Hematopoietic stem cell niche generation and maintenance are distinguishable by an epitranscriptomic program.

The niche is typically considered as a pre-established structure sustaining stem cells. Therefore, the regulation of its formation remains largely unexplored. Whether distinct molecular mechanisms control the establishment versus maintenance of a stem cell niche is unknown. To address this, we compared perinatal and adult bone marrow mesenchymal stromal cells (MSCs), a key component of the hematopoietic stem cell (HSC) niche. MSCs exhibited enrichment in genes mediating m6A mRNA methylation at the perinatal stage and downregulated the expression of Mettl3, the m6A methyltransferase, shortly after birth. Deletion of Mettl3 from developing MSCs but not osteoblasts led to excessive osteogenic differentiation and a severe HSC niche formation defect, which was significantly rescued by deletion of Klf2, an m6A target. In contrast, deletion of Mettl3 from MSCs postnatally did not affect HSC niche. Stem cell niche generation and maintenance thus depend on divergent molecular mechanisms, which may be exploited for regenerative medicine.

Mechanism of noncoding RNA-associated N6-methyladenosine recognition by an RNA processing complex during IgH DNA recombination.

Immunoglobulin heavy chain (IgH) locus-associated G-rich long noncoding RNA (SµGLT) is important for physiological and pathological B cell DNA recombination. We demonstrate that the METTL3 enzyme-catalyzed N6-methyladenosine (m6A) RNA modification drives recognition and 3' end processing of SµGLT by the RNA exosome, promoting class switch recombination (CSR) and suppressing chromosomal translocations. The recognition is driven by interaction of the MPP6 adaptor protein with nuclear m6A reader YTHDC1. MPP6 and YTHDC1 promote CSR by recruiting AID and the RNA exosome to actively transcribe SµGLT. Direct suppression of m6A modification of SµGLT or of m6A reader YTHDC1 reduces CSR. Moreover, METTL3, an essential gene for B cell development in the bone marrow and germinal center, suppresses IgH-associated aberrant DNA breaks and prevents genomic instability. Taken together, we propose coordinated and central roles for MPP6, m6A modification, and m6A reader proteins in controlling long noncoding RNA processing, DNA recombination, and development in B cells.

Substance use disorder approaches in US primary care clinics with national reputations as workforce innovators.

BACKGROUND: Over the last decade, primary care clinics in the United States have responded both to national policies encouraging clinics to support substance use disorders (SUD) service expansion and to regulations aiming to curb the opioid epidemic. OBJECTIVE: To characterize approaches to SUD service expansion in primary care clinics with national reputations as workforce innovators. METHODS: Comparative case studies were conducted to characterize different approaches among 12 primary care clinics purposively and iteratively recruited from a national registry of workforce innovators. Observational field notes and qualitative interviews from site visits were coded and analysed to identify and characterize clinic attributes. RESULTS: Codes describing clinic SUD expansion approaches emerged from our analysis. Clinics were characterized as: avoidant (n = 3), contemplative (n = 5) and responsive (n = 4). Avoidant clinics were resistant to planning SUD service expansion; had no or few on-site behavioural health staff; and lacked on-site medication treatment (previously termed medication-assisted therapy) waivered providers. Contemplative clinics were planning or had partially implemented SUD services; members expressed uncertainties about expansion; had co-located behavioural healthcare providers, but no on-site medication treatment waivered and prescribing providers. Responsive clinics had fully implemented SUD; members used non-judgmental language about SUD services; had both co-located SUD behavioural health staff trained in SUD service provision and waivered medication treatment physicians and/or a coordinated referral pathway. CONCLUSIONS: Efforts to support SUD service expansion should tailor implementation supports based on specific clinic training and capacity building needs. Future work should inform the adaption of evidence-based practices that are responsive to resource constraints to optimize SUD treatment access.

