Could chlorophyllins improve the safety profile of beta-d-N4-hydroxycytidine versus N-hydroxycytidine, the active ingredient of the SARS-CoV-2 antiviral molnupiravir?

Could natural plant pigment (chlorophyll) derivatives (chlorophyllins) improve the safety of the antiviral Molnupiravir, used to treat COVID-19 disease? Molnupiravir, a specific SARS-CoV-2 antiviral, may cause adverse genetic changes and thereby create potential host cell damage (through genotoxicity and DNA stressors). In our opinion, this side effect of treatment could be reduced if the antiviral was taken as a combined therapy with chlorophyllins. Specifically, we hypothesise that chlorophyllins might improve the overall effectiveness of molnupiravir, typically used to treat patients suffering from COVID-19. Chlorophyllins, antioxidants derived from natural plant chlorophyll, are safe, effective and non-toxic antioxidants that could combat possible genotoxic flow-on effects of molnupiravir. In addition, as they possess antiviral properties, treatment with chlorophyllins may enhance the overall antiviral effect via a mechanism different to molnupiravir.

Effect of Brief Produce Exposure and Unconstrained Grocery Gift Cards on Caregiver Influence on Diet of Elementary Age Children: A Randomized Clinical Trial.

Importance: Children's diets affect health trajectories but are difficult to influence, especially for resource-constrained families. Objective: To assess the effectiveness of providing 4 weeks of grocery gift cards and small produce boxes to caregivers on their ability to support healthy shifts in children's diets. Design, Setting, and Participants: This 2-group randomized clinical trial was conducted from May to July 2021, with 4 weeks of intervention and follow-up at 8 weeks. Resources were provided curbside at 3 schools, 3 housing sites, and 1 after-school site for use at home. Participants consisted of 1 index child ages 5 to 11 years with 1 index caregiver from 68 low-income families. Data were analyzed from July 2021 through March 2022. Interventions: During each week for 4 weeks, caregivers were offered 10-lb (4.5 kg) boxes of fruits and vegetables, $10.00 grocery gift cards, an additional $10.00 gift card over the last 3 weeks triggered by a task completion, and a 1-time choice of a $25.00 food preparation tool. Main Outcomes and Measures: Index child and caregiver diets were measured together over the phone at baseline, 4 weeks, and 8 weeks using the 2019 to 2020 Texas School Physical Activity and Nutrition (SPAN) tool, which measures the number of times food items were eaten over the prior day to report a SPAN Healthy Eating Index (SHEI) score and subscores for specific categories of foods (range, 0-57, with higher scores indicating a more healthful diet). Results: Among 68 children (mean [SD] age, 8.2 [1.7] years; 35 [51.5%] girls) and caregivers (mean [SD] age, 37.9 [7.9] years; 63 mothers [92.6%]) from primarily low-income families, 26 caregivers were Hispanic or Latino (38.2%), while 18 caregivers were Black (26.4%), 25 caregivers were White (36.7%), and 24 caregivers had more than 1 race (35.3%). Most families were below the federal poverty level (41 of 60 families that reported income [68.3%]). Per participating caregiver, a mean (SD) 2.7 [1.4] fruit and vegetable boxes and $42.35 ($25.46) worth of gift cards were picked up over 4 weeks. Mean (SE) child SPAN SHEI increased from 32.03 (0.62) times/d to 33.75 (0.69) times/d at 4 weeks (ie, postintervention) and 34.03 (0.69) times/d 4 weeks later (ie, at 8 weeks of follow-up). Mean (SE) child fruit and vegetable intake increased from 5.31 (0.47) times/d to 5.78 (0.51) times/d postintervention and 6.03 (0.51) times/d at follow-up. Children in the control group did not have improved diet (overall mean [SE] SHEI: 31.48 [0.58] times/d at baseline, 31.68 [0.54] times/d postintervention, and 31.81 [0.52] times/d at follow-up; mean [SE] fruit and vegetable intake: 5.21 [0.45] times/d at baseline, 4.77 [0.45] times/d postintervention, and 4.68 [0.41] times/d at follow-up). Compared with children in the control group, mean SHEI was increased for children in the intervention group by 2.07 times/d postintervention and 2.23 times/d at follow-up. Improvements as a function of program dose were statistically significant for child SHEI (P = .01) and fruit and vegetable intake (P = .03). No significant changes in caregiver diets were found. Conclusions and Relevance: This study found that easily accessed fruits and vegetables and unconstrained grocery store cards provided directly to caregivers over 4 weeks resulted in improvements in child diet, which were sustained over 4 additional weeks. Future work may investigate whether diet improvement from a brief intervention optimized for caregiver flexibility reflects a natural maximum or potential for greater improvements on extension. Trial Registration: ClinicalTrials.gov Identifier: NCT04827654.

