A Myc-dependent division timer complements a cell-death timer to regulate T cell and B cell responses.

T lymphocytes and B lymphocytes integrate activating signals to control the size of their proliferative response. Here we report that such control was achieved by timed changes in the production rate of cell-cycle-regulating proto-oncoprotein Myc, with division cessation occurring when Myc levels fell below a critical threshold. The changing pattern of the level of Myc was not affected by cell division, which identified the regulating mechanism as a cell-intrinsic, heritable temporal controller. Overexpression of Myc in stimulated T cells and B cells did not sustain cell proliferation indefinitely, as a separate 'time-to-die' mechanism, also heritable, was programmed after lymphocyte activation and led to eventual cell loss. Together the two competing cell-intrinsic timed fates created the canonical T cell and B cell immune-response pattern of rapid growth followed by loss of most cells. Furthermore, small changes in these timed processes by regulatory signals, or by oncogenic transformation, acted in synergy to greatly enhance cell numbers over time.

Class-Switch Recombination Occurs Infrequently in Germinal Centers.

Class-switch recombination (CSR) is a DNA recombination process that replaces the immunoglobulin (Ig) constant region for the isotype that can best protect against the pathogen. Dysregulation of CSR can cause self-reactive BCRs and B cell lymphomas; understanding the timing and location of CSR is therefore important. Although CSR commences upon T cell priming, it is generally considered a hallmark of germinal centers (GCs). Here, we have used multiple approaches to show that CSR is triggered prior to differentiation into GC B cells or plasmablasts and is greatly diminished in GCs. Despite finding a small percentage of GC B cells expressing germline transcripts, phylogenetic trees of GC BCRs from secondary lymphoid organs revealed that the vast majority of CSR events occurred prior to the onset of somatic hypermutation. As such, we have demonstrated the existence of IgM-dominated GCs, which are unlikely to occur under the assumption of ongoing switching.

Transcription Factor PU.1 Promotes Conventional Dendritic Cell Identity and Function via Induction of Transcriptional Regulator DC-SCRIPT.

Dendritic cells (DCs) are can be broadly divided into conventional (cDC) and plasmacytoid (pDC) subsets. Despite the importance of this lineage diversity, its genetic basis is not fully understood. We found that conditional ablation of the Ets-family transcription factor PU.1 in DC-restricted progenitors led to increased pDC production at the expense of cDCs. PU.1 controlled many of the cardinal functions of DCs, such as antigen presentation by cDCs and type I interferon production by pDCs. Conditional ablation of PU.1 de-repressed the pDC transcriptional signature in cDCs. The combination of genome-wide mapping of PU.1 binding and gene expression analysis revealed a key role for PU.1 in maintaining cDC identity through the induction of the transcriptional regulator DC-SCRIPT. PU.1 activated DC-SCRIPT expression, which in turn promoted cDC formation, particularly of cDC1s, and repressed pDC development. Thus, cDC identity is regulated by a transcriptional node requiring PU.1 and DC-SCRIPT.

Transcriptional profiling of mouse B cell terminal differentiation defines a signature for antibody-secreting plasma cells.

When B cells encounter an antigen, they alter their physiological state and anatomical localization and initiate a differentiation process that ultimately produces antibody-secreting cells (ASCs). We have defined the transcriptomes of many mature B cell populations and stages of plasma cell differentiation in mice. We provide a molecular signature of ASCs that highlights the stark transcriptional divide between B cells and plasma cells and enables the demarcation of ASCs on the basis of location and maturity. Changes in gene expression correlated with cell-division history and the acquisition of permissive histone modifications, and they included many regulators that had not been previously implicated in B cell differentiation. These findings both highlight and expand the core program that guides B cell terminal differentiation and the production of antibodies.

Long-Lived Plasma Cells Have a Sweet Tooth.

Long-lived plasma cells (LLPCs) are durable antibody-producing cells that are key to immunity. Bhattacharya and colleagues find that LLPCs derive their enhanced survival capacity from a higher rate of glucose import. Some of this glucose sustains the cells through glycolysis, while the bulk is required for antibody glycosylation.

The Helix-Loop-Helix Protein ID2 Governs NK Cell Fate by Tuning Their Sensitivity to Interleukin-15.

