Expression of the essential Kinase PfCDPK1 from Plasmodium falciparum in Toxoplasma gondii facilitates the discovery of novel antimalarial drugs

We have previously shown that genetic disruption of Toxoplasma gondii calcium-dependent protein kinase 3 (TgCDPK3) affects calcium ionophore-induced egress. We examined whether Plasmodium falciparum CDPK1 (PfCDPK1), the closest homolog of TgCDPK3 in the malaria parasite P. falciparum, could complement a TgCDPK3 mutant strain. PfCDPK1 is essential and plays critical roles in merozoite development, motility, and secretion. We show that expression of PfCDPK1 in the TgCDPK3 mutant strain rescues the egress defect. This phenotypic complementation requires the localization of PfCDPK1 to the plasma membrane and kinase activity. Interestingly, PfCDPK1-expressing Toxoplasma becomes more sensitive to egress inhibition by purfalcamine, a potent inhibitor of PfCDPK1 with low activity against TgCDPK3. Based on this result, we tested eight small molecules previously determined to inhibit the kinase activity of recombinant PfCDPK1 for their abilities to inhibit ionophore-induced egress in the PfCDPK1-expressing strain. While two of these chemicals did not inhibit egress, we found that six drugs affected this process selectively in PfCDPK1-expressing Toxoplasma. Using mutant versions of PfCDPK1 and TgCDPK3, we show that the selectivities of dasatinib and PLX-4720 are regulated by the gatekeeper residue in the ATP binding site. Importantly, we have confirmed that the three most potent inhibitors of egress in the PfCDPK1-expressing strain effectively kill P. falciparum. Thus, we have established and validated a recombinant strain of Toxoplasma that can be used as a surrogate for the discovery and analysis of PfCDPK1-specific inhibitors that can be developed as antimalarials

An Integrated Mechanistic Model of Mindfulness-Oriented Recovery Enhancement for Opioid-Exposed Mother–Infant Dyads

A growing body of neurobiological and psychological research sheds light on the mechanisms underlying the development and maintenance of opioid use disorder and its relation to parenting behavior. Perinatal opioid use is associated with risks for women and children, including increased risk of child maltreatment. Drawing from extant data, here we provide an integrated mechanistic model of perinatal opioid use, parenting behavior, infant attachment, and child well-being to inform the development and adaptation of behavioral interventions for opioid-exposed mother–infant dyads. The model posits that recurrent perinatal opioid use may lead to increased stress sensitivity and reward dysregulation for some mothers, resulting in decreased perceived salience of infant cues, disengaged parenting behavior, disrupted infant attachment, and decreased child well-being. We conclude with a discussion of Mindfulness-Oriented Recovery Enhancement as a means of addressing mechanisms undergirding perinatal opioid use, parenting, and attachment, presenting evidence on the efficacy and therapeutic mechanisms of mindfulness. As perinatal opioid use increases in the United States, empirically informed models can be used to guide treatment development research and address this growing concern

Foreground Predictions for the Cosmic Microwave Background Power Spectrum from Measurements of Faint Inverted Radio Sources at 5 GHz

We present measurements of a population of matched radio sources at 1.4 and 5 GHz down to a flux limit of 1.5 mJy in 7 sq. degs. of the NOAO Deep Field South. We find a significant fraction of sources with inverted spectral indices that all have 1.4 GHz fluxes less than 10 mJy, and are therefore too faint to have been detected and included in previous radio source count models that are matched at multiple frequencies. Combined with the matched source population at 1.4 and 5 GHz in 1 sq. deg. in the ATESP survey, we update models for the 5 GHz differential number counts and distributions of spectral indices in 5 GHz flux bins that can be used to estimate the unresolved point source contribution to the cosmic microwave background temperature anisotropies. We find a shallower logarithmic slope in the 5 GHz differential counts than in previously published models for fluxes < 100 mJy as well as larger fractions of inverted spectral indices at these fluxes. Because the Planck flux limit for resolved sources is larger than 100 mJy in all channels, our modified number counts yield at most a 10% change in the predicted Poisson contribution to the Planck temperature power spectrum. For a flux cut of 5 mJy with the South Pole Telescope and a flux cut of 20 mJy with the Atacama Cosmology Telescope we predict a ~30% and ~10% increase, respectively, in the radio source Poisson power in the lowest frequency channels of each experiment relative to that predicted by previous models.Comment: 14 pages, 9 figures, includes ApJ proof correction

MGMT enrichment and second gene co-expression in hematopoietic progenitor cells using separate or dual-gene lentiviral vectors