Primary care clinics in the US have been encouraged to expand addiction services to increase treatment access and respond to the opioid epidemic. This study uses structured observations and depth interviews to assess and compare how primary care clinics with innovative workforces have responded to the growing need for substance use disorder treatment. Each of the clinics studied represents a 'case.' We systematically compared cases to understand how and if addiction services were expanded. Twelve clinic 'cases' were coded and characterized based on a continuum of receptivity ranging from avoidant (i.e., resistant), contemplative (i.e., organization members plan to implement change) and responsive (i.e. expansions implemented). Our analysis characterized three clinics as avoidant to expanding addiction services reporting no plans to respond to calls to expand addiction services. Five clinics were characterized as contemplative, meaning they recognized the need but still had reservations and concerns about the expansion. Four clinics were characterized as responsive to addiction service expansion and had several organizational-wide strategies to assess, intervene and treat patients with addictions. Despite national and state-based policies to entice clinics to expand addiction services there was a diversity of approaches observed in clinics. Avoidant and contemplative clinics may need implementation support to build capacity for this type of delivery expansion.

Mental health nurses experience of the introduction and practice of the Safewards model: a qualitative descriptive study.

BACKGROUND: A lack of safety experienced by patients and staff in acute psychiatric units is a major concern and containment methods used to manage conflict have the potential to cause harm and upset to both staff and patients. To ensure safety for all, it is highly desirable to reduce levels of conflict and containment and the Safewards model is an evidence-based model aimed at reducing conflict and containment rates by improving nurse-patient relationships and safety. METHODS: The aim of this study was to explore mental health nurses' experience of the introduction and practice of three Safewards interventions; reassurance, soft words and discharge messages. A qualitative descriptive research design utilising a purposive sample (n = 21) of registered psychiatric nurses (n = 16) and managers (n = 5) in an acute psychiatric unit in Ireland. Following a 12-week implementation of Safewards, three focus groups were conducted, two with nursing staff and one with nurse managers. Data were analysed using Braun and Clarke thematic analysis framework which supported the identification of four themes: introducing Safewards, challenges of Safewards, impact of Safewards and working towards success. RESULTS: The findings indicate that the process of implementation was inadequate in the training and education of staff, and that poor support from management led to poor staff adherence and acceptance of the Safewards interventions. The reported impact of Safewards on nursing practice and patient experience were mixed. Overall, engagement and implementation under the right conditions are essential for success and while some participants perceived that the interventions already existed in practice, participants agreed Safewards enhanced their communication skills and relationships with patients. CONCLUSION: The implementation of Safewards requires effective leadership and support from management, mandatory training for all staff, and the involvement of staff and patients during implementation. Future research should focus on the training and education required for successful implementation of Safewards and explore the impact of Safewards on nursing practice and patient experience.

Tracking the embryonic stem cell transition from ground state pluripotency.

Mouse embryonic stem (ES) cells are locked into self-renewal by shielding from inductive cues. Release from this ground state in minimal conditions offers a system for delineating developmental progression from naïve pluripotency. Here, we examine the initial transition process. The ES cell population behaves asynchronously. We therefore exploited a short-half-life Rex1::GFP reporter to isolate cells either side of exit from naïve status. Extinction of ES cell identity in single cells is acute. It occurs only after near-complete elimination of naïve pluripotency factors, but precedes appearance of lineage specification markers. Cells newly departed from the ES cell state display features of early post-implantation epiblast and are distinct from primed epiblast. They also exhibit a genome-wide increase in DNA methylation, intermediate between early and late epiblast. These findings are consistent with the proposition that naïve cells transition to a distinct formative phase of pluripotency preparatory to lineage priming.

Reprint of: The role of the pharmacist in mental health: An investigation of the impact of pharmacist-led interventions on psychotropic medication adherence in patients with diabetes.