An Ecologically Framed Comparison of The Potential for Zoonotic Transmission of Non-Human and Human-Infecting Species of Malaria Parasite.

The threats, both real and perceived, surrounding the development of new and emerging infectious diseases of humans are of critical concern to public health and well-being. Among these risks is the potential for zoonotic transmission to humans of species of the malaria parasite, Plasmodium, that have been considered historically to infect exclusively non-human hosts. Recently observed shifts in the mode, transmission, and presentation of malaria among several species studied are evidenced by shared vectors, atypical symptoms, and novel host-seeking behavior. Collectively, these changes indicate the presence of environmental and ecological pressures that are likely to influence the dynamics of these parasite life cycles and physiological make-up. These may be further affected and amplified by such factors as increased urban development and accelerated rate of climate change. In particular, the extended host-seeking behavior of what were once considered non-human malaria species indicates the specialist niche of human malaria parasites is not a limiting factor that drives the success of blood-borne parasites. While zoonotic transmission of non-human malaria parasites is generally considered to not be possible for the vast majority of Plasmodium species, failure to consider the feasibility of its occurrence may lead to the emergence of a potentially life-threatening blood-borne disease of humans. Here, we argue that recent trends in behavior among what were hitherto considered to be non-human malaria parasites to infect humans call for a cross-disciplinary, ecologically-focused approach to understanding the complexities of the vertebrate host/mosquito vector/malaria parasite triangular relationship. This highlights a pressing need to conduct a multi-species investigation for which we recommend the construction of a database to determine ecological differences among all known Plasmodium species, vectors, and hosts. Closing this knowledge gap may help to inform alternative means of malaria prevention and control.

The effects of dry needling to the thoracolumbar junction multifidi on measures of regional and remote flexibility and pain sensitivity: A randomized controlled trial.

BACKGROUND: Dry needling (DN) has been consistently shown to decrease pain sensitivity and increase flexibility local to the site of treatment, however it is unclear whether these effects are limited to the region of treatment or can be observed remote to the area of treatment. OBJECTIVE: To determine the immediate, short-term effects of DN to the thoracolumbar junction on regional and remote flexibility, and to observe if changes in pain sensitivity can occur remote to site of treatment. DESIGN: Double-blind randomized clinical trial. METHODS: Fifty-four subjects with low back pain and decreased length in at least one hamstring were randomized to receive either DN or sham DN to the T12 and L1 multifidi. Participants underwent regional (fingertip-to-floor) and remote flexibility (passive knee extension, passive straight leg raise) and pressure pain threshold (PPT) testing of the upper and lower extremity before, immediately after and 1 day after treatment. ANCOVAs were used to analyze flexibility data, with the covariate of pre-treatment values. Paired t-tests were used for difference in remote pain sensitivity. RESULTS: Statistically larger improvements in regional flexibility, but not remote flexibility, were observed immediately post-treatment in those who received DN than in those receiving sham DN (p = .0495; adjusted difference 1.2, 95% CI 0.002-2.3). Differences between upper and lower extremity PPT were not significant. CONCLUSION: DN can potentially have immediate changes in regional flexibility, but effects are not sustained at 24-h follow-up. DN may not affect remote flexibility or segmental pain sensitivity.

Effect of Layperson-Delivered, Empathy-Focused Program of Telephone Calls on Loneliness, Depression, and Anxiety Among Adults During the COVID-19 Pandemic: A Randomized Clinical Trial.