The inhibitor of DNA binding 2 (Id2) is essential for natural killer (NK) cell development with its canonical role being to antagonize E-protein function and alternate lineage fate. Here we have identified a key role for Id2 in regulating interleukin-15 (IL-15) receptor signaling and homeostasis of NK cells by repressing multiple E-protein target genes including Socs3. Id2 deletion in mature NK cells was incompatible with their homeostasis due to impaired IL-15 receptor signaling and metabolic function and this could be rescued by strong IL-15 receptor stimulation or genetic ablation of Socs3. During NK cell maturation, we observed an inverse correlation between E-protein target genes and Id2. These results shift the current paradigm on the role of ID2, indicating that it is required not only to antagonize E-proteins during NK cell commitment, but constantly required to titrate E-protein activity to regulate NK cell fitness and responsiveness to IL-15.

Transgender and gender diverse curriculum in medical imaging programs: a case study.

BACKGROUND: Transgender and gender diverse (TGD) individuals face barriers, including harassment and discrimination, when accessing healthcare services. Medical imaging procedures require personal information to be shared, such as date of last menstrual cycle and/or pregnancy status; some imaging exams are also invasive or intimate in nature. Terminology is based on binary sex creating an inherently cis-heteronormative environment. TGD patients fear being outed and often feel a need to function as educators and advocates for their care. Incorporation of inclusive healthcare curriculum related to TGD populations is an effective means of educating new health providers and promotes safer and more inclusive spaces in healthcare settings. Educators face barriers which hinder the creation and implementation of TGD content. The purpose of this study was to examine the impacts educators are faced with when creating and delivering TGD content in their medical imaging curriculum. METHODS: A case study of medical imaging programs at a Canadian post-secondary institute was undertaken. Data was collected via semi-structured interviews with faculty. Relevant institutional documents such as strategic plans, policies/procedures, websites, and competency profiles were accessed. Framework analysis was used to analyze the data. RESULTS: The study found seven themes that influence the development of TGD curriculum as follows: familiarity and comfort with the curriculum and content change process; collaboration with other healthcare programs; teaching expertise; management of course workload and related. duties; connections to the TGD community; knowledge of required TGD content and existing gaps in curriculum; and access to supports. CONCLUSIONS: Understanding educators' perspectives can lead to an increased sense of empowerment for them to create and incorporate TGD curriculum in the future. Many post- secondary institutions are incorporating an inclusive lens to educational plans; this research can be used in future curriculum design projects. The goal is improved medical imaging experiences for the TGD population.

Spirituality in Addiction Recovery: A Narrative Review.

In the area of addiction, Canada has been in a public health crisis since 2016. Addiction takes a toll on an individual's self-worth and identity. In this narrative literature review, the distinct nature of spirituality was addressed. Next, individualized conceptualizations of spirituality were outlined. Subsequently, the importance of fellowship in addiction recovery was detailed. Next, the significance of being of service was presented. Meaningful and authentic spirituality were discussed in the context of recovery identity. Lastly, spirituality as a personal journey is described. A narrative literature review of 70 manuscripts published between 1999 and 2021 was undertaken to determine multiple approaches to treating addiction recovery in the context of spiritual development. An understanding of spirituality can inform counsellors regarding spiritual development in addiction recovery. Implications for counselling include a roadmap to support clients developing an individualized spiritual connection and operating as a functional system.

A qualitative study examining newly diagnosed breast cancer patients' experiences of participating in the Alberta Moving Beyond Breast Cancer (AMBER) prospective cohort study.

BACKGROUND: Decisions to participate in cancer trials are associated with uncertainty, distress, wanting to help find a cure, the hope for benefit, and altruism. There is a gap in the literature regarding research examining participation in prospective cohort studies. The aim of this study was to examine the experiences of newly diagnosed women with breast cancer participating in the AMBER Study to identify potential strategies to support patients' recruitment, retention, and motivation. METHODS: Newly diagnosed breast cancer patients were recruited from the Alberta Moving Beyond Breast Cancer (AMBER) cohort study. Data were collected using semi-structured conversational interviews with 21 participants from February to May 2020. Transcripts were imported into NVivo software for management, organization, and coding. Inductive content analysis was undertaken. RESULTS: Five main concepts associated with recruitment, retention, and motivation to participate were identified. These main concepts included: (1) personal interest in exercise and nutrition; (2) investment in individual results; (3) personal and professional interest in research; (4) burden of assessments; (5) importance of research staff. CONCLUSIONS: Breast cancer survivors participating in this prospective cohort study had numerous reasons for participating and these reasons could be considered in future studies to enhance participant recruitment and retention. Improving recruitment and retention in prospective cancer cohort studies could result in more valid and generalizable study findings that could improve the care of cancer survivors.