OBJECTIVE: To evaluate the impact of targeted telepharmacist mental health adherence interventions for patients with type 2 diabetes. METHODS: This retrospective review involved the analysis of a telepharmacist-led mental health intervention. The subjects included: patients aged 18 years or more with type 2 diabetes, enrolled in an adherence service and who had been prescribed psychotropics. Psychotropic medication adherence was measured using the proportion of days covered (PDC) 6 months before and after the telepharmacist-led medication intervention. RESULTS: A total of 8167 patients (67% women), with a mean age of 63 ± 11 years, were included in the study. Most alerts for psychotropics were for selective serotonin reuptake inhibitors (SSRIs) (53%, n = 4334), selective norepinephrine reuptake inhibitors (SNRIs) (22%, n = 1775), second-generation antipsychotics (11%, n = 909), and bupropion (10%, n = 782). Alerts with the greatest volume of PDCs (above 85%) at postintervention follow up included SSRIs (51%, n = 2228), SNRIs (50%, n = 884), and second-generation antipsychotics (47%, n = 424). Before the alert, the mean PDC was 66% ± 12% across all medications studied. Post intervention, the mean PDC rose to 79% ± 19. A paired t-test revealed statistically significant improvement in adherence overall, with the greatest change observed in these alert groups: SSRIs (P < 0.001), alpha-2 antagonist (P < 0.001), SNRIs (P < 0.001), and bupropion (P < 0.001). CONCLUSION: This retrospective review showed that pharmacist-led targeted, adherence interventions greatly improved psychotropic medication adherence in adult patients with type 2 diabetes. Future work is warranted to investigate the impact on type 2 diabetes medication adherence and health markers (e.g., HbA1c values) in larger and more diverse populations of patients with comorbid type 2 diabetes and a mental health condition.

The role of the pharmacist in mental health: An investigation of the impact of pharmacist-led interventions on psychotropic medication adherence in patients with diabetes.

OBJECTIVE: To evaluate the impact of targeted telepharmacist mental health adherence interventions for patients with type 2 diabetes. METHODS: This retrospective review involved the analysis of a telepharmacist-led mental health intervention. The subjects included: patients aged 18 years or more with type 2 diabetes, enrolled in an adherence service and who had been prescribed psychotropics. Psychotropic medication adherence was measured using the proportion of days covered (PDC) 6 months before and after the telepharmacist-led medication intervention. RESULTS: A total of 8167 patients (67% women), with a mean age of 63 ± 11 years, were included in the study. Most alerts for psychotropics were for selective serotonin reuptake inhibitors (SSRIs) (53%, n = 4334), selective norepinephrine reuptake inhibitors (SNRIs) (22%, n = 1775), second-generation antipsychotics (11%, n = 909), and bupropion (10%, n = 782). Alerts with the greatest volume of PDCs (above 85%) at postintervention follow up included SSRIs (51%, n = 2228), SNRIs (50%, n = 884), and second-generation antipsychotics (47%, n = 424). Before the alert, the mean PDC was 66% ± 12% across all medications studied. Post intervention, the mean PDC rose to 79% ± 19. A paired t-test revealed statistically significant improvement in adherence overall, with the greatest change observed in these alert groups: SSRIs (P < 0.001), alpha-2 antagonist (P < 0.001), SNRIs (P < 0.001), and bupropion (P < 0.001). CONCLUSION: This retrospective review showed that pharmacist-led targeted, adherence interventions greatly improved psychotropic medication adherence in adult patients with type 2 diabetes. Future work is warranted to investigate the impact on type 2 diabetes medication adherence and health markers (e.g., HbA1c values) in larger and more diverse populations of patients with comorbid type 2 diabetes and a mental health condition.

Internalization of Western Ideals on Appearance and Self-Esteem in Jamaican Undergraduate Students.

Beauty ideals in the Caribbean are shifting with increased exposure to Western and European standards of appearance. Previous research has shown a consistent link between internalization of Western beauty ideals and depressive symptoms and other forms of psychological disturbance among diverse populations including Caribbeans. We examined the association between internalization of Western beauty ideals and depressive symptoms as well as the potential mediating role of self-esteem on this relation in N = 222 students (155 females, 79 males) attending a tertiary institution in Kingston, Jamaica. Internalization of Western ideals was inversely associated with self-esteem (r =- .35, p < .01) and positively associated with depressive symptoms (r =.13, p < .05). In a model adjusted for age and sex, results revealed a significant indirect effect of internalization of Western ideals of appearance on depressive symptoms via self-esteem (estimate= .21, SE = .05, 95% confidence interval [.13, .32]). The potent effects of culture must be better understood as intercontinental travel becomes less important as a mechanism for cultural exposure and exchange, and there is a significant increase of digital and internet access in the Caribbean. The current study suggest that Caribbeans are at significant risk for internalizing Western ideals of beauty, subsequently diminishing their self-esteem, and ultimately increasing depression symptomatology. The benefits and consequences of cultural exchange should continue to be a topic for research studies.

