Dementia resources for eating, activity, and meaningful inclusion (DREAM) toolkit co-development: process, output, and lessons learned.

Promoting wellbeing of persons with dementia and their families is a priority of research and practice. Engaging diverse partners, including persons with dementia and their families, to co-develop interventions promotes relevant and impactful solutions. We describe the process, output, and lessons learned from the dementia resources for eating, activity, and meaningful inclusion (DREAM) project, which co-developed tools/resources with persons with dementia, care partners, community service providers, health care professionals, and researchers with the aim of increasing supports for physical activity, healthy eating, and wellbeing of persons with dementia. Our process included: (1) Engaging and maintaining the DREAM Steering Team; (2) Setting and navigating ways of engagement; (3) Selecting the priority audience and content; (4) Drafting the toolkit; (5) Iterative co-development of tools and resources; (6) Usability testing; and (7) Implementation and evaluation. In virtual meetings, the DREAM Steering Team confirmed the toolkit audiences (primary: community service providers; secondary: persons with dementia and care partners) and identified and evolved content areas. An environmental scan identified few existing, high-quality resources aligned with content areas. The Steering Team, additional multi-perspective partners, and external contractors iteratively co-developed new tools/resources to meet gaps over a 4-month virtual process that included virtual meetings, email exchange of documents and feedback, and one-on-one calls by telephone or email. The final DREAM toolkit includes a website with seven learning modules (on the diversity of dementia, rights and inclusion of persons living with dementia, physical activity, healthy eating, dementia-inclusive practices), a learning manual, six videos, nine handouts, and four wallet cards ( www.dementiawellness.ca ). Our co-development participants rated the process highly in relation to the principles and enablers of authentic partnership even though all engagement was virtual. Through use of the co-developed DREAM toolkit, we anticipate community service providers will gain the knowledge and confidence needed to provide dementia-inclusive wellness programs and services that benefit persons with dementia and their families.

A chemically controlled Cas9 switch enables temporal modulation of diverse effectors.

CRISPR-Cas9 has yielded a plethora of effectors, including targeted transcriptional activators, base editors and prime editors. Current approaches for inducibly modulating Cas9 activity lack temporal precision and require extensive screening and optimization. We describe a versatile, chemically controlled and rapidly activated single-component DNA-binding Cas9 switch, ciCas9, which we use to confer temporal control over seven Cas9 effectors, including two cytidine base editors, two adenine base editors, a dual base editor, a prime editor and a transcriptional activator. Using these temporally controlled effectors, we analyze base editing kinetics, showing that editing occurs within hours and that rapid early editing of nucleotides predicts eventual editing magnitude. We also reveal that editing at preferred nucleotides within target sites increases the frequency of bystander edits. Thus, the ciCas9 switch offers a simple, versatile approach to generating chemically controlled Cas9 effectors, informing future effector engineering and enabling precise temporal effector control for kinetic studies.

Prevalence of current nutrition care practices for disease-related malnutrition in Canadian hospitals.

Disease-related malnutrition is common in hospital patients. The Health Standards Organization Canadian Malnutrition Prevention, Detection, and Treatment Standard was published in 2021. The purpose of this study was to determine the current state of nutrition care in hospitals prior to implementation of the Standard. An online survey was distributed to hospitals across Canada via email. A representative reported on nutrition best practices based on the Standard at the hospital level. Descriptive and bivariate statistics were completed for selected variables based on size and type of hospital. One hundred and forty-three responses from nine provinces were received (56% community, 23% academic, and 21% other). Malnutrition risk screening was being completed on admission in 74% (n = 106/142) of hospitals, although not all units participated in screening all patients. Nutrition-focused physical exam is completed as part of a nutrition assessment in 74% (n = 101/139) of sites. Flagging a malnutrition diagnosis (n = 38/104) and physician documentation (18/136) were sporadic. Academic and medium (100-499 beds) and large hospitals (500+ beds) were more likely to have a physician document a malnutrition diagnosis. Some, but not all, best practices are occurring in Canadian hospitals on a regular basis. This demonstrates a need for continued knowledge mobilization of the Standard.

Loneliness and resilience are associated with nutrition risk after the first wave of COVID-19 in community-dwelling older Canadians.

