Co-visualise the impact of Sickle Cell: How can we use design thinking to investigate and visualise the impact of Sickle Cell?

Background: This study leverages the principles of design thinking and system thinking to investigate and visualise the impact of Sickle Cell. Sickle cell is a genetic disorder that affects millions of people worldwide and disproportionately affects people of African and Caribbean descent. Patients with this condition face a range of physical, emotional, and social challenges due to the unpredictable nature of the disease. The fundamental principle explored in this study is designing for empathy. Design for Sickle Cell (D4SC) initiative was developed to bridge the gap between art, design and science within the Sickle Cell landscape. Aim: This design research project aims to investigate and visualise the condition's impact from a multi-stakeholder perspective by developing art and design prototypes that can inform an innovative Sickle Cell exhibition. Methods: Techniques from social constructionism, phenomenological qualitative research and user-experience research were utilised to create a novel methodology for this small-scale study. The methodology involved a multi-step process combining the double diamond, system thinking, and action research frameworks to gather insights that guided the development of this design research practice. Semi-structured interviews were conducted with Sickle Cell experts consisting of patients, a healthcare practitioner and support staff. Common themes were generated from their experience of the condition's impact. D4SC collaborated with Imperial's Invisible Warrior Project, RCABlack, a PhD Archivist Researcher and Photographer at the Slade School of Fine Art, to develop visual prototypes for the exhibition. Results: The findings from these interviews informed the development of a range of workshops and prototypes. The prototypes were tested by Sickle Cell experts and healthcare designers, who provided feedback on the concept. The results showed that the design outputs and exhibition were well-received and had the potential to improve education and awareness of the condition and promote empathy. Design research in healthcare has the potential to create innovative solutions. With the use of a multidisciplinary approach, it can yield a positive impact. Conclusions: The project highlights the importance of design thinking, system thinking and collaboration in developing innovative healthcare solutions for this complex health condition. The study demonstrates the value of a multi-stakeholder approach to designing for empathy. It shows the potential of visualising the impact of Sickle Cell to promote understanding and awareness of the condition. To facilitate the further advancement of the concepts developed in this study, securing funding an

CAPS-1 requires its C2, PH, MHD1 and DCV domains for dense core vesicle exocytosis in mammalian CNS neurons

CAPS (calcium-dependent activator protein for secretion) are multi-domain proteins involved in regulated exocytosis of synaptic vesicles (SVs) and dense core vesicles (DCVs). Here, we assessed the contribution of different CAPS-1 domains to its subcellular localization and DCV exocytosis by expressing CAPS-1 mutations in four functional domains in CAPS-1/-2 null mutant (CAPS DKO) mouse hippocampal neurons, which are severely impaired in DCV exocytosis. CAPS DKO neurons showed normal development and no defects in DCV biogenesis and their subcellular distribution. Truncation of the CAPS-1 C-terminus (CAPS Δ654-1355) impaired CAPS-1 synaptic enrichment. Mutations in the C2 (K428E or G476E) or pleckstrin homology (PH; R558D/K560E/K561E) domain did not. However, all mutants rescued DCV exocytosis in CAPS DKO neurons to only 20% of wild type CAPS-1 exocytosis capacity. To assess the relative importance of CAPS for both secretory pathways, we compared effect sizes of CAPS-1/-2 deficiency on SV and DCV exocytosis. Using the same (intense) stimulation, DCV exocytosis was impaired relatively strong (96% inhibition) compared to SV exocytosis (39%). Together, these data show that the CAPS-1 C-terminus regulates synaptic enrichment of CAPS-1. All CAPS-1 functional domains are required, and the C2 and PH domain together are not sufficient, for DCV exocytosis in mammalian CNS neurons

Tetanus insensitive VAMP2 differentially restores synaptic and dense core vesicle fusion in tetanus neurotoxin treated neurons

The SNARE proteins involved in the secretion of neuromodulators from dense core vesicles (DCVs) in mammalian neurons are still poorly characterized. Here we use tetanus neurotoxin (TeNT) light chain, which cleaves VAMP1, 2 and 3, to study DCV fusion in hippocampal neurons and compare the effects on DCV fusion to those on synaptic vesicle (SV) fusion. Both DCV and SV fusion were abolished upon TeNT expression. Expression of tetanus insensitive (TI)-VAMP2 restored SV fusion in the presence of TeNT, but not DCV fusion. Expression of TI-VAMP1 or TI-VAMP3 also failed to restore DCV fusion. Co-transport assays revealed that both TI-VAMP1 and TI-VAMP2 are targeted to DCVs and travel together with DCVs in neurons. Furthermore, expression of the TeNT-cleaved VAMP2 fragment or a protease defective TeNT in wild type neurons did not affect DCV fusion and therefore cannot explain the lack of rescue of DCV fusion by TI-VAMP2. Finally, to test if two different VAMPs might both be required in the DCV secretory pathway, Vamp1 null mutants were tested. However, VAMP1 deficiency did not reduce DCV fusion. In conclusion, TeNT treatment combined with TI-VAMP2 expression differentially affects the two main regulated secretory pathways: while SV fusion is normal, DCV fusion is absent

VPS35 depletion does not impair presynaptic structure and function

The endosomal system is proposed as a mediator of synaptic vesicle recycling, but the molecular recycling mechanism remains largely unknown. Retromer is a key protein complex which mediates endosomal recycling in eukaryotic cells, including neurons. Retromer is important for brain function and mutations in retromer genes are linked to neurodegenerative diseases. In this study, we aimed to determine the role of retromer in presynaptic structure and function. We assessed the role of retromer by knocking down VPS35, the core subunit of retromer, in primary hippocampal mouse neurons. VPS35 depletion led to retromer dysfunction, measured as a decrease in GluA1 at the plasma membrane, and bypassed morphological defects previously described in chronic retromer depletion models. We found that retromer is localized at the mammalian presynaptic terminal. However, VPS35 depletion did not alter the presynaptic ultrastructure, synaptic vesicle release or retrieval. Hence, we conclude that retromer is present in the presynaptic terminal but it is not essential for the synaptic vesicle cycle. Nonetheless, the presynaptic localization of VPS35 suggests that retromer-dependent endosome sorting could take place for other presynaptic cargo