Sex Differences in Left Bundle Branch Area Pacing Versus Biventricular Pacing for Cardiac Resynchronization Therapy.

BACKGROUND: Women respond more favorably to biventricular pacing (BIVP) than men. Sex differences in atrioventricular and interventricular conduction have been described in BIVP studies. Left bundle branch area pacing (LBBAP) offers advantages due to direct capture of the conduction system. We hypothesized that men could respond better to LBBAP than BIVP. OBJECTIVES: This study aims to describe the sex differences in response to LBBAP vs BIVP as the initial cardiac resynchronization therapy (CRT). METHODS: In this multicenter prospective registry, we included patients with left ventricular ejection fraction ≤35% and left bundle branch block or a left ventricular ejection fraction ≤40% with an expected right ventricular pacing exceeding 40% undergoing initial CRT with LBBAP or BIVP. The composite primary outcome was heart failure-related hospitalization and all-cause mortality. The primary safety outcome included all procedure-related complications. RESULTS: There was no significant difference in the primary outcome when comparing men and women receiving LBBAP (P = 0.46), whereas the primary outcome was less frequent in women in the BIVP group than men treated with BIVP (P = 0.03). The primary outcome occurred less frequently in men undergoing LBBAP (29.9%) compared to those treated with BIVP (46.5%) (P = 0.004). In women, the incidence of the primary endpoint was 24.14% in the LBBAP group and 36.2% in the BIVP group; however, this difference was not statistically significant (P = 0.23). Complication rates remained consistent across all groups. CONCLUSIONS: Men and women undergoing LBBAP for CRT had similar clinical outcomes. Men undergoing LBBAP showed a lower risk of heart failure-related hospitalizations and all-cause mortality compared to men undergoing BIVP, whereas there was no difference between LBBAP and BIVP in women.

UDP-6-glucose dehydrogenase in hormonally responsive breast cancers.

Survival for metastatic breast cancer is low and thus, continued efforts to treat and prevent metastatic progression are critical. Estrogen is shown to promote aggressive phenotypes in multiple cancer models irrespective of estrogen receptor (ER) status. Similarly, UDP-Glucose 6-dehydrogenase (UGDH) a ubiquitously expressed enzyme involved in extracellular matrix precursors, as well as hormone processing increases migratory and invasive properties in cancer models. While the role of UGDH in cellular migration is defined, how it intersects with and impacts hormone signaling pathways associated with tumor progression in metastatic breast cancer has not been explored. Here we demonstrate that UGDH knockdown blunts estrogen-induced tumorigenic phenotypes (migration and colony formation) in ER+ and ER- breast cancer in vitro. Knockdown of UGDH also inhibits extravasation of ER- breast cancer ex vivo, primary tumor growth and animal survival in vivo in both ER+ and ER- breast cancer. We also use single cell RNA-sequencing to demonstrate that our findings translate to a human breast cancer clinical specimen. Our findings support the role of estrogen and UGDH in breast cancer progression provide a foundation for future studies to evaluate the role of UGDH in therapeutic resistance to improve outcomes and survival for breast cancer patients.

Multispectral and thermal infrared data, visual scores for severity of common rust symptoms, and genotypic single nucleotide polymorphism data of three F2-derived biparental doubled-haploid maize populations.

Three F2-derived biparental doubled haploid (DH) maize populations were generated for genetic mapping of resistance to common rust. Each of the three populations has the same susceptible parent, but a different resistance donor parent. Population 1 and 3 consist of 320 lines each, population 2 consists of 260 lines. The DH lines were evaluated for their susceptibility to common rust in two years and with two replications in each year. For phenotyping, a visual score (VS) for susceptibility was assigned. Additionally, unmanned aerial vehicle (UAV) derived multispectral and thermal infrared data was recorded and combined in different vegetation indices ("remote sensing", RS). The DH lines were genotyped with the DarTseq method, to obtain data on single nucleotide polymorphisms (SNPs). After quality control, 9051 markers remained. Missing values were "imputed" by the empirical mean of the marker scores of the respective locus. We used the data for comparison of genome-wide association studies and genomic prediction when based on different phenotyping methods, that is either VS or RS data. The data may be interesting for reuse for instance for benchmarking genomic prediction models, for phytopathological studies addressing common rust, or for specifications of vegetation indices.