Lessons learned from clinical course design in the pandemic: Pedagogical implications from a qualitative analysis.

AIMS: The purpose of this study was to examine clinical pedagogy based on experiences of changes and adaptations to clinical courses that occurred in nursing education during the pandemic. Beyond learning how to manage nursing education during a pandemic or other crisis, we uncover the lessons to be learned for overall improvement of nursing education. DESIGN: Qualitative descriptive analysis using semi-structured interview data with baccalaureate nursing students. METHODS: Data were collected in the spring of 2021 using semi-structured interview with 15 participants. Transcribed text was analysed using thematic content analysis. The COREQ checklist was used to guide our reporting. RESULTS: Three themes were identified related to course design in clinical courses for nursing students: the role and limitations of simulation, competency evaluations and career implications. Students expressed some concern over not 'finishing hours', loss of in-person clinical experiences and their reduced exposure to different clinical settings. CONCLUSION: To prepare work-ready nurses, educators need to keep in mind the trends, issues and demands of future healthcare systems. Simulation may have been a temporary measure to achieve clinical competence during the pandemic but needs to be of high-quality and cannot meet all the expected learning outcomes of clinical courses. Exposure to different patients, families and communities will ensure that the future nursing workforce has experience, socialization, competence, and desire to work in various clinical settings. Competency evaluation similarly needs to be robust and objective and consider the role and perception of hours completed. PATIENT OR PUBLIC CONTRIBUTION: No patient or public contribution. Participants were nursing students.

The philosophy of Hans-Georg Gadamer: An exemplar of the complicated relationship between philosophy and nursing practice.

Philosophy has a complicated relationship with nursing practice. Selected concepts from Hans-Georg Gadamer's Truth and Method specifically prejudice, conversation, and language are articulated. An exemplar involving nursing practice at an outpatient clinic for women seeking pre- and postbreast cancer care is offered to explicate these concepts. We considered the fit of Gadamer's philosophy, particularly the concept of conversation, within a public health nursing practice context in home and community settings of the client/family and offered tentative conclusions. To extend the discussion of the relationship between philosophy and nursing practice, we posed questions developed to provide deeper insight into this complicated relationship.

Two of a kind: transmissible Schwann cell cancers in the endangered Tasmanian devil (Sarcophilus harrisii).

Devil facial tumour disease (DFTD) comprises two genetically distinct transmissible cancers (DFT1 and DFT2) endangering the survival of the Tasmanian devil (Sarcophilus harrisii) in the wild. DFT1 first arose from a cell of the Schwann cell lineage; however, the tissue-of-origin of the recently discovered DFT2 cancer is unknown. In this study, we compared the transcriptome and proteome of DFT2 tumours to DFT1 and normal Tasmanian devil tissues to determine the tissue-of-origin of the DFT2 cancer. Our findings demonstrate that DFT2 expresses a range of Schwann cell markers and exhibits expression patterns consistent with a similar origin to the DFT1 cancer. Furthermore, DFT2 cells express genes associated with the repair response to peripheral nerve damage. These findings suggest that devils may be predisposed to transmissible cancers of Schwann cell origin. The combined effect of factors such as frequent nerve damage from biting, Schwann cell plasticity and low genetic diversity may allow these cancers to develop on rare occasions. The emergence of two independent transmissible cancers from the same tissue in the Tasmanian devil presents an unprecedented opportunity to gain insight into cancer development, evolution and immune evasion in mammalian species.

Haemopedia RNA-seq: a database of gene expression during haematopoiesis in mice and humans.

During haematopoiesis, haematopoietic stem cells differentiate into restricted potential progenitors before maturing into the many lineages required for oxygen transport, wound healing and immune response. We have updated Haemopedia, a database of gene-expression profiles from a broad spectrum of haematopoietic cells, to include RNA-seq gene-expression data from both mice and humans. The Haemopedia RNA-seq data set covers a wide range of lineages and progenitors, with 57 mouse blood cell types (flow sorted populations from healthy mice) and 12 human blood cell types. This data set has been made accessible for exploration and analysis, to researchers and clinicians with limited bioinformatics experience, on our online portal Haemosphere: https://www.haemosphere.org. Haemosphere also includes nine other publicly available high-quality data sets relevant to haematopoiesis. We have added the ability to compare gene expression across data sets and species by curating data sets with shared lineage designations or to view expression gene vs gene, with all plots available for download by the user.