The DUX4 homeodomains mediate inhibition of myogenesis and are functionally exchangeable with the Pax7 homeodomain.

Facioscapulohumeral muscular dystrophy (FSHD) is caused by inappropriate expression of the double homeodomain protein DUX4. DUX4 has bimodal effects, inhibiting myogenic differentiation and blocking MyoD at low levels of expression, and killing myoblasts at high levels. Pax3 and Pax7, which contain related homeodomains, antagonize the cell death phenotype of DUX4 in C2C12 cells, suggesting some type of competitive interaction. Here, we show that the effects of DUX4 on differentiation and MyoD expression require the homeodomains but do not require the C-terminal activation domain of DUX4. We tested the set of equally related homeodomain proteins (Pax6, Pitx2c, OTX1, Rax, Hesx1, MIXL1 and Tbx1) and found that only Pax3 and Pax7 display phenotypic competition. Domain analysis on Pax3 revealed that the Pax3 homeodomain is necessary for phenotypic competition, but is not sufficient, as competition also requires the paired and transcriptional activation domains of Pax3. Remarkably, substitution mutants in which DUX4 homeodomains are replaced by Pax7 homeodomains retain the ability to inhibit differentiation and to induce cytotoxicity.

Parallel single-cell sequencing links transcriptional and epigenetic heterogeneity.

We report scM&T-seq, a method for parallel single-cell genome-wide methylome and transcriptome sequencing that allows for the discovery of associations between transcriptional and epigenetic variation. Profiling of 61 mouse embryonic stem cells confirmed known links between DNA methylation and transcription. Notably, the method revealed previously unrecognized associations between heterogeneously methylated distal regulatory elements and transcription of key pluripotency genes.

Genomic Modifiers of Natural Killer Cells, Immune Responsiveness and Lymphoid Tissue Remodeling Together Increase Host Resistance to Viral Infection.

The MHC class I D(k) molecule supplies vital host resistance during murine cytomegalovirus (MCMV) infection. Natural killer (NK) cells expressing the Ly49G2 inhibitory receptor, which specifically binds D(k), are required to control viral spread. The extent of D(k)-dependent host resistance, however, differs significantly amongst related strains of mice, C57L and MA/My. As a result, we predicted that relatively small-effect modifier genetic loci might together shape immune cell features, NK cell reactivity, and the host immune response to MCMV. A robust D(k)-dependent genetic effect, however, has so far hindered attempts to identify additional host resistance factors. Thus, we applied genomic mapping strategies and multicolor flow cytometric analysis of immune cells in naive and virus-infected hosts to identify genetic modifiers of the host immune response to MCMV. We discovered and validated many quantitative trait loci (QTL); these were mapped to at least 19 positions on 16 chromosomes. Intriguingly, one newly discovered non-MHC locus (Cmv5) controlled splenic NK cell accrual, secondary lymphoid organ structure, and lymphoid follicle development during MCMV infection. We infer that Cmv5 aids host resistance to MCMV infection by expanding NK cells needed to preserve and protect essential tissue structural elements, to enhance lymphoid remodeling and to increase viral clearance in spleen.

ELF5 Drives Lung Metastasis in Luminal Breast Cancer through Recruitment of Gr1+ CD11b+ Myeloid-Derived Suppressor Cells.