Effect of Ambrosia arborescens Mill. ethanolic extract on breast cancer induced in rats.

Background and Aims: Ambrosia arborescens Mill. (A. arborescens) is an aromatic plant used in traditional medicine as an anti-inflammatory, anti-tussive, anti-rheumatic, and anti-diarrheal agent. This study aimed to evaluate the effect of A. arborescens Mill. on a Rattus norvegicus var. albinus-induced breast cancer model. Materials and Methods: We collected A. arborescens from the province of Julcán, La Libertad Region, Per, and prepared an ethanolic extract using pulverized leaves macerated in 96° ethanol for 72 h with magnetic stirring. In the evaluation of anticancer activity, four experimental groups with 10 female rats each were formed: Group I (Control-7,12-dimethylbenz[a]anthracene [DMBA]), which received DMBA (single dose) and physiological saline solution for 4 months, and Groups II, III, and IV, which received DMBA (single dose) and 200, 400, and 600 mg/kg/day of the ethanolic extract of A. arborescens, respectively, for 4 months. Results: The DMBA control group presented histological characteristics of ductal carcinoma in situ with necrotic and inflammatory areas, whereas the A. arborescens extract group showed a decrease in tumor volume and recovery of the ductal duct. Conclusion: Ethanol extract of A. arborescens leaves decreases tumor development in rats with induced breast cancer, and this effect is dose-dependent.

Use of remote sensing for linkage mapping and genomic prediction for common rust resistance in maize.

Breeding for disease resistance is a central component of strategies implemented to mitigate biotic stress impacts on crop yield. Conventionally, genotypes of a plant population are evaluated through a labor-intensive process of assigning visual scores (VS) of susceptibility (or resistance) by specifically trained staff, which limits manageable volumes and repeatability of evaluation trials. Remote sensing (RS) tools have the potential to streamline phenotyping processes and to deliver more standardized results at higher through-put. Here, we use a two-year evaluation trial of three newly developed biparental populations of maize doubled haploid lines (DH) to compare the results of genomic analyses of resistance to common rust (CR) when phenotyping is either based on conventional VS or on RS-derived (vegetation) indices. As a general observation, for each population × year combination, the broad sense heritability of VS was greater than or very close to the maximum heritability across all RS indices. Moreover, results of linkage mapping as well as of genomic prediction (GP), suggest that VS data was of a higher quality, indicated by higher -logp values in the linkage studies and higher predictive abilities for genomic prediction. Nevertheless, despite the qualitative differences between the phenotyping methods, each successfully identified the same genomic region on chromosome 10 as being associated with disease resistance. This region is likely related to the known CR resistance locus Rp1. Our results indicate that RS technology can be used to streamline genetic evaluation processes for foliar disease resistance in maize. In particular, RS can potentially reduce costs of phenotypic evaluations and increase trialing capacities.

Single-cell RNA sequencing comparison of the human metastatic prostate spine tumor microenvironment.

Spinal column tumors can be difficult to process for single-cell omic studies, given the heterogeneity in tissue. Here, we present a protocol for operating room-to-benchtop single-cell processing of clinical specimens from a prostate cancer patient. We describe steps for sample homogenization, red blood cell lysis, cryopreservation, and single-cell sequencing analysis. This protocol can be used to identify prognostic markers and therapeutic targets for patients with osseous spine metastases and better inform eligibility for clinical trials.

Attosecond-pump attosecond-probe x-ray spectroscopy of liquid water.

Attosecond-pump/attosecond-probe experiments have long been sought as the most straightforward method for observing electron dynamics in real time. Although there has been much success with overlapped near-infrared femtosecond and extreme ultraviolet attosecond pulses combined with theory, true attosecond-pump/attosecond-probe experiments have been limited. We used a synchronized attosecond x-ray pulse pair from an x-ray free-electron laser to study the electronic response to valence ionization in liquid water through all x-ray attosecond transient absorption spectroscopy (AX-ATAS). Our analysis showed that the AX-ATAS response is confined to the subfemtosecond timescale, eliminating any hydrogen atom motion and demonstrating experimentally that the 1b1 splitting in the x-ray emission spectrum is related to dynamics and is not evidence of two structural motifs in ambient liquid water.

Impact of intracardiac echocardiography versus transesophageal echocardiography guidance on left atrial appendage occlusion procedures: A meta-analysis.