The Work of Preparing Canadian Nurses for A Licensure Exam Originating from the USA: A Nurse Educator's Journey into the Institutional Organization of the NCLEX-RN.

The licensing exam for registered nurses in Canada has recently been changed from a Canadian developed, owned and delivered exam to the National Council Licensure Examination for Registered Nurses (NCLEX-RN) which originates from the United States. Rationale for this exam change focused on transitioning to a computer-based exam that has increased writing dates, with increased security, validated psychometrics, increased exam result delivery, and an anticipated decrease in expense to students. Concerns have arisen around the acceptance, implementation and delivery of this exam to Canadian nursing students that reflects the broad Canadian landscape of education and nursing practice. The experience of a Canadian nurse educator in working to facilitate students' transition to this exam is addressed using an institutional ethnographic lens. Finally, we come to conclusions about the importance of countries utilizing licensing exams that reflect their nursing education and practice.

Mining the Plasma Cell Transcriptome for Novel Cell Surface Proteins.

Antibody Secreting Cells (ASCs) are a fundamental component of humoral immunity, however, deregulated or excessive antibody production contributes to the pathology of autoimmune diseases, while transformation of ASCs results in the malignancy Multiple Myeloma (MM). Despite substantial recent improvements in treating these conditions, there is as yet no widely used ASC-specific therapeutic approach, highlighting a critical need to identify novel methods of targeting normal and malignant ASCs. Surface molecules specifically expressed by the target cell population represent ideal candidates for a monoclonal antibody-based therapy. By interrogating the ASC gene signature that we previously defined we identified three surface proteins, Plpp5, Clptm1l and Itm2c, which represent potential targets for novel MM treatments. Plpp5, Clptm1l and Itm2c are highly and selectively expressed by mouse and human ASCs as well as MM cells. To investigate the function of these proteins within the humoral immune system we have generated three novel mouse strains, each carrying a loss-of-function mutation in either Plpp5, Clptm1l or Itm2c. Through analysis of these novel strains, we have shown that Plpp5, Clptm1l and Itm2c are dispensable for the development, maturation and differentiation of B-lymphocytes, and for the production of antibodies by ASCs. As adult mice lacking either protein showed no apparent disease phenotypes, it is likely that targeting these molecules on ASCs will have minimal on-target adverse effects.

IL4 and IL21 cooperate to induce the high Bcl6 protein level required for germinal center formation.

Bcl6 (B-cell lymphoma 6) is a transcriptional repressor and critical mediator of the germinal center reaction during a T-cell-dependent antibody response, where it enables somatic hypermutation of immunoglobulin genes and inhibits terminal differentiation via repression of Blimp1. It can also contribute to the development of diffuse large B-cell lymphoma when expressed inappropriately. Bcl6 regulation is mediated both at the transcriptional and post-transcriptional levels, and in particular a strong signal through the B-cell receptor causes rapid proteasomal degradation of Bcl6. Despite the importance of Bcl6 in both immunity and cancer, little is known about how other extrinsic factors regulate Bcl6 in B cells. Here we show that Bcl6 is indeed highly unstable in B cells after a B-cell receptor (BCR) signal, but that the T-cell-derived cytokines interleukin 4 (IL4) and IL21 counteract BCR-mediated degradation, preserving Bcl6 protein levels. Stat6, downstream of IL4, can induce Bcl6 transcription directly. In vivo, B-cell intrinsic loss of IL4 or IL21 signaling reduces the magnitude or duration of the GC response, respectively, while their combined loss almost completely eliminates the GC response. This work provides key insights into the effect mediated by T-follicular helper cytokines on Bcl6 regulation.

Immunology of a Transmissible Cancer Spreading among Tasmanian Devils.

Devil facial tumor disease (DFTD) is a transmissible cancer that has killed most of the Tasmanian devil (Sarcophilus harrissii) population. Since the first case appeared in the mid-1990s, it has spread relentlessly across the Tasmanian devil's geographic range. As Tasmanian devils only exist in Tasmania, Australia, DFTD has the potential to cause extinction of this species. The origin of DFTD was a Schwann cell from a female devil. The disease is transmitted when devils bite each other around the facial areas, a behavior synonymous with this species. Every devil that is 'infected' with DFTD dies from the cancer. Once the DFTD cells have been transmitted, they appear to develop into a cancer without inducing an immune response. The DFTD cancer cells avoid allogeneic recognition because they do not express MHC class I molecules on the cell surface. A reduced genetic diversity and the production of immunosuppressive cytokines may also contribute.

