Modeling of Noisy Spindle Dynamics Reveals Separable Contributions to Achieving Correct Orientation

The mechanisms by which the mammalian mitotic spindle is guided to a predefined orientation through microtubule-cortex interactions have recently received considerable interest, but there has been no dynamic model that describes spindle movements toward the preferred axis in human cells. Here, we develop a dynamic model based on stochastic activity of cues anisotropically positioned around the cortex of the mitotic cell and we show that the mitotic spindle does not reach equilibrium before chromosome segregation. Our model successfully captures the characteristic experimental behavior of noisy spindle rotation dynamics in human epithelial cells, including a weak underlying bias in the direction of rotation, suppression of motion close to the alignment axis, and the effect of the aspect ratio of the interphase cell shape in defining the final alignment axis. We predict that the force exerted per cue has a value that minimizes the deviation of the spindle from the predefined axis. The model has allowed us to systematically explore the parameter space around experimentally relevant configurations, and predict the mechanistic function of a number of established regulators of spindle orientation, highlighting how physical modeling of a noisy system can lead to functional biological understanding. We provide key insights into measurable parameters in live cells that can help distinguish between mechanisms of microtubule and cortical-cue interactions that jointly control the final orientation of the spindle.     

Evaluating the Performance of Time-Gated Live-Cell Microscopy with Lanthanide Probes

Probes and biosensors that incorporate luminescent Tb(III) or Eu(III) complexes are promising for cellular imaging because time-gated microscopes can detect their long-lifetime (approximately milliseconds) emission without interference from short-lifetime (approximately nanoseconds) fluorescence background. Moreover, the discrete, narrow emission bands of Tb(III) complexes make them uniquely suited for multiplexed imaging applications because they can serve as Förster resonance energy transfer (FRET) donors to two or more differently colored acceptors. However, lanthanide complexes have low photon emission rates that can limit the image signal/noise ratio, which has a square-root dependence on photon counts. This work describes the performance of a wide-field, time-gated microscope with respect to its ability to image Tb(III) luminescence and Tb(III)-mediated FRET in cultured mammalian cells. The system employed a UV-emitting LED for low-power, pulsed excitation and an intensified CCD camera for gated detection. Exposure times of ∼1 s were needed to collect 5–25 photons per pixel from cells that contained micromolar concentrations of a Tb(III) complex. The observed photon counts matched those predicted by a theoretical model that incorporated the photophysical properties of the Tb(III) probe and the instrument’s light-collection characteristics. Despite low photon counts, images of Tb(III)/green fluorescent protein FRET with a signal/noise ratio ≥ 7 were acquired, and a 90% change in the ratiometric FRET signal was measured. This study shows that the sensitivity and precision of lanthanide-based cellular microscopy can approach that of conventional FRET microscopy with fluorescent proteins. The results should encourage further development of lanthanide biosensors that can measure analyte concentration, enzyme activation, and protein-protein interactions in live cells.     

Stimulation of pulses of 13,14-dihydro-15-keto-PGF2α (PGFM) with estradiol-17β and changes in circulating progesterone concentrations within a PGFM pulse in heifers

A single physiologic dose (0.1 mg) of estradiol-17β in sesame-oil vehicle or vehicle alone (n = 8) was given to heifers on day 14 after ovulation to study the effect on circulating 13-14-dihydro-15-keto-PGF2α (PGFM), PGFM pulses, and changes in progesterone concentrations within a PGFM pulse. Blood samples were collected hourly for 16 h after treatment. The estradiol group had a greater mean concentration of PGFM, greater number of heifers with PGFM pulses and number of pulses/heifer, and greater prominence of the PGFM pulses. Changes in progesterone concentrations were not detected during the 16 h sampling session in the vehicle group, indicating that the heifers were in preluteolysis. Progesterone decreased after 12 h in the estradiol group, indicating a luteolytic effect of the estradiol-induced PGF secretion as represented by PGFM concentrations. Intrapulse changes in progesterone were detected during a PGFM pulse in the estradiol group (P < 0.006), but not in the vehicle group. Progesterone increased (P < 0.01) between Hours −2 and −1 of an estradiol-induced PGFM pulse (Hour 0 = peak of pulse), decreased (P < 0.004) between Hours −1 and 0, and increased (P < 0.01) or rebounded between Hours 0 and 1. Results were compatible with previous reports of a role for estradiol in the induction of PGFM pulses in cattle and demonstrated intrapulse changes in progesterone concentrations during an induced PGFM pulse.     