BACKGROUND: Intracardiac echocardiography (ICE) is increasingly used during left atrial appendage occlusion (LAAO) as an alternative to transesophageal echocardiography (TEE). The objective of this study is to evaluate the impact of ICE versus TEE guidance during LAAO on procedural characteristics and acute outcomes, as well the presence of peri-device leaks and residual septal defects during follow-up. METHODS: All studies comparing ICE-guided versus TEE-guided LAAO were identified. The primary outcomes were procedural efficacy and occurrence of procedure-related complications. Secondary outcomes included lab efficiency (defined as a reduction in in-room time), procedural time, fluoroscopy time, and presence of peri-device leaks and residual interatrial septal defects (IASD) during follow-up. RESULTS: Twelve studies (n = 5637) were included. There were no differences in procedural success (98.3% vs. 97.8%; OR 0.73, 95% CI 0.42-1.27, p = .27; I2 = 0%) or adverse events (4.5% vs. 4.4%; OR 0.81 95% CI 0.56-1.16, p = .25; I2 = 0%) between the ICE-guided and TEE-guided groups. ICE guidance reduced in in-room time (mean-weighted 28.6-min reduction in in-room time) without differences in procedural time or fluoroscopy time. There were no differences in peri-device leak (OR 0.93, 95% CI 0.68-1.27, p = 0.64); however, an increased prevalence of residual IASD was observed with ICE-guided versus TEE-guided LAAO (46.3% vs. 34.2%; OR 2.23, 95% CI 1.05-4.75, p = 0.04). CONCLUSION: ICE guidance is associated with similar procedural efficacy and safety, but could result in improved lab efficiency (as established by a significant reduction in in-room time). No differences in the rate of periprocedural leaks were found. A higher prevalence of residual interatrial septal defects was observed with ICE guidance.

Gallbladder plication as a rare complication of endoscopic sleeve gastroplasty: A case report.

BACKGROUND: Endoscopic sleeve gastroplasty (ESG) is a minimally invasive procedure used in the treatment of obesity, with a complication rate of less than 2% of cases. There have been only two reported cases worldwide of gallbladder injuries as a major complication of ESG. CASE SUMMARY: We present the case of a 34-year-old patient who developed a complication after ESG. The patient experienced epigastric and right hypochondrium pain 12 h after the procedure, and a positive Murphy's sign was identified on physical examination. Laboratory results showed a leukocyte count of 17 × 103/µL, and computed tomography indicated the presence of free fluid in the pelvic cavity and perihepatic recesses as well as a possible suture in the wall of the Hartmann's pouch toward the anterior surface of the stomach. A diagnostic laparoscopy was performed, revealing plication of the Hartmann's pouch wall to the anterior stomach wall. Laparoscopic cholecystectomy and lavage were carried out. The patient had a stable recovery and was discharged 72 h after surgery, tolerating oral intake. CONCLUSION: Gallbladder plication should be suspected if signs and symptoms consistent with acute cholecystitis occur after ESG.

Influence of plasma kinetic energy during the pulsed laser deposition of borophene films on silicon (100).

Developing borophene films with good structural stability on non-metallic substrates to maximize their potential in photosensitivity, gas detection, photothermia, energy storage, and deformation detection, among others has been challenging in recent years. Herein, we succeeded in the pulsed laser deposition of multilayered borophene films on Si (100) with ß12 or χ3 bonding by tuning the mean kinetic energy in the plasma during the deposition process. Raman and X-ray photoelectron spectroscopies confirm ß12 and χ3 bonding in the films. Borophene films with ß12 bonding were obtained by tuning a high mean kinetic energy in the plasma, while borophene with χ3 bonding required a relatively low mean kinetic energy. Atomic force microscopy (AFM) micrographs revealed a granular and directional growth of the multilayered borophene films following the linear atomic terraces from the (100) silicon substrate. AFM nanofriction was used to access the borophene surfaces and to reveal the pull-off force and friction coefficient of the films where the surface oxide showed a significant contribution. To summarize, we show that it is possible to deposit multilayered borophene thin films with different bondings by tuning the mean kinetic energy during pulsed laser deposition. The characterization of the plasma during borophene deposition accompanies our findings, providing support for the changes in kinetic energy.

UDP-glucose dehydrogenase (UGDH) in clinical oncology and cancer biology.