Importance: Loneliness is a risk factor for many clinical conditions, but there are few effective interventions deployable at scale. Objective: To determine whether a layperson-delivered, empathy-focused program of telephone calls could rapidly improve loneliness, depression, and anxiety in at-risk adults. Design, Setting, and Participants: From July 6 to September 24, 2020, we recruited and followed up 240 adults who were assigned to receive calls (intervention group) or no calls (control group) via block randomization. Loneliness, depression, and anxiety were measured using validated scales at enrollment and after 4 weeks. Intention-to-treat analyses were conducted. Meals on Wheels Central Texas (MOWCTX) clients received calls in their homes or wherever they might have been when the call was received. The study included MOWCTX clients who fit their service criteria, including being homebound and expressing a need for food. A total of 296 participants were screened, of whom 240 were randomized to intervention or control. Interventions: Sixteen callers, aged 17 to 23 years, were briefly trained in empathetic conversational techniques. Each called 6 to 9 participants over 4 weeks daily for the first 5 days, after which clients could choose to drop down to fewer calls but no less than 2 calls a week. Main Outcomes and Measures: Primary outcome was loneliness (3-item UCLA Loneliness Scale, range 3-9; and 6-item De Jong Giervald Loneliness [De Jong] Scale, range 0-6). Secondary outcomes were depression (Personal Health Questionnaire for Depression), anxiety (Generalized Anxiety Disorder scale), and self-rated health (Short Form Health Survey Questionnaire). Results: The 240 participants were aged 27 to 101 years, with 63% aged at least 65 years (n = 149 of 232), 56% living alone (n = 135 of 240), 79% women (n = 190 of 240), 39% Black or African American (n = 94 of 240), and 22% Hispanic or Latino (n = 52 of 240), and all reported at least 1 chronic condition. Of 240 participants enrolled, 13 were lost to follow-up in the intervention arm and 1 in the control arm. Postassessment differences between intervention and control after 4 weeks showed an improvement of 1.1 on the UCLA Loneliness Scale (95% CI, 0.5-1.7; P < .001; Cohen d of 0.48), and improvement of 0.32 on De Jong (95% CI, -0.20 to 0.81; P = .06; Cohen d, 0.17) for loneliness; an improvement of 1.5 on the Personal Health Questionnaire for Depression (95% CI, 0.22-2.7; P < .001; Cohen d, 0.31) for depression; and an improvement of 1.8 on the Generalized Anxiety Disorder scale (95% CI, 0.44 to 3.2; P < .001; Cohen d, 0.35) for anxiety. General physical health on the Short Form Health Questionnaire Survey showed no change, but mental health improved by 2.6 (95% CI, 0.81 to 4.4; P = .003; Cohen d of 0.46). Conclusions and Relevance: A layperson-delivered, empathy-oriented telephone call program reduced loneliness, depression, and anxiety compared with the control group and improved the general mental health of participants within 4 weeks. Future research can determine whether effects on depression and anxiety can be extended to maximize clinical relevance. Trial Registration: ClinicalTrials.gov Identifier: NCT04595708.

Transcriptome and regulatory maps of decidua-derived stromal cells inform gene discovery in preterm birth.

While a genetic component of preterm birth (PTB) has long been recognized and recently mapped by genome-wide association studies (GWASs), the molecular determinants underlying PTB remain elusive. This stems in part from an incomplete availability of functional genomic annotations in human cell types relevant to pregnancy and PTB. We generated transcriptome (RNA-seq), epigenome (ChIP-seq of H3K27ac, H3K4me1, and H3K4me3 histone modifications), open chromatin (ATAC-seq), and chromatin interaction (promoter capture Hi-C) annotations of cultured primary decidua-derived mesenchymal stromal/stem cells and in vitro differentiated decidual stromal cells and developed a computational framework to integrate these functional annotations with results from a GWAS of gestational duration in 56,384 women. Using these resources, we uncovered additional loci associated with gestational duration and target genes of associated loci. Our strategy illustrates how functional annotations in pregnancy-relevant cell types aid in the experimental follow-up of GWAS for PTB and, likely, other pregnancy-related conditions.

Malignant hyperthermia susceptibility: utilization of genetic results in an electronic medical record to increase safety.