Effectiveness of Group Problem Management Plus,a brief psychological intervention for adults affected by humanitarian disasters in Nepal: A cluster randomized controlled trial

BackgroundGlobally, 235 million people are impacted by humanitarian emergencies worldwide, presenting increased risk of experiencing a mental disorder. Our objective was to test the effectiveness of a brief group psychological treatment delivered by trained facilitators without prior professional mental health training in a disaster-prone setting.Methods and findingsWe conducted a cluster randomized controlled trial (cRCT) from November 25, 2018 through September 30, 2019. Participants in both arms were assessed at baseline, midline (7 weeks post-baseline, which was approximately 1 week after treatment in the experimental arm), and endline (20 weeks post-baseline, which was approximately 3 months posttreatment). The intervention was Group Problem Management Plus (PM+), a psychological treatment of 5 weekly sessions, which was compared with enhanced usual care (EUC) consisting of a family psychoeducation meeting with a referral option to primary care providers trained in mental healthcare. The setting was 72 wards (geographic unit of clustering) in eastern Nepal, with 1 PM+ group per ward in the treatment arm. Wards were eligible if they were in disaster-prone regions and residents spoke Nepali. Wards were assigned to study arms based on covariate constrained randomization. Eligible participants were adult women and men 18 years of age and older who met screening criteria for psychological distress and functional impairment. Outcomes were measured at the participant level, with assessors blinded to group assignment. The primary outcome was psychological distress assessed with the General Health Questionnaire (GHQ-12). Secondary outcomes included depression symptoms, posttraumatic stress disorder (PTSD) symptoms, “heart–mind” problems, social support, somatic symptoms, and functional impairment. The hypothesized mediator was skill use aligned with the treatment’s mechanisms of action. A total of 324 participants were enrolled in the control arm (36 wards) and 319 in the Group PM+ arm (36 wards). The overall sample (N = 611) had a median age of 45 years (range 18–91 years), 82% of participants were female, 50% had recently experienced a natural disaster, and 31% had a chronic physical illness. Endline assessments were completed by 302 participants in the control arm (36 wards) and 303 participants in the Group PM+ arm (36 wards). At the midline assessment (immediately after Group PM+ in the experimental arm), mean GHQ-12 total score was 2.7 units lower in Group PM+ compared to control (95% CI: 1.7, 3.7, p < 0.001), with standardized mean difference (SMD) of −0.4 (95% CI: −0.5, −0.2). At 3 months posttreatment (primary endpoint), mean GHQ-12 total score was 1.4 units lower in Group PM+ compared to control (95% CI: 0.3, 2.5, p = 0.014), with SMD of −0.2 (95% CI: −0.4, 0.0). Among the secondary outcomes, Group PM+ was associated with endline with a larger proportion attaining more than 50% reduction in depression symptoms (29.9% of Group PM+ arm versus 17.3% of control arm, risk ratio = 1.7, 95% CI: 1.2, 2.4, p = 0.002). Fewer participants in the Group PM+ arm continued to have “heart–mind” problems at endline (58.8%) compared to the control arm (69.4%), risk ratio = 0.8 (95% CI, 0.7, 1.0, p = 0.042). Group PM+ was not associated with lower PTSD symptoms or functional impairment. Use of psychosocial skills at midline was estimated to explain 31% of the PM+ effect on endline GHQ-12 scores. Adverse events in the control arm included 1 suicide death and 1 reportable incidence of domestic violence; in the Group PM+ arm, there was 1 death due to physical illness. Study limitations include lack of power to evaluate gender-specific effects, lack of long-term outcomes (e.g., 12 months posttreatment), and lack of cost-effectiveness information.ConclusionsIn this study, we found that a 5-session group psychological treatment delivered by nonspecialists modestly reduced psychological distress and depression symptoms in a setting prone to humanitarian emergencies. Benefits were partly explained by the degree of psychosocial skill use in daily life. To improve the treatment benefit, future implementation should focus on approaches to enhance skill use by PM+ participants.Trial registrationClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT03747055","term_id":"NCT03747055"}}NCT03747055.

Mark Jordans and co-workers evaluate a group therapy intervention in adults affected by psychological distress in Nepal.