Value of Schwartz Rounds in promoting the emotional well-being of healthcare workers: a qualitative study.

OBJECTIVES: Schwartz Rounds are forums that enable healthcare staff to reflect on emotional and social dimensions of their work. In this study, we aimed to explore the experiences of Schwartz Rounds on emotional aspects of care and practice within a clinical environment. DESIGN: Using qualitative methods, we interviewed participants individually and in focus groups. Interviews were recorded, transcribed and analysed by thematic analysis. SETTING: The study was based at a public health service Te Whatu Ora Counties Manukau in Auckland, New Zealand's largest, most ethnically diverse population. PARTICIPANTS: Participants were panellists who took part in successive Schwartz Rounds over a 10-month period. There were 17 participants with a range of experience (1-30 years) and occupations including clinical, allied, technical and administrative staff from medical specialties of plastic surgery, pain services, emergency medicine, intensive care, organ donation services, COVID-19 response and palliative care services. RESULTS: Three themes were identified: the need for emotional processing, valuing guided reflection and realising our humanity. The third theme 'realising our humanity' comprised altruism, connection and compassion. Schwartz Rounds provided staff with clear benefits: emotionally resonant experiences within an environment of psychological safety and connection to the wider organisation. The daunting nature of emotional disclosure was mitigated by a supportive audience. CONCLUSION: There is an organisational imperative to ensure that staff have opportunities to process intense emotions associated with healthcare work. Schwartz Rounds are one means to attend to the emotional welfare of healthcare staff, enabling them to gain different perspectives in the care of their patients and colleagues within system constraints.

Risk factors for perioperative adverse events in children with myotonic dystrophy.

BACKGROUND: This study was conducted to identify patient-related, surgical, and anesthetic factors that would help predict adverse events and allow for better planning of perioperative care in children with myotonic dystrophy. METHODS: This is a retrospective chart review from a large tertiary pediatric hospital. Data were collected on demographics, disease severity, surgical procedure, and anesthetic technique. Perioperative adverse events were recorded. RESULTS: Records on 27 patients having 78 anesthetics over a 17.5-year period were reviewed. The overall frequency of postoperative respiratory complications was 10%. Significant risk factors were high muscular impairment rating scale (MIRS) grade (P = 0.007), at least 2300 cytosine, thymine, guanine (CTG) repeats on the protein kinase gene of chromosome 19q (P = 0.009), a longer duration of surgery (RR = 14.0 for surgery lasting at least 1 h; P = 0.002), perioperative morphine use (RR = 7.7, 95% CI 2.2-12.8; P = 0.005), intubation (P = 0.02), and the use of muscle relaxant without reversal (RR = 15.5, P = 0.0002). Using a multivariate risk model, only MIRS grade and the use of muscle relaxant without reversal were shown to be significant independent risk factors (RR = 24.9, P < 0.0001). CONCLUSIONS: The MIRS is a statistically significant and clinically useful tool for predicting high perioperative risk. Patients with a high MIRS grade should therefore be considered for postoperative intensive care. The use of muscle relaxant without reversal was also shown to be a significant risk factor. Patients who require morphine infusions postoperatively might also be most safely managed in a high dependency unit.

Bone morphogenetic protein 4 modulates c-Kit expression and differentiation potential in murine embryonic aorta-gonad-mesonephros haematopoiesis in vitro.

The transforming growth factor-beta-related factor bone morphogenetic protein 4 (BMP4) is expressed in the human embryonic aorta-gonad-mesonephros (AGM) coincident with the emergence of haematopoietic cells and influences postnatal mammalian haematopoietic stem cells in vitro. To investigate the role of BMP4 in mammalian embryonic haematopoiesis, cells were isolated from murine AGM and two populations of CD34(+) cells with different levels of c-Kit expression and multipotency were identified. CD34(+)/c-Kit(high) cells express CD45 and are haematopoietic-restricted progenitors. In contrast, CD34(+)/c-Kit(low) cells are Flk1+/CD45(neg) and generate adherent colonies in ex vivo culture that resemble haemangioblast colonies identified in other systems. The addition of BMP4 to AGM cells resulted in expansion of the CD34(+)/c-Kit(low) cell pool within 48 h, via a combination of down modulation of the c-Kit receptor in CD34(+)/c-Kit(high) cells and proliferation. In long-term culture, BMP4 increased the growth/survival of CD34(+)/c-Kit(high) haematopoietic progenitors, effects that were blocked by BMP inhibitors. CD34(+)/c-Kit(high) progenitors cultured with BMP4 also generated adherent colonies typical of c-Kit(low) cells. These results suggest that BMP4 regulates c-Kit expression and differentiation potential in CD34(+) AGM cells and supports a role for BMP signalling in the maintenance of multipotency during embryonic haematopoiesis, providing an insight into stem cell homeostasis within the mammalian haematopoietic niche.

Gammaretrovirus-mediated correction of SCID-X1 is associated with skewed vector integration site distribution in vivo.