During pregnancy, the ETS transcription factor ELF5 establishes the milk-secreting alveolar cell lineage by driving a cell fate decision of the mammary luminal progenitor cell. In breast cancer, ELF5 is a key transcriptional determinant of tumor subtype and has been implicated in the development of insensitivity to anti-estrogen therapy. In the mouse mammary tumor virus-Polyoma Middle T (MMTV-PyMT) model of luminal breast cancer, induction of ELF5 levels increased leukocyte infiltration, angiogenesis, and blood vessel permeability in primary tumors and greatly increased the size and number of lung metastasis. Myeloid-derived suppressor cells, a group of immature neutrophils recently identified as mediators of vasculogenesis and metastasis, were recruited to the tumor in response to ELF5. Depletion of these cells using specific Ly6G antibodies prevented ELF5 from driving vasculogenesis and metastasis. Expression signatures in luminal A breast cancers indicated that increased myeloid cell invasion and inflammation were correlated with ELF5 expression, and increased ELF5 immunohistochemical staining predicted much shorter metastasis-free and overall survival of luminal A patients, defining a group who experienced unexpectedly early disease progression. Thus, in the MMTV-PyMT mouse mammary model, increased ELF5 levels drive metastasis by co-opting the innate immune system. As ELF5 has been previously implicated in the development of antiestrogen resistance, this finding implicates ELF5 as a defining factor in the acquisition of the key aspects of the lethal phenotype in luminal A breast cancer.

The Potential Effects of Caffeinated Beverages on Insulin Sensitivity.

Caffeinated beverages, most commonly tea and coffee, may have important effects on insulin regulation that may give their consumption an important role among nutritional factors in the development of diseases of glucose and insulin metabolism, such as diabetes and atherosclerotic vascular diseases. These beverages include compounds that may have contradictory effects on insulin and glucose: Caffeine impairs insulin sensitivity, but polyphenolic molecules within tea, coffee, and cocoa augment the effects of insulin. In addition, epidemiologic associations exist between greater consumption of such beverages and lower risk of diabetes. The beneficial effects of such beverages might be enhanced by changing the process of their preparation and substitution of other substances commonly added to caffeinated beverages that impair the effect of insulin, such as sugar or milk.

Endoscopic Browplasty.

The endoscopic brow lift has become an established procedure that can safely and reliably rejuvenate the upper third of the face. The authors discuss relevant anatomy and considerations for patient selection to optimize surgical outcomes. A detailed review of surgical technique is presented, and the potential complications and means to reduce them are discussed.

Single-cell genome-wide bisulfite sequencing for assessing epigenetic heterogeneity.

We report a single-cell bisulfite sequencing (scBS-seq) method that can be used to accurately measure DNA methylation at up to 48.4% of CpG sites. Embryonic stem cells grown in serum or in 2i medium displayed epigenetic heterogeneity, with '2i-like' cells present in serum culture. Integration of 12 individual mouse oocyte datasets largely recapitulated the whole DNA methylome, which makes scBS-seq a versatile tool to explore DNA methylation in rare cells and heterogeneous populations.

ELF5 isoform expression is tissue-specific and significantly altered in cancer.

BACKGROUND: E74-like factor 5 (ELF5) is an epithelial-specific member of the E26 transforming sequence (ETS) transcription factor family and a critical regulator of cell fate in the placenta, pulmonary bronchi, and milk-producing alveoli of the mammary gland. ELF5 also plays key roles in malignancy, particularly in basal-like and endocrine-resistant forms of breast cancer. Almost all genes undergo alternative transcription or splicing, which increases the diversity of protein structure and function. Although ELF5 has multiple isoforms, this has not been considered in previous studies of ELF5 function. METHODS: RNA-sequencing data for 6757 samples from The Cancer Genome Atlas were analyzed to characterize ELF5 isoform expression in multiple normal tissues and cancers. Extensive in vitro analysis of ELF5 isoforms, including a 116-gene quantitative polymerase chain reaction panel, was performed in breast cancer cell lines. RESULTS: ELF5 isoform expression was found to be tissue-specific due to alternative promoter use but altered in multiple cancer types. The normal breast expressed one main isoform, while in breast cancer there were subtype-specific alterations in expression. Expression of other ETS factors was also significantly altered in breast cancer, with the basal-like subtype demonstrating a distinct ETS expression profile. In vitro inducible expression of the full-length isoforms 1 and 2, as well as isoform 3 (lacking the Pointed domain) had similar phenotypic and transcriptional effects. CONCLUSIONS: Alternative promoter use, conferring differential regulatory responses, is the main mechanism governing ELF5 action rather than differential transcriptional activity of the isoforms. This understanding of expression and function at the isoform level is a vital first step in realizing the potential of transcription factors such as ELF5 as prognostic markers or therapeutic targets in cancer.