AbstractThe DNA repair gene O6-methylguanine-DNA methyltransferase (MGMT) allows efficient in vivo enrichment of transduced hematopoietic stem cells (HSC). Thus, linking this selection strategy to therapeutic gene expression offers the potential to reconstitute diseased hematopoietic tissue with gene-corrected cells. However, different dual-gene expression vector strategies are limited by poor expression of one or both transgenes. To evaluate different co-expression strategies in the context of MGMT-mediated HSC enrichment, we compared selection and expression efficacies in cells cotransduced with separate single-gene MGMT and GFP lentivectors to those obtained with dual-gene vectors employing either encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) or foot and mouth disease virus (FMDV) 2A elements for co-expression strategies. Each strategy was evaluated in vitro and in vivo using equivalent multiplicities of infection (MOI) to transduce 5-fluorouracil (5-FU) or Lin−Sca-1+c-kit+ (LSK)-enriched murine bone marrow cells (BMCs). The highest dual-gene expression (MGMT+GFP+) percentages were obtained with the FMDV-2A dual-gene vector, but half of the resulting gene products existed as fusion proteins. Following selection, dual-gene expression percentages in single-gene vector cotransduced and dual-gene vector transduced populations were similar. Equivalent MGMT expression levels were obtained with each strategy, but GFP expression levels derived from the IRES dual-gene vector were significantly lower. In mice, vector-insertion averages were similar among cells enriched after dual-gene vectors and those cotransduced with single-gene vectors. These data demonstrate the limitations and advantages of each strategy in the context of MGMT-mediated selection, and may provide insights into vector design with respect to a particular therapeutic gene or hematologic defect

Transcriptional profiling of the ductus arteriosus: Comparison of rodent microarrays and human RNA sequencing

DA closure is crucial for the transition from fetal to neonatal life. This closure is supported by changes to the DA’s signaling and structural properties that distinguish it from neighboring vessels. Examining transcriptional differences between these vessels is key to identifying genes or pathways responsible for DA closure. Several microarray studies have explored the DA transcriptome in animal models but varied experimental designs have led to conflicting results. Thorough transcriptomic analysis of the human DA has yet to be performed. A clear picture of the DA transcriptome is key to guiding future research endeavors, both to allow more targeted treatments in the clinical setting, and to understand the basic biology of DA function. In this review, we use a cross-species cross-platform analysis to consider all available published rodent microarray data and novel human RNAseq data in order to provide high priority candidate genes for consideration in future DA studies

Bayesian modelling of clusters of galaxies from multi-frequency pointed Sunyaev--Zel'dovich observations

We present a Bayesian approach to modelling galaxy clusters using multi-frequency pointed observations from telescopes that exploit the Sunyaev--Zel'dovich effect. We use the recently developed MultiNest technique (Feroz, Hobson & Bridges, 2008) to explore the high-dimensional parameter spaces and also to calculate the Bayesian evidence. This permits robust parameter estimation as well as model comparison. Tests on simulated Arcminute Microkelvin Imager observations of a cluster, in the presence of primary CMB signal, radio point sources (detected as well as an unresolved background) and receiver noise, show that our algorithm is able to analyse jointly the data from six frequency channels, sample the posterior space of the model and calculate the Bayesian evidence very efficiently on a single processor. We also illustrate the robustness of our detection process by applying it to a field with radio sources and primordial CMB but no cluster, and show that indeed no cluster is identified. The extension of our methodology to the detection and modelling of multiple clusters in multi-frequency SZ survey data will be described in a future work.Comment: 12 pages, 7 figures, submitted to MNRA

Mindfulness-Oriented Recovery Enhancement vs Supportive Group Therapy for Co-occurring Opioid Misuse and Chronic Pain in Primary Care: A Randomized Clinical Trial

