Chromatin and transcription factor profiling in rare stem cell populations using CUT&Tag.

Muscle stem cells (MuSCs) are a rare stem cell population that provides myofibers with a remarkable capacity to regenerate after tissue injury. Here, we have adapted the Cleavage Under Target and Tagmentation technology to the mapping of the chromatin landscape and transcription factor binding in 50,000 activated MuSCs isolated from injured mouse hindlimb muscles. We have applied this same approach to human CD34+ hematopoietic stem and progenitor cells. This protocol could be adapted to any rare stem cell population. For complete details on the use and execution of this protocol, please refer to Robinson et al. (2021).

The Bromodomains of the mammalian SWI/SNF (mSWI/SNF) ATPases Brahma (BRM) and Brahma Related Gene 1 (BRG1) promote chromatin interaction and are critical for skeletal muscle differentiation.

Skeletal muscle regeneration is mediated by myoblasts that undergo epigenomic changes to establish the gene expression program of differentiated myofibers. mSWI/SNF chromatin remodeling enzymes coordinate with lineage-determining transcription factors to establish the epigenome of differentiated myofibers. Bromodomains bind to acetylated lysines on histone N-terminal tails and other proteins. The mutually exclusive ATPases of mSWI/SNF complexes, BRG1 and BRM, contain bromodomains with undefined functional importance in skeletal muscle differentiation. Pharmacological inhibition of mSWI/SNF bromodomain function using the small molecule PFI-3 reduced differentiation in cell culture and in vivo through decreased myogenic gene expression, while increasing cell cycle-related gene expression and the number of cells remaining in the cell cycle. Comparative gene expression analysis with data from myoblasts depleted of BRG1 or BRM showed that bromodomain function was required for a subset of BRG1- and BRM-dependent gene expression. Reduced binding of BRG1 and BRM after PFI-3 treatment showed that the bromodomain is required for stable chromatin binding at target gene promoters to alter gene expression. Our findings demonstrate that mSWI/SNF ATPase bromodomains permit stable binding of the mSWI/SNF ATPases to promoters required for cell cycle exit and establishment of muscle-specific gene expression.

Negative elongation factor regulates muscle progenitor expansion for efficient myofiber repair and stem cell pool repopulation.

Negative elongation factor (NELF) is a critical transcriptional regulator that stabilizes paused RNA polymerase to permit rapid gene expression changes in response to environmental cues. Although NELF is essential for embryonic development, its role in adult stem cells remains unclear. In this study, through a muscle-stem-cell-specific deletion, we showed that NELF is required for efficient muscle regeneration and stem cell pool replenishment. In mechanistic studies using PRO-seq, single-cell trajectory analyses and myofiber cultures revealed that NELF works at a specific stage of regeneration whereby it modulates p53 signaling to permit massive expansion of muscle progenitors. Strikingly, transplantation experiments indicated that these progenitors are also necessary for stem cell pool repopulation, implying that they are able to return to quiescence. Thus, we identified a critical role for NELF in the expansion of muscle progenitors in response to injury and revealed that progenitors returning to quiescence are major contributors to the stem cell pool repopulation.

Epigenetic Regulation of Adult Myogenesis.

Skeletal muscle regeneration is an efficient stem cell-based repair system that ensures healthy musculature. For this repair system to function continuously throughout life, muscle stem cells must contribute to the process of myofiber repair as well as repopulation of the stem cell niche. The decision made by the muscle stem cells to commit to the muscle repair or to remain a stem cell depends upon patterns of gene expression, a process regulated at the epigenetic level. Indeed, it is well accepted that dynamic changes in epigenetic landscapes to control DNA accessibility and expression is a critical component during myogenesis for the effective repair of damaged muscle. Changes in the epigenetic landscape are governed by various posttranslational histone tail modifications, nucleosome repositioning, and DNA methylation events which collectively allow the control of changes in transcription networks during transitions of satellite cells from a dormant quiescent state toward terminal differentiation. This chapter focuses upon the specific epigenetic changes that occur during muscle stem cell-mediated regeneration to ensure myofiber repair and continuity of the stem cell compartment. Furthermore, we explore open questions in the field that are expected to be important areas of exploration as we move toward a more thorough understanding of the epigenetic mechanism regulating muscle regeneration.

An analysis of the use of Pharmacy Curriculum Outcomes Assessment (PCOA) scores within one professional program.

OBJECTIVE: The Pharmacy Curriculum Outcomes Assessment (PCOA) is a recent assessment requirement for US pharmacy professional programs. This study analyses PCOA scores for uses described in the 2016 Standards with data from one professional program. METHODS: PCOA data were analyzed for two consecutive classes (n=215) of pharmacy students at the end of their didactic curriculum to explore relationships among PCOA scores, grade point average (GPA), and North American Pharmacist Licensure Examination (NAPLEX) scores utilizing regression analyses. RESULTS: Decisions about student learning based on PCOA scores and GPA indicated remediation would have been prescribed for approximately 7% of students. In comparison, NAPLEX scores revealed a 1% failure rate among the study sample. Relationships between PCOA scores and GPA (r=0.47) and NAPLEX (r=0.51) were moderate to large, respectively. GPA explained a larger portion of unique variance (14%) than PCOA (8%) in NAPLEX scores. CONCLUSIONS: In this sample of students, academic decisions would have varied depending upon the learning assessment, which is consistent with a moderate correlation between GPA and PCOA scores. Although PCOA scores correlate with GPA and NAPLEX, PCOA scores explained a smaller portion of unique variance in NAPLEX scores than GPA. The ongoing establishment of validity evidence of PCOA scores is important for meaningful interpretation of scores for the intended uses.

Educating for safety in the pharmacy curriculum.

This overview of the Educating for Safety supplement issue explores the context and urgency of the problem of unsafe care, what we have learned about improving both safety and quality in health care, and the implications of this for educators. This supplement issue is a response to the charge of the AACP Council of Deans (COD) and the Council of Faculties (COF) Medication Safety Task Force to address the role of colleges and schools of pharmacy in responding to the national patient safety agenda. The articles included are intended to serve as a nexus for pharmacy education in developing curricula and promoting best practices as they relate to the importance of medication safety.

Unleashing the potential: women's development and ways of knowing as a perspective for veterinary medical education.

Women now dominate student enrollment in colleges of veterinary medicine in the USA, as well as in other countries. Projections indicate that this will remain a constant. The implications for teaching, learning, mentoring, leadership, professional development, student and faculty diversity, and curriculum structure are enormous. This article provides the groundwork for examining gender diversity in veterinary medical education. Women's development and ways of knowing are identified as paramount for understanding and benefiting students and faculty in their higher education experiences and in their professional lives. Seminal research focusing on women's development and ways of knowing is introduced, summarized, and contrasted to male-centered models, and implications for teaching practice are considered. Our underlying premise is that research about women's moral and intellectual development is relevant to veterinary education and supports the adoption of student-centered approaches to teaching and learning.