UDP-glucose-6-dehydrogenase (UGDH) is a cytosolic, hexameric enzyme that converts UDP-glucose to UDP-glucuronic acid (UDP-GlcUA), a key reaction in hormone and xenobiotic metabolism and in the production of extracellular matrix precursors. In this review, we classify UGDH as a molecular indicator of tumor progression in multiple cancer types, describe its involvement in key canonical cancer signaling pathways, and identify methods to inhibit UGDH, its substrates, and its downstream products. As such, we position UGDH as an enzyme to be exploited as a potential prognostication marker in oncology and a therapeutic target in cancer biology.

Genetic variation among elite inbred lines suggests potential to breed for BNI-capacity in maize.

Biological nitrification inhibition (BNI) is a plant function where root systems release antibiotic compounds (BNIs) specifically aimed at suppressing nitrifiers to limit soil-nitrate formation in the root zone. Little is known about BNI-activity in maize (Zea mays L.), the most important food, feed, and energy crop. Two categories of BNIs are released from maize roots; hydrophobic and hydrophilic BNIs, that determine BNI-capacity in root systems. Zeanone is a recently discovered hydrophobic compound with BNI-activity, released from maize roots. The objectives of this study were to understand/quantify the relationship between zeanone activity and hydrophobic BNI-capacity. We assessed genetic variability among 250 CIMMYT maize lines (CMLs) characterized for hydrophobic BNI-capacity and zeanone activity, towards developing genetic markers linked to this trait in maize. CMLs with high BNI-capacity and ability to release zeanone from roots were identified. GWAS was performed using 27,085 SNPs (with unique positions on the B73v.4 reference genome, and false discovery rate = 10), and phenotypic information for BNI-capacity and zeanone production from root systems. Eighteen significant markers were identified; three associated with specific BNI-activity (SBNI), four with BNI-activity per plant (BNIPP), another ten were common between SBNI and BNIPP, and one with zeanone release. Further, 30 annotated genes were associated with the significant SNPs; most of these genes are involved in pathways of "biological process", and one (AMT5) in ammonium regulation in maize roots. Although the inbred lines in this study were not developed for BNI-traits, the identification of markers associated with BNI-capacity suggests the possibility of using these genomic tools in marker-assisted selection to improve hydrophobic BNI-capacity in maize.

Single-Layer Graphene/Germanium Interface Representing a Schottky Junction Studied by Photoelectron Spectroscopy.

We investigated the interfacial electronic structure of the bidimensional interface of single-layer graphene on a germanium substrate. The procedure followed a well-established approach using ultraviolet (UPS) and X-ray (XPS) photoelectron spectroscopy. The direct synthesis of the single-layer graphene on the surface of (110) undoped Ge substrates was conducted via chemical vapor deposition (CVD). The main graphitic properties of the systems were identified, and it was shown that the Ge substrate affected the electronic structure of the single-layer graphene, indicating the electronic coupling between the graphene and the Ge substrate. Furthermore, the relevant features associated with the Schottky contact's nature, the energy level's alignments, and the energy barrier's heights for electron and hole injection were obtained in this work. The results are useful, given the possible integration of single-layer graphene on a Ge substrate with the complementary metal-oxide-semiconductor (CMOS) technology.

Sustained Apnea for Epicardial Access With Right Ventriculography: The SAFER Epicardial Approach.

BACKGROUND: Epicardial access (EA) has emerged as an increasingly important approach for the treatment of ventricular arrhythmias and to perform other interventional cardiology procedures. EA is frequently underutilized because the current approach is challenging and carries a high risk of life-threatening complications. OBJECTIVE: The purpose of this study was to determine the efficacy and safety of the SAFER (Sustained Apnea for Epicardial Access With Right Ventriculography) epicardial approach. METHODS: Consecutive patients who underwent EA with the SAFER technique were included in this multicenter study. The primary efficacy outcome was the successful achievement of EA. The primary safety outcomes included right ventricular (RV) perforation, major hemorrhagic pericardial effusion (HPE), and bleeding requiring surgical intervention. Secondary outcomes included procedural characteristics and any complications. Our results were compared with those from previous studies describing other EA techniques to assess differences in outcomes. RESULTS: A total of 105 patients undergoing EA with the SAFER approach from June 2021 to February 2023 were included. EA was used for ventricular tachycardia ablation in 98 patients (93.4%), left atrial appendage closure in 6 patients (5.7%), and phrenic nerve displacement in 1 patient (0.9%). EA was successful in all subjects (100%). The median time to EA was 7 minutes (IQR: 5-14 minutes). No cases of RV perforation, HPE, or need of surgical intervention were observed in this cohort. Comparing our results with previous studies about EA, the SAFER epicardial approach resulted in a significant reduction in major pericardial bleeding. CONCLUSIONS: The SAFER epicardial approach is a simple, efficient, effective, and low-cost technique easily reproducible across multiple centers. It is associated with lower complication rates than previously reported techniques for EA.