Nutrition risk is linked to hospitalization, frailty, depression, and death. Loneliness during the coronavirus disease 2019 (COVID-19) pandemic may have heightened nutrition risk. We sought to determine prevalence of high nutrition risk and whether loneliness, mental health, and assistance with meal preparation/delivery were associated with risk in community-dwelling older adults (65+ years) after the first wave of COVID-19 in association analyses and when adjusting for meaningful covariates. Data were collected from 12 May 2020 to 19 August 2020. Descriptive statistics, association analyses, and linear regression analyses were conducted. For our total sample of 272 participants (78 ± 7.3 years old, 70% female), the median Seniors in the Community: Risk evaluation for Eating and Nutrition (SCREEN-8) score (nutrition risk) was 35 [1st quartile, 3rd quartile: 29, 40], and 64% were at high risk (SCREEN-8 < 38). Fifteen percent felt lonely two or more days a week. Loneliness and meal assistance were associated with high nutrition risk in association analyses. In multivariable analyses adjusting for other lifestyle factors, loneliness was negatively associated with SCREEN-8 scores (-2.92, 95% confidence interval [-5.51, -0.34]), as was smoking (-3.63, [-7.07, -0.19]). Higher SCREEN-8 scores were associated with higher education (2.71, [0.76, 4.66]), living with others (3.17, [1.35, 4.99]), higher self-reported health (0.11, [0.05, 0.16]), and resilience (1.28, [0.04, 2.52]). Loneliness, but not mental health and meal assistance, was associated with nutrition risk in older adults after the first wave of COVID-19. Future research should consider longitudinal associations among loneliness, resilience, and nutrition.

Qualitative analysis of a virtual research meeting summarises expert-based strategies to promote hydration in residential care during COVID-19 and beyond.

OBJECTIVES: Poor fluid intake is a complex and long-standing issue in residential care, further exacerbated by COVID-19 infection control procedures. There is no consensus on how best to prevent dehydration in residents who vary in their primary reasons for insufficient fluid intake for a variety of reasons. The objectives of this research were to determine expert and provider perspectives on: (1) how COVID-19 procedures impacted hydration in residential care and potential solutions to mitigate these challenges and (2) strategies that could target five types of residents based on an oral hydration typology focused on root causes of low fluid intake. DESIGN: Qualitative study based on virtual group discussion. The discussion was audiorecorded with supplementary field notes. Qualitative content analysis was completed. SETTING: Residential care. PARTICIPANTS: 27 invited researcher and provider experts. RESULTS: Challenges that have potentially impacted hydration of residents because of COVID-19 procedures were categorised as resident (eg, apathy), staff (eg, new staff) and home-related (eg, physical distancing in dining rooms). Potential solutions were offered, such as fun opportunities (eg, popsicle) for distanced interactions; training new staff on how to approach specific residents and encourage drinking; and automatically providing water at meals. Several strategies were mapped to the typology of five types of residents with low intake (eg, sipper) and categorised as: supplies (eg, vessels with graduated markings), timing (eg, identify best time of day for drinking), facility context (eg, identify preferred beverages), socialisation (eg, promote drinking as a social activity) and education (eg, educate cognitively well on water consumption goals). CONCLUSIONS: COVID-19 has necessitated new procedures and routines in residential care, some of which can be optimised to promote hydration. A variety of strategies to meet the hydration needs of different subgroups of residents can be compiled into multicomponent interventions for future research.

Feasibility and Acceptability Testing of Evidence-Based Hydration Strategies for Residential Care.

The current study examined stakeholder perspectives on the perceived effectiveness, feasibility, and acceptability of 20 evidence-based strategies appropriate for residential care via an online survey (N = 162). Most participants worked in long-term care (83%), were direct care providers (62%), worked in food/nutrition roles (55%), and identified as female (94%). Strategies that were rated as effective, feasible, and likely to be used in the future were social drinking events, increased drink options at meals, and pre-thickened drinks. Participants also listed their top strategies for inclusion in a multicomponent intervention. Responses to open-ended questions provided insight on implementation, compliance, and budget constraints. Participant perspectives provide insight into developing a multicomponent intervention. Strategies prioritized for such an intervention include: staff education, social drinking opportunities, drinks trolley, volunteer support, improved beverage availability, hydration reminders, offering preferred beverages, and prompting residents to drink using various cues. [Research in Gerontological Nursing, 15(1), 27-38.].

Telephone Administration of the Automated Self-Administered 24-hour Dietary Assessment in