We treated 10 children with X-linked SCID (SCID-X1) using gammaretrovirus-mediated gene transfer. Those with sufficient follow-up were found to have recovered substantial immunity in the absence of any serious adverse events up to 5 years after treatment. To determine the influence of vector integration on lymphoid reconstitution, we compared retroviral integration sites (RISs) from peripheral blood CD3(+) T lymphocytes of 5 patients taken between 9 and 30 months after transplantation with transduced CD34(+) progenitor cells derived from 1 further patient and 1 healthy donor. Integration occurred preferentially in gene regions on either side of transcription start sites, was clustered, and correlated with the expression level in CD34(+) progenitors during transduction. In contrast to those in CD34(+) cells, RISs recovered from engrafted CD3(+) T cells were significantly overrepresented within or near genes encoding proteins with kinase or transferase activity or involved in phosphorus metabolism. Although gross patterns of gene expression were unchanged in transduced cells, the divergence of RIS target frequency between transduced progenitor cells and post-thymic T lymphocytes indicates that vector integration influences cell survival, engraftment, or proliferation.

Lentiviral vectors containing an enhancer-less ubiquitously acting chromatin opening element (UCOE) provide highly reproducible and stable transgene expression in hematopoietic cells.

Ubiquitously acting chromatin opening elements (UCOEs) consist of methylation-free CpG islands encompassing dual divergently transcribed promoters of housekeeping genes that have been shown to confer resistance to transcriptional silencing and to produce consistent and stable transgene expression in tissue culture systems. To develop improved strategies for hematopoietic cell gene therapy, we have assessed the potential of the novel human HNRPA2B1-CBX3 UCOE (A2UCOE) within the context of a self-inactivating (SIN) lentiviral vector. Unlike viral promoters, the enhancer-less A2UCOE gave rise to populations of cells that expressed a reporter transgene at a highly reproducible level. The efficiency of expression per vector genome was also markedly increased in vivo compared with vectors incorporating either spleen focus-forming virus (SFFV) or cytomegalovirus (CMV) promoters, suggesting a relative resistance to silencing. Furthermore, an A2UCOE-IL2RG vector fully restored the IL-2 signaling pathway within IL2RG-deficient human cells in vitro and successfully rescued the X-linked severe combined immunodeficiency (SCID-X1) phenotype in a mouse model of this disease. These data indicate that the A2UCOE displays highly reliable transcriptional activity within a lentiviral vector, largely overcoming insertion-site position effects and giving rise to therapeutically relevant levels of gene expression. These properties are achieved in the absence of classic enhancer activity and therefore may confer a high safety profile.

Capture and generation of adenovirus specific T cells for adoptive immunotherapy.

Adenoviral infections represent a major cause of morbidity and mortality following haematopoietic stem cell transplantation. Current anti-viral agents are virostatic and it is evident that elimination of adenovirus (ADV) infection is only achieved by recovery of cellular immunity. Using an interferon-gamma (IFN-gamma) secretion and capture assay to isolate ADV-specific T cells, followed by a 2 week expansion and restimulation protocol, we generated ADV T cells that may be used for cellular immunotherapy. In contrast to virus-specific T cells for cytomegalovirus or Epstein-Barr virus, the ADV response was dominated by CD4(+) T cells and the majority of captured cells exhibited an effector/memory immunophenotype. Highly specific antigen responses were demonstrated by intracellular IFN-gamma expression and cytotoxicity assays when the expanded cells underwent restimulation with ADV-pulsed target cells. Although T cells were initially generated in response to ADV species C, the expanded populations also showed strong activity against ADV species B, suggesting cross-reactivity across ADV species; a finding that has important clinical consequences in the paediatric setting, where the majority of infections are caused by ADV type B and C. The protocols can be readily translated to generate ADV-specific T cells suitable for clinical use and offer an effective immunotherapeutic strategy to control ADV infection.

Successful reconstitution of immunity in ADA-SCID by stem cell gene therapy following cessation of PEG-ADA and use of mild preconditioning.

Gene therapy is a promising treatment option for monogenic diseases, but success has been seen in only a handful of studies thus far. We now document successful reconstitution of immune function in a child with the adenosine deaminase (ADA)-deficient form of severe combined immunodeficiency (SCID) following hematopoietic stem cell (HSC) gene therapy. An ADA-SCID child who showed a poor response to PEG-ADA enzyme replacement was enrolled into the clinical study. Following cessation of enzyme replacement therapy, autologous CD34(+) HSCs were transduced with an ADA-expressing gammaretroviral vector. Gene-modified cells were reinfused following one dose of preconditioning chemotherapy. Two years after the procedure, immunological and biochemical correction has been maintained with progressive increase in lymphocyte numbers, reinitiation of thymopoiesis, and systemic detoxification of ADA metabolites. Sustained vector marking with detection of polyclonal vector integration sites in multiple cell lineages and detection of ADA activity in red blood cells suggests transduction of early hematopoietic progenitors. No serious side effects were seen either as a result of the conditioning procedure or due to retroviral insertion. Gene therapy is an effective treatment option for the treatment of ADA-SCID.