Aim: This manuscript describes implementation of clinical decision support for providers concerned with perioperative complications of malignant hyperthermia susceptibility. Materials & methods: Clinical decision support for malignant hyperthermia susceptibility was implemented in 2018 based around our pre-emptive genotyping platform. We completed a brief descriptive review of patients who underwent pre-emptive testing, focused particularly on RYR1 and CACNA1S genes. Results: To date, we have completed pre-emptive genetic testing on more than 10,000 patients; 13 patients having been identified as a carrier of a pathogenic or likely pathogenic variant of RYR1 or CACNA1S. Conclusion: An alert system for malignant hyperthermia susceptibility - as an extension of our pre-emptive genomics platform - was implemented successfully. Implementation strategies and lessons learned are discussed herein.

Single-Copy Knock-In Loci for Defined Gene Expression in Caenorhabditis elegans.

We have generated a single-copy knock-in loci for defined gene expression (SKI LODGE) system to insert any DNA by CRISPR/Cas9 at defined safe harbors in the Caenorhabditis elegans genome. Utilizing a single crRNA guide, which also acts as a Co-CRISPR enrichment marker, any DNA sequence can be introduced as a single copy, regulated by different tissue-specific promoters. The SKI LODGE system provides a fast, economical, and effective approach for generating single-copy ectopic transgenes in C. elegans.

Impact of Pharmacist-Driven Heart Failure in-Home Counseling on 30-Day Readmission Rates.

PURPOSE/OBJECTIVE: This study examines the impact of a pharmacist-driven intervention specific to heart failure patients with the goal of reducing readmission rates and improving quality of life in this population. FINDINGS/CONCLUSIONS: A total of 21 patients were included in the study. Twelve patients were female and 9 were male. The mean age was 76 years with a range of 65-93 years. Two of the 21 patients were readmitted within 30 days. One of the 2 readmitted patients died soon after admission was in the final stages of his or her disease and passed away soon after; it is unlikely for a home visit to have altered their path. This study did not meet the goal sample size due to some unforeseen limitations. However, the limited data that were obtained suggest a strong basis for further research. IMPLICATIONS FOR CASE MANAGEMENT PRACTICE: During a patient's transition in care from hospital to home, he or she is most vulnerable for complications and readmission. Intervention during this time will not only improve patient care and quality of life but also contribute to a notable cost savings for each prevented readmission. Pharmacist intervention, as part of the health care team, during this tenuous time has shown to make a valuable impact.

Knee mechanics during a change of direction movement in division I athletes following full return to sport from anterior cruciate ligament reconstruction.

OBJECTIVES: The objective of this study was to determine whether Division I athletes following anterior cruciate ligament reconstruction (ACLR), and who are medically cleared to return to sport, demonstrate faulty knee mechanics during a change of direction maneuver, as evidenced by the presence of genu valgum on VICON® motion analysis system of 5° or greater. STUDY DESIGN: Cross-sectional study. SETTING: University. PARTICIPANTS: Ten Division I athletes performed a 90° change of direction movement on both the ACLR and unaffected limbs. MAIN OUTCOME MEASURES: Knee mechanics were assessed using the VICON® motion analysis system. Knee position was analyzed in the frontal plane and peak genu valgum position was determined during the functional movement. RESULTS: Eight out of ten athletes (80%) demonstrated genu valgum ≥ 5° on the ACLR limb during a 90° cutting movement. Fourteen out of twenty (70%) of the knees assessed demonstrated genu valgum ≥ 5° during a 90° cutting movement. CONCLUSION: Athletes who have fully returned to sport following an ACLR demonstrate substantial genu valgum during a cutting maneuver, which suggests an increased risk for future injury. These results show an increased need during ACL rehabilitation to focus on existing biomechanical deficits during sport-specific movements.

THE RELATIONSHIP BETWEEN SEGMENTAL ROLLING ABILITY AND LUMBAR MULTIFIDUS ACTIVATION TIME.

