent transmission were implemented, including education and auditing of staff and isolation and cohorting of CRE patients with dedicated nursing staff and shared medical equipment. Trends were evaluated using weighted linear or Poisson regression. CRE transmission cases were included in a case-control study to evaluate risk factors for acquisition. A real-time polymerase chain reaction assay was used to detect the bla(KPC) gene, and pulsed-field gel electrophoresis was performed to assess the genetic relatedness of isolates. RESULTS: Ninety-nine CRE transmission cases, 16 admission cases (from 7 acute care hospitals), and 29 CRE bacteremia episodes were identified. Significant reductions were observed in CRE prevalence (49% vs. 8%), percentage of patients screened with newly detected CRE (44% vs. 0%), and CRE bacteremia episodes (2.5 vs. 0.0 per 1,000 patient-days). Cases were more likely to have received ß-lactams, have diabetes, and require mechanical ventilation. All tested isolates were KPC-producing K. pneumoniae, and nearly all isolates were genetically related. CONCLUSION: CRE transmission can be reduced in LTACHs through surveillance testing and targeted interventions. Sustainable reductions within and across healthcare facilities may require a regional public health approach.

Female reproductive status and circulating blood leukocyte expression of selected metabolic or signaling genes involved in sex steroid metabolism.

OBJECTIVE: To examine the blood leukocyte expression of 22 sex steroid metabolic/signaling genes according to female reproductive status. METHODS: Michigan Fisheaters' Cohort participants underwent blood collection during the luteal phase of the menstrual cycle or randomly in non-menstruating participants. Gene expression (GE) was measured using Taqman hydrolysis probes and quantitative RT-PCR. Repeatability of four genes was determined in a subgroup. RESULTS: Five premenstrual, 57 premenopausal (20 users of systemic hormonal contraception), and 43 postmenopausal females participated. After Bonferroni correction for multiple comparisons of median GE between groups, three findings remained significant: greater GE of AhR in postmenopausal women than in premenopausal non-users of systemic hormonal contraception; and greater GE of ESR2 and HSD17B7 in premenstrual girls compared to postmenopausal women. Modest intra-class correlations were identified for CYP 19, ESR1, and ESR2 GE measured both in 2007 and 2010, but no intra-class correlation over the same time period was found for CYP17. CONCLUSIONS: There was little differential variation of blood leukocyte sex steroid ge between premenopausal women in the luteal phase of the menstrual cycle and postmenopausal women for most genes analyzed, but it will be necessary to make statistical adjustments in future epidemiologic studies in two circumstances: 1) when comparing AhR GE in premenopausal women non-users of systemic hormone contraception with postmenopausal women and 2) when comparing ESR2 and HSD17B7 GE in studies that include premenstrual girls. Developmental differences may explain the differential GE found in ESR2 and HSD17B7 in premenstrual girls compared with postmenopausal women.