Progesterone drives mammary secretory differentiation via RankL-mediated induction of Elf5 in luminal progenitor cells.

Progesterone-RankL paracrine signaling has been proposed as a driver of stem cell expansion in the mammary gland, and Elf5 is essential for the differentiation of mammary epithelial progenitor cells. We demonstrate that Elf5 expression is induced by progesterone and that Elf5 and progesterone cooperate to promote alveolar development. The progesterone receptor and Elf5 are expressed in a mutually exclusive pattern, and we identify RankL as the paracrine mediator of the effects of progesterone on Elf5 expression in CD61+ progenitor cells and their consequent differentiation. Blockade of RankL action prevented progesterone-induced side branching and the expansion of Elf5(+) mature luminal cells. These findings describe a mechanism by which steroid hormones can produce the expansion of steroid hormone receptor-negative mammary epithelial cells.

Osteoclast differentiation factor RANKL controls development of progestin-driven mammary cancer.

Breast cancer is one of the most common cancers in humans and will on average affect up to one in eight women in their lifetime in the United States and Europe. The Women's Health Initiative and the Million Women Study have shown that hormone replacement therapy is associated with an increased risk of incident and fatal breast cancer. In particular, synthetic progesterone derivatives (progestins) such as medroxyprogesterone acetate (MPA), used in millions of women for hormone replacement therapy and contraceptives, markedly increase the risk of developing breast cancer. Here we show that the in vivo administration of MPA triggers massive induction of the key osteoclast differentiation factor RANKL (receptor activator of NF-κB ligand) in mammary-gland epithelial cells. Genetic inactivation of the RANKL receptor RANK in mammary-gland epithelial cells prevents MPA-induced epithelial proliferation, impairs expansion of the CD49f(hi) stem-cell-enriched population, and sensitizes these cells to DNA-damage-induced cell death. Deletion of RANK from the mammary epithelium results in a markedly decreased incidence and delayed onset of MPA-driven mammary cancer. These data show that the RANKL/RANK system controls the incidence and onset of progestin-driven breast cancer.

Follow-up Care Education and Information: Identifying Cancer Survivors in Need of More Guidance.

Cancer survivors engage in cancer screenings and protective health behaviors at suboptimal rates despite their increased risk for future illness. Survivorship care plans and other educational strategies to prepare cancer survivors to adopt engaged roles in managing long-term follow-up care and health risks are needed. In a sample of cancer survivors, we identified patient characteristics and psychosocial predictors associated with increased follow-up care informational needs. Cross-sectional surveys were administered to early-stage breast and prostate survivors (N = 278; 68 % breast) at least 2 years post treatment from four community hospital programs in New Jersey between May 2012 and July 2013. Patient demographics, medical history, psychosocial characteristics (i.e., worries about the future, fear of disease recurrence, and patient activation), and perceptions of oncology and primary care were assessed. African-American survivors (AOR = 2.69, 95 % confidence interval [CI] 1.27-5.68) and survivors with higher comorbidity (AOR =1.16, CI 1.01-1.33) were more likely to want additional information to guide follow-up care. Adjusting for race and comorbidities, survivors who wanted more information to guide their follow-up care reported greater worries about the future (p < 0.05) and fears about disease recurrence (p < 0.05) compared to those who did not want additional information. Results emphasize the need to develop cancer survivorship educational strategies that are both responsive to the needs of specific populations (e.g., African-American survivors and patients with multiple comorbidities) and the psychosocial profiles that motivate requests for more extensive follow-up guidance.