BACKGROUND: Segmental rolling has been utilized as an assessment and intervention tool to identify and affect dysfunction of the upper quarter, core, and lower quarter. One theory to explain dysfunctional segmental rolling is a lack of segmental spinal control / stabilization. Faulty muscle firing sequencing has been related to poor spinal stability, however to date, no assessment tool exists to evaluate a patient's motor coordination of local and global musculature. PURPOSE: The purpose of this study was to assess the temporal sequence of lumbar multifidus activation associated with anterior deltoid activation, and to determine if faulty sequencing was associated with the inability to segmentally roll in subjects without mobility restrictions. The authors hypothesized that in individuals who could not roll, a multifidus muscle onset latency relative to a prime mover activation would be present. In addition, a subset of the individuals with an inability to roll were utilized for a pilot study examining the ability to address the firing pattern with corrective exercise. METHODS: Twenty healthy subjects (13 females, 7 males), ages 19-25, participated in the study. Each subject underwent an upper and lower quarter screen and assessment of thoracic spine mobility. Subjects were excluded from the study if they had previous spine surgery, or were currently experiencing back pain. In addition, subjects who had any disease, disorders, or pathology that would hinder participation in segmental rolling or who had spinal movement contraindications were excluded. Since shoulder flexion is performed during the study, participants who had shoulder pathology or contraindications to upper extremity movement were excluded as well. Subjects with less than 50 degrees of trunk rotation were excluded from the study due to a possible physical mobility limitation that would prevent proficient segmental rolling. Included subjects were assessed on their ability to segmentally roll. Subjects who could complete the rolling task were placed in cohort A ("can roll"), and subjects who could not roll were placed in cohort B ("can't roll").Electromyographic (EMG) activity of the multifidus was recorded adjacent to the lamina of the L4 vertebrae using intramuscular fine-wire electrodes. EMG activity of the anterior deltoid was also recorded with a surface electrode during a single arm movement into shoulder flexion. While in a standing position, subjects were instructed to move their right upper arm into flexion as quickly as possible. Subjects flexed their shoulder to 90 degrees for three trials while muscle activity was recorded. Data were high-pass filtered at 30 Hz to remove baseline artifact, and the onset EMG times was selected as the point at which EMG increased two SD above baseline levels. Onset of the multifidus muscle was reported relative to that of the prime mover (anterior deltoid). Muscle onset latency was defined as the time difference between the onset of contraction of the multifidus and the anterior deltoid. RESULTS: Nine subjects were placed in cohort A, 11 subjects were placed in cohort B. The mean firing time of the lumbar multifidus for the cohort A was 16.67msec before the anterior deltoid, and the mean firing time of the lumbar multifidus for cohort B was 57.36msec after the anterior deltoid. There was a statistically significant difference (p<0.00) in the firing time between cohorts A and B. CONCLUSIONS: In subjects who could segmentally roll, the multifidus muscle activation always preceded that of the prime mover muscle activation. In subjects who could not segmentally roll, the results of this study confirm that there is a multifidus muscle onset latency relative to the activation of the anterior deltoid. The inability to segmentally roll may be related to faulty sequencing of lumbar multifidus firing.

Conditional Ablation of Progesterone Receptor Membrane Component 2 Causes Female Premature Reproductive Senescence.

The nonclassical progesterone receptors progesterone receptor membrane component (PGRMC) 1 and PGRMC2 have been implicated in regulating cell survival of endometrial and ovarian cells in vitro and are abundantly expressed in these cell types. The objective of this study was to determine if Pgrmc1 and Pgrmc2 are essential for normal female reproduction. To accomplish this objective, Pgrmc1 and/or Pgrmc2 floxed mice (Pgrmc2fl/fl and Pgrmc1/2fl/fl) were crossed with Pgr-cre mice, which resulted in the conditional ablation of Pgrmc1 and/or Pgrmc2 from female reproductive tissues (i.e.,Pgrmc2d/d and Pgrmc1/2d/d mice). A breeding trial revealed that conditional ablation of Pgrmc2 initially led to subfertility, with Pgrmc2d/d female mice producing 47% fewer pups/litter than Pgrmc2fl/fl mice (P = 0.001). Pgrmc2d/d mice subsequently underwent premature reproductive senescence by parities 2 to 5, producing 37.8% fewer litters overall during the trial compared with Pgrmc2fl/fl mice (P = 0.020). Similar results were observed with Pgrmc1/2d/d mice. Based on ovarian morphology and serum P4, the subfertility/infertility was not due to faulty ovulation or luteal insufficiency. Rather an analysis of midgestation implantation sites revealed that postimplantation embryonic death was the major cause of the subfertility/infertility. As with our previous report of Pgrmc1d/d mice, Pgrmc2d/d and Pgrmc1/2d/d mice developed endometrial cysts consistent with accelerated aging of this tissue. Given the timing of postimplantation embryonic demise, uterine decidualization may be disrupted in mice deficient in PGRMC2 or PGRMC1/2. Overall, this study revealed that Pgrmc1 and/or Pgrmc2 are required for the maintenance of uterine histoarchitecture and normal female reproductive lifespan.