Importance: Successful treatment of opioid misuse among people with chronic pain has proven elusive. Guidelines recommend nonopioid therapies, but the efficacy of mindfulness-based interventions for opioid misuse is uncertain. Objective: To evaluate the efficacy of Mindfulness-Oriented Recovery Enhancement (MORE) for the reduction of opioid misuse and chronic pain. Design, Setting, and Participants: This interviewer-blinded randomized clinical trial enrolled patients from primary care clinics in Utah between January 4, 2016, and January 16, 2020. The study included 250 adults with chronic pain receiving long-term opioid therapy who were misusing opioid medications. Interventions: Treatment with MORE (comprising training in mindfulness, reappraisal, and savoring positive experiences) or supportive group psychotherapy (control condition) across 8 weekly 2-hour group sessions. Main Outcomes and Measures: Primary outcomes were (1) opioid misuse assessed by the Drug Misuse Index (self-report, interview, and urine screen) and (2) pain severity and pain-related functional interference, assessed by subscale scores on the Brief Pain Inventory through 9 months of follow-up. Secondary outcomes were opioid dose, emotional distress, and ecological momentary assessments of opioid craving. The minimum intervention dose was defined as 4 or more completed sessions of MORE or supportive group psychotherapy. Results: Among 250 participants (159 women [63.6%]; mean [SD] age, 51.8 [11.9] years), 129 were randomized to the MORE group and 121 to the supportive psychotherapy group. Overall, 17 participants (6.8%) were Hispanic or Latino, 218 (87.2%) were White, and 15 (6.0%) were of other races and/or ethnicities (2 American Indian, 3 Asian, 1 Black, 2 Pacific Islander, and 7 did not specify). At baseline, the mean duration of pain was 14.7 years (range, 1-60 years), and the mean (SD) morphine-equivalent opioid dose was 101.0 (266.3) mg (IQR, 16.0-90.0 mg). A total of 203 participants (81.2%) received the minimum intervention dose (mean [SD], 5.7 [2.2] sessions); at 9 months, 92 of 250 participants (36.8%) discontinued the study. The overall odds ratio for reduction in opioid misuse through the 9-month follow-up period in the MORE group compared with the supportive psychotherapy group was 2.06 (95% CI, 1.17-3.61; P = .01). At 9 months, 36 of 80 participants (45.0%) in the MORE group were no longer misusing opioids compared with 19 of 78 participants (24.4%) in the supportive psychotherapy group. Mixed models demonstrated that MORE was superior to supportive psychotherapy through 9 months of follow-up for pain severity (between-group effect: 0.49; 95% CI, 0.17-0.81; P = .003) and pain-related functional interference (between-group effect: 1.07; 95% CI, 0.64-1.50; P < .001). Participants in the MORE group reduced their opioid dose to a greater extent than those in the supportive psychotherapy group. The MORE group also had lower emotional distress and opioid craving. Conclusions and Relevance: In this randomized clinical trial, among adult participants in a primary care setting, the MORE intervention led to sustained improvements in opioid misuse and chronic pain symptoms and reductions in opioid dosing, emotional distress, and opioid craving compared with supportive group psychotherapy. Despite attrition caused by the COVID-19 pandemic and the vulnerability of the sample, MORE appeared to be efficacious for reducing opioid misuse among adults with chronic pain. Trial Registration: ClinicalTrials.gov Identifier: NCT0260253

Endoleaks after endovascular graft treatment of aortic aneurysms: Classification, risk factors, and outcome

AbstractPurpose: Incomplete endovascular graft exclusion of an abdominal aortic aneurysm results in an endoleak. To better understand the pathogenesis, significance, and fate of endoleaks, we analyzed our experience with endovascular aneurysm repair. Methods:Between November 1992 and May 1997, 47 aneurysms were treated. In a phase I study, patients received either an endovascular aortoaortic graft (11) or an aortoiliac, femorofemoral graft (8). In phase II, procedures and grafts were modified to include aortofemoral, femorofemoral grafts (28) that were inserted with juxtarenal proximal stents, sutured endovascular distal anastomoses within the femoral artery, and hypogastric artery coil embolization. Endoleaks were detected by arteriogram, computed tomographic scan, or duplex ultrasound. Classification systems to describe anatomic, chronologic, and physiologic endoleak features were developed, and aortic characteristics were correlated with endoleak incidence. Results: Endoleaks were discovered in 11 phase I patients (58%) and only six phase II patients (21%; p < 0.05). Aneurysm neck lengths 2 cm or less increased the incidence of endoleaks (p < 0.05). Although not significant, aneurysms with patent side branches or severe neck calcification had a higher rate of endoleaks than those without these features (47% vs 29% and 57% vs 33%, respectively), and patients with iliac artery occlusive disease had a lower rate of endoleaks than those without occlusive disease (18% vs 42%). Endoleak classifications revealed that most endoleaks were immediate, without outflow, and persistent (71% each), proximal (59%), and had aortic inflow (88%). One patient with a persistent endoleak had aneurysm rupture and died. Conclusions: Endoleaks complicate a significant number of endovascular abdominal aortic aneurysm repairs and may permit aneurysm growth and rupture. The type of graft used, the technique of graft insertion, and aortic anatomic features all affect the rate of endoleaks. Anatomic, chronologic, and physiologic classifications can facilitate endoleak reporting and improve understanding of their pathogenesis, significance, and fate. (J Vasc Surg 1998;27:69-80.

Learning and Reflection in the Midst of Persistent Challenges on Practicing School Leaders’ Time

Among the many scarce resources that principals must conserve and use wisely, time may present one of the more persistent challenges to their leadership practices. A simultaneous and equally challenging practice for school leaders is engaging in systematic reflection that serves to mitigate time constraints and emotional upheavals of the job. Reflection, and resistance to it, emerged as a theme during a yearlong-program that emerged from a school-university partnership, focused on the development of district-level coaches to support experienced principals in becoming more reflective leaders for continuous school improvement. The program uncovered principals’ resistance to the pauses in their practices that reflection requires. These principals and coaches also reported emotional reactions to persisting mandates on school accountability. This paper offers some insights into how reflection may balance the ongoing emotional dynamics and time constraints of schooling, and the degree to which the mentoring program supported principal protégés in this effort