A high throughput embryonic stem cell screen identifies Oct-2 as a bifunctional regulator of neuronal differentiation.

Neuronal differentiation is a complex process that involves a plethora of regulatory steps. To identify transcription factors that influence neuronal differentiation we developed a high throughput screen using embryonic stem (ES) cells. Seven-hundred human transcription factor clones were stably introduced into mouse ES (mES) cells and screened for their ability to induce neuronal differentiation of mES cells. Twenty-four factors that are capable of inducing neuronal differentiation were identified, including four known effectors of neuronal differentiation, 11 factors with limited evidence of involvement in regulating neuronal differentiation, and nine novel factors. One transcription factor, Oct-2, was studied in detail and found to be a bifunctional regulator: It can either repress or induce neuronal differentiation, depending on the particular isoform. Ectopic expression experiments demonstrate that isoform Oct-2.4 represses neuronal differentiation, whereas Oct-2.2 activates neuron formation. Consistent with a role in neuronal differentiation, Oct-2.2 expression is induced during differentiation, and cells depleted of Oct-2 and its homolog Oct-1 have a reduced capacity to differentiate into neurons. Our results reveal a number of transcription factors potentially important for mammalian neuronal differentiation, and indicate that Oct-2 may serve as a binary switch to repress differentiation in precursor cells and induce neuronal differentiation later during neuronal development.

Mitogen-activated Tasmanian devil blood mononuclear cells kill devil facial tumour disease cells.

Devil facial tumour disease (DFTD) is a transmissible cancer that has brought the host species, the Tasmanian devil, to the brink of extinction. The cancer cells avoid allogeneic immune recognition by downregulating cell surface major histocompatibility complex (MHC) I expression. This should prevent CD8(+) T cell, but not natural killer (NK) cell, cytotoxicity. The reason why NK cells, normally reactive to MHC-negative cells, are not activated to kill DFTD cells has not been determined. The immune response of wild devils to DFTD, if it occurs, is uncharacterised. To investigate this, we tested 12 wild devils with DFTD, and found suggestive evidence of low levels of antibodies against DFTD cells in one devil. Eight of these devils were also analysed for cytotoxicity, however, none showed evidence for cytotoxicity against cultured DFTD cells. To establish whether mimicking activation of antitumour responses could induce cytotoxic activity against DFTD, Tasmanian devil peripheral blood mononuclear cells (PBMCs) were treated with either the mitogen Concanavalin A, the Toll-like receptor agonist polyinosinic:polycytidylic acid or recombinant Tasmanian devil IL-2. All induced the PBMC cells to kill cultured DFTD cells, suggesting that activation does not occur after encounter with DFTD cells in vivo, but can be induced. The identification of agents that activate cytotoxicity against DFTD target cells is critical for developing strategies to protect against DFTD. Such agents could function as adjuvants to induce functional immune responses capable of targeting DFTD cells and tumours in vivo.

Transcription factor IRF4 regulates germinal center cell formation through a B cell-intrinsic mechanism.

In response to antigenic stimulation, mature B cells interact with follicular helper T cells in specialized structures called germinal centers (GCs), which leads to the development of memory B cells and Ab-secreting plasma cells. The transcription factor IFN regulatory factor 4 (IRF4) is essential for the formation of follicular helper T cells and thus GCs, although whether IRF4 plays a distinct role in GC B cells remains contentious. RNAseq analysis on ex vivo-derived mouse B cell populations showed that Irf4 was lowly expressed in naive B cells, highly expressed in plasma cells, but absent from GC B cells. In this study, we used conditional deletion of Irf4 in mature B cells as well as wild-type and Irf4-deficient mixed bone marrow chimeric mice to investigate how and where IRF4 plays its essential role in GC formation. Strikingly, GC formation was severely impaired in mice in which Irf4 was conditionally deleted in mature B cells, after immunization with protein Ags or infection with Leishmania major. This effect was evident as early as day 5 following immunization, before the development of GCs, indicating that Irf4 was required for the development of early GC B cells. This defect was B cell intrinsic because Irf4-deficient B cells in chimeric mice failed to participate in the GC in response to L. major or influenza virus infection. Taken together, these data demonstrate a B cell-intrinsic requirement for IRF4 for not only the development of Ab secreting plasma cells but also for GC formation.