Progesterone receptor membrane component 1 promotes survival of human breast cancer cells and the growth of xenograft tumors.

Triple negative breast cancers (TNBCs) are highly aggressive and grow in response to sex steroid hormones despite lacking expression of the classical estrogen (E2) and progesterone (P4) receptors. Since P4 receptor membrane component 1 (PGRMC1) is expressed in breast cancer tumors and is known to mediate P4-induced cell survival, this study was designed to determine the expression of PGRMC1 in TNBC tumors and the involvement of PGRMC1 in regulating proliferation and survival of TNBC cells in vitro and the growth of TNBC tumors in vivo. For the latter studies, the MDA-MB-231 (MDA) cell line derived from TNBC was used. These cells express PGRMC1 but lack expression of the classical P4 receptor. A lentiviral-based shRNA approach was used to generate a stably transfected PGRMC1-deplete MDA line for comparison to the PGRMC1-intact MDA line. The present studies demonstrate that PGRMC1: 1) is expressed in TNBC cells; 2) mediates the ability of P4 to suppress TNBC cell mitosis in vitro; 3) is required for P4 to reduce the apoptotic effects of doxorubicin in vitro; and 4) facilitates TNBC tumor formation and growth in vivo. Taken together, these findings indicate that PGRMC1 plays an important role in regulating the growth and survival of TNBC cells in vitro and ultimately in the formation and development of these tumors in vivo. Thus, PGRMC1 may be a therapeutic target for TNBCs.

Differences in metacognitive regulation in introductory biology students: when prompts are not enough.

Strong metacognition skills are associated with learning outcomes and student performance. Metacognition includes metacognitive knowledge-our awareness of our thinking-and metacognitive regulation-how we control our thinking to facilitate learning. In this study, we targeted metacognitive regulation by guiding students through self-evaluation assignments following the first and second exams in a large introductory biology course (n = 245). We coded these assignments for evidence of three key metacognitive-regulation skills: monitoring, evaluating, and planning. We found that nearly all students were willing to take a different approach to studying but showed varying abilities to monitor, evaluate, and plan their learning strategies. Although many students were able to outline a study plan for the second exam that could effectively address issues they identified in preparing for the first exam, only half reported that they followed their plans. Our data suggest that prompting students to use metacognitive-regulation skills is effective for some students, but others need help with metacognitive knowledge to execute the learning strategies they select. Using these results, we propose a continuum of metacognitive regulation in introductory biology students. By refining this model through further study, we aim to more effectively target metacognitive development in undergraduate biology students.

Drosophila muller f elements maintain a distinct set of genomic properties over 40 million years of evolution.

The Muller F element (4.2 Mb, ~80 protein-coding genes) is an unusual autosome of Drosophila melanogaster; it is mostly heterochromatic with a low recombination rate. To investigate how these properties impact the evolution of repeats and genes, we manually improved the sequence and annotated the genes on the D. erecta, D. mojavensis, and D. grimshawi F elements and euchromatic domains from the Muller D element. We find that F elements have greater transposon density (25-50%) than euchromatic reference regions (3-11%). Among the F elements, D. grimshawi has the lowest transposon density (particularly DINE-1: 2% vs. 11-27%). F element genes have larger coding spans, more coding exons, larger introns, and lower codon bias. Comparison of the Effective Number of Codons with the Codon Adaptation Index shows that, in contrast to the other species, codon bias in D. grimshawi F element genes can be attributed primarily to selection instead of mutational biases, suggesting that density and types of transposons affect the degree of local heterochromatin formation. F element genes have lower estimated DNA melting temperatures than D element genes, potentially facilitating transcription through heterochromatin. Most F element genes (~90%) have remained on that element, but the F element has smaller syntenic blocks than genome averages (3.4-3.6 vs. 8.4-8.8 genes per block), indicating greater rates of inversion despite lower rates of recombination. Overall, the F element has maintained characteristics that are distinct from other autosomes in the Drosophila lineage, illuminating the constraints imposed by a heterochromatic milieu.