Failure of SCID-X1 gene therapy in older patients.

Gene therapy has been shown to be a highly effective treatment for infants with typical X-linked severe combined immunodeficiency (SCID-X1, gammac-deficiency). For patients in whom previous allogeneic transplantation has failed, and others with attenuated disease who may present later in life, the optimal treatment strategy in the absence of human leukocyte antigen (HLA)-matched donors is unclear. Here we report the failure of gene therapy in 2 such patients, despite effective gene transfer to bone marrow CD34(+) cells, suggesting that there are intrinsic host-dependent restrictions to efficacy. In particular, there is likely to be a limitation to initiation of normal thymopoiesis, and we therefore suggest that intervention for these patients should be considered as early as possible.

Impaired dendritic-cell homing in vivo in the absence of Wiskott-Aldrich syndrome protein.

Regulated migration and spatial localization of dendritic cells (DCs) are critical events during the initiation of physiologic immune responses and maintenance of tolerance. Here we have used cells deficient in the Wiskott-Aldrich syndrome protein (WASp) to demonstrate the importance of dynamic remodeling of the actin cytoskeleton for these trafficking processes to occur in vitro and in vivo. On fibronectin-coated surfaces, WASp-null immature murine DCs exhibited defects both of attachment and detachment, resulting in impaired net translocation compared with normal cells. The chemokinetic response to CCL21, which is critical for normal lymphatic trafficking, was also abrogated in the absence of WASp. In vivo in both fluorescein isothiocyanate (FITC) and oxazolone contact hypersensitivity models, WASp-null Langerhans cell (LC) migration was compromised, as judged by exit from the skin as well as by homing to the draining lymph node (LN). Furthermore, following systemic challenge with lipopolysaccharide (LPS) or toxoplasma-derived antigen, WASp-null DCs showed incomplete redistribution to T-cell areas in the spleen. Instead, they were retained ectopically in the marginal zone. DC trafficking in vivo is therefore dependent on a normally regulated actin cytoskeleton, which performs an essential function during maintenance of physiologic immunity and when disturbed may contribute significantly to the immunopathology of Wiskott-Aldrich Syndrome.

Gene therapy of X-linked severe combined immunodeficiency by use of a pseudotyped gammaretroviral vector.

BACKGROUND: X-linked severe combined immunodeficiency (SCID-X1) is caused by mutations in the common cytokine-receptor gamma chain (gamma(c)), resulting in disruption of development of T lymphocytes and natural-killer cells. B-lymphocyte function is also intrinsically compromised. Allogeneic bone-marrow transplantation is successful if HLA-matched family donors are available, but HLA-mismatched procedures are associated with substantial morbidity and mortality. We investigated the application of somatic gene therapy by use of a gibbon-ape-leukaemia-virus pseudotyped gammaretroviral vector. METHODS: Four children with SCID-X1 were enrolled. Autologous CD34-positive haemopoietic bone-marrow stem cells were transduced ex vivo and returned to the patients without preceding cytoreductive chemotherapy. The patients were monitored for integration and expression of the gamma(c) vector and for functional immunological recovery. FINDINGS: All patients have shown substantial improvements in clinical and immunological features, and prophylactic medication could be withdrawn in two. No serious adverse events have been recorded. T cells responded normally to mitogenic and antigenic stimuli, and the T-cell-receptor (TCR) repertoire was highly diverse. Where assessable, humoral immunity, in terms of antibody production, was also restored and associated with increasing rates of somatic mutation in immunoglobulin genes. INTERPRETATION: Gene therapy for SCID-X1 is a highly effective strategy for restoration of functional cellular and humoral immunity.

SAP mediates specific cytotoxic T-cell functions in X-linked lymphoproliferative disease.

Cytotoxic T cells (CTLs) and natural killer cells play a major role in the immune response to Epstein-Barr virus (EBV) infection. In X-linked lymphoproliferative (XLP) disease, a severe immunodeficiency, immunodysregulatory phenomena are observed following EBV infection, suggesting that defects exist in these effector populations. The gene defective in XLP is SAP (signaling lymphocytic activation molecule [SLAM]-associated protein), an adaptor protein that mediates signals through SLAM and other immunoglobulin superfamily receptors including 2B4. We generated EBV-specific T-cell lines from controls and XLP patients and examined CTL function in response to different stimuli. We show that XLP patients can generate EBV-T-cell lines that are phenotypically similar to those from controls. XLP patient EBV-T-cell lines showed a significant decrease in interferon-gamma (IFN-gamma) production in response to 2B4 and autologous EBV-transformed lymphoblastoid cell line (LCL) stimulation but not in response to SLAM. Furthermore, XLP EBV-T-cell lines demonstrated markedly decreased cytotoxic activity against autologous LCLs. By retroviral gene transfer of the SAP gene into XLP EBV-T-cell lines, we show reconstitution of IFN-gamma production and of cytotoxic activity confirming SAP-dependent defects. These studies demonstrate that in XLP the lack of SAP affects specific signaling pathways resulting in severe disruption of CTL function.