Bladder cancer patients have increased NETosis and impaired DNaseI-mediated NET degradation that can be therapeutically restored in vitro.

Background: Neutrophils, key players of the immune system, also promote tumor development through the formation of neutrophil extracellular traps (NETs) in a process called NETosis. NETs are extracellular networks of DNA, histones and cytoplasmic and granular proteins (calprotectin, myeloperoxidase, elastase, etc.) released by neutrophils upon activation. NETs regulate tumor growth while promoting angiogenesis and invasiveness, and tumor cells also stimulate NETosis. Although NETosis seems to be increased in cancer patients, an increase of NETs in plasma may also be mediated by an impaired degradation by plasma DNaseI, as evidenced in several immunological disorders like lupus nephritis. However, this has never been evidenced in bladder cancer (BC) patients. Herein, we aimed to evaluate the occurrence of increased NETosis in plasma and tumor tissue of BC patients, to ascertain whether it is mediated by a reduced DNaseI activity and degradation, and to in vitro explore novel therapeutic interventions. Methods: We recruited 71 BC patients from whom we obtained a plasma sample before surgery and a formalin-fixed paraffin embedded tumor tissue sample, and 64 age- and sex-matched healthy controls from whom we obtained a plasma sample. We measured NETs markers (cell-free fDNA, calprotectin, nucleosomes and neutrophil elastase) and the DNaseI activity in plasma with specific assays. We also measured NETs markers in BC tissue by immunofluorescence. Finally, we evaluated the ability of BC and control plasma to degrade in vitro-generated NETs, and evaluated the performance of the approved recombinant human DNaseI (rhDNaseI, Dornase alfa, Pulmozyme®, Roche) to restore the NET-degradation ability of plasma. In vitro experiments were performed in triplicate. Statistical analysis was conducted with Graphpad (v.8.0.1). Results: NETosis occurs in BC tissue, more profusely in the muscle-invasive subtype (P<0.01), that with the worst prognosis. Compared to controls, BC patients had increased NETosis and a reduced DNaseI activity in plasma (P<0.0001), which leads to an impairment to degrade NETs (P<0.0001). Remarkably, this can be therapeutically restored with rhDNaseI to the level of healthy controls. Conclusion: To the best of our knowledge, this is the first report demonstrating that BC patients have an increased NETosis systemically and in the tumor microenvironment, in part caused by an impaired DNaseI-mediated NET degradation. Remarkably, this defect can be therapeutically restored in vitro with the approved Dornase alfa, thus Pulmozyme® could become a potential therapeutic tool to locally reduce BC progression.

Sports drinks effect on salivary volume and pH in children after exercise: a randomized clinical study.

BACKGROUND: The aim of this study was to examine the sports' related oral health behavior and the effect of sports drinks on the salivary volume and acidity of 6 to 14 years old children. METHODS: Sixty-eight children with a median age of 8 years old from Waterford Tennis Association camp participated in the study. Each child was randomly assigned to either the control group that consumed water or the experimental group that consumed a sports drink. Salivary volume and acidity were measured for all groups before exercise, right after exercise and after consuming the rehydrating agent. Salivary volume was measured by dripping into a pre-measured (mL) plastic medicine cup while salivary acidity was measured using an electric pH meter with 0.01 sensitivity. RESULTS: Oral health behavior did not differ between the two groups. No statistically significant difference was detected in the salivary volume before and after exercise. A statistically significant increase (P=0.005) was found in the salivary volume before (1.73ml) and after re-hydration (2.92ml) regardless of the drink consumed (P=0.813). Salivary pH increased significantly (P=0.012) before (7.06) and after exercise (7.73), and dropped significantly (P=0.001) after the consumption of the rehydration drink (6.63) among the same group. The pH decrease was greater in the sports drink group (P=0.013). CONCLUSIONS: No difference in the children's salivary volume was found between the two groups. However, consumption of sports drinks reduces significantly salivary pH and thus, water should be the drink of choice for rehydration in children.

Influence of physical fitness of coronary disease patients on vagal reentry and heart rate recovery after exercise with and without fluid replacement.

BACKGROUND: Fluid replacement during exercise has been studied as an important strategy to enhance recovery, however, studies are needed to investigate the effect of this strategy in different physical profiles. The aim of the study was to investigate the influence of physical fitness of coronary artery disease (CAD) patients on vagal reentry and heart rate recovery after exercise performed with and without fluid replacement. METHODS: Nonrandomized crossover clinical trial. Thirty-three CAD patients were submitted to (I) cardiopulmonary exercise test: to divide the sample into lower and higher VO2  peak groups; (II) control protocol (CP): composed of rest, aerobic exercise and passive recovery; (III) hydration protocol (HP): composed of the same activities as the CP, but with water intake during exercise. The recovery was evaluated by vagal reentry and heart rate recovery immediately after exercise. RESULTS: The results did not show significant differences between the higher and lower VO2  peak groups. In addition, the hydration strategy adopted was not able to cause significant changes between control and HPs, regardless of group. However, a time effect was observed, suggesting anticipation of vagal reactivation and heart rate reduction in HP. CONCLUSIONS: Physical fitness did not influence vagal reentry and heart rate recovery in CAD patients after exercise. However, the hydration strategy seems to have anticipated vagal reentry and produced a more efficient reduction in heart rate regardless of the individuals' physical fitness, but these results should be analyzed with caution due to the absence of significant differences between groups and protocols.

Incidental Diagnosis of Metastatic Breast Cancer in a Man With 99m Tc-PSMA SPECT/CT.

ABSTRACT: Breast cancer in men is a rare and unsuspected malignancy. A 48-year-old man begins with disabling low back pain. The CT scan reported a compression fracture in L2 and diffuse skeletal lesions suggestive of metastatic disease. The serum prostate-specific antigen was 6.2 ng/mL. He was referred for SPECT/CT with 99m Tc-EDDA/HYNIC-inhibitor prostate-specific membrane antigen due to clinical suspicion of prostate cancer. SPECT/CT with 99m Tc-EDDA/HYNIC-inhibitor prostate-specific membrane antigen showed a primary lesion in the left breast and multiple bone lesions. Biopsy confirmed infiltrating ductal carcinoma with positive hormone receptors and indeterminate HER2 (human epidermal growth factor receptor 2).

Evaluating Communication Training at AVMA COE-Accredited Institutions and the Need to Consider Diversity within Simulated Client Pools.

The push for competency-based veterinary medical education by accrediting bodies has led to the inclusion of non-technical skills within curricula. Communication, self-awareness, and cultural humility are considered essential for post-graduate success. To facilitate skills development, veterinary educators have incorporated a variety of modalities including lecture, group discussions, virtual and peer-assisted learning, role play, video review of consultations, and simulated clients (SCs). The overarching goal is developing students into self-reflective practitioners through exposure to clinical scenarios that enhance and embody diversity. Decision making about case management is subject to stereotypes, bias, and assumptions. Racial and ethnic disparities reported in health care can adversely impact patient outcomes. This study was conducted to evaluate communication training and diversity among SC pools within veterinary colleges. A questionnaire was electronically disseminated to assistant/associate deans and/or directors of curriculum/education at 54 American Veterinary Medical Association Council on Education-accredited or provisionally accredited colleges of veterinary medicine. Twenty-one institutions are represented within the data set. Participating institutions summarized their communication curricula: 18 (85.71%) used SCs. Over 55% of these did not track SC demographic data or social identities; among institutions that did track, SCs were primarily monolingual English-speaking (77%), non-disabled (94.2%), white (90.4%), non-Hispanic/Latinx (98.6%) women (57%) over age 56 (64%). Sixteen institutions agreed with the statement "I do not feel that our SC pool is adequately diverse." Respondents shared that lack of time and capacity for recruitment were barriers to diversifying SC pools and proposed strategies to improve outreach.