Refinement of animal models of sepsis and septic shock.

This report aims to facilitate the implementation of the Three Rs (replacement, reduction, and refinement) in the use of animal models or procedures involving sepsis and septic shock, an area where there is the potential of high levels of suffering for animals. The emphasis is on refinement because this has the greatest potential for immediate implementation. Specific welfare issues are identified and discussed, and practical measures are proposed to reduce animal use and suffering as well as reducing experimental variability and increasing translatability. The report is based on discussions and submissions from a nonregulatory expert working group consisting of veterinarians, animal technologists, and scientists with expert knowledge relevant to the field.

Progesterone receptor membrane component 1 deficiency attenuates growth while promoting chemosensitivity of human endometrial xenograft tumors.

Endometrial cancer is the leading gynecologic cancer in women in the United States with 52,630 women predicted to be diagnosed with the disease in 2014. The objective of this study was to determine if progesterone (P4) receptor membrane component 1 (PGRMC1) influenced endometrial cancer cell viability in response to chemotherapy in vitro and in vivo. A lentiviral-based shRNA knockdown approach was used to generate stable PGRMC1-intact and PGRMC1-deplete Ishikawa endometrial cancer cell lines that also lacked expression of the classical progesterone receptor (PGR). Progesterone treatment inhibited mitosis of PGRMC1-intact, but not PGRMC1-deplete cells, suggesting that PGRMC1 mediates the anti-mitotic actions of P4. To test the hypothesis that PGRMC1 attenuates chemotherapy-induced apoptosis, PGRMC1-intact and PGRMC1-deplete cells were treated in vitro with vehicle, P4 (1 µM), doxorubicin (Dox, 2 µg/ml), or P4 + Dox for 48 h. Doxorubicin treatment of PGRMC1-intact cells resulted in a significant increase in cell death; however, co-treatment with P4 significantly attenuated Dox-induced cell death. This response to P4 was lost in PGRMC1-deplete cells. To extend these observations in vivo, a xenograft model was employed where PGRMC1-intact and PGRMC1-deplete endometrial tumors were generated following subcutaneous and intraperitoneal inoculation of immunocompromised NOD/SCID and nude mice, respectively. Tumors derived from PGRMC1-deplete cells grew slower than tumors from PGRMC1-intact cells. Mice harboring endometrial tumors were then given three treatments of vehicle (1:1 cremophor EL: ethanol + 0.9% saline) or chemotherapy [Paclitaxel (15 mg/kg, i.p.) followed after an interval of 30 minutes by CARBOplatin (50 mg/kg)] at five day intervals. In response to chemotherapy, tumor volume decreased approximately four-fold more in PGRMC1-deplete tumors when compared with PGRMC1-intact control tumors, suggesting that PGRMC1 promotes tumor cell viability during chemotherapeutic stress. In sum, these in vitro and in vivo findings demonstrate that PGRMC1 plays a prominent role in the growth and chemoresistance of human endometrial tumors.

A migrating ciliary gate compartmentalizes the site of axoneme assembly in Drosophila spermatids.

BACKGROUND: In most cells, the cilium is formed within a compartment separated from the cytoplasm. Entry into the ciliary compartment is regulated by a specialized gate located at the base of the cilium in a region known as the transition zone. The transition zone is closely associated with multiple structures of the ciliary base, including the centriole, axoneme, and ciliary membrane. However, the contribution of these structures to the ciliary gate remains unclear. RESULTS: Here we report that, in Drosophila spermatids, a conserved module of transition zone proteins mutated in Meckel-Gruber syndrome (MKS), including Cep290, Mks1, B9d1, and B9d2, comprise a ciliary gate that continuously migrates away from the centriole to compartmentalize the growing axoneme tip. We show that Cep290 is essential for transition zone composition, compartmentalization of the axoneme tip, and axoneme integrity and find that MKS proteins also delimit a centriole-independent compartment in mouse spermatids. CONCLUSIONS: Our findings demonstrate that the ciliary gate can migrate away from the base of the cilium, thereby functioning independently of the centriole and of a static interaction with the axoneme to compartmentalize the site of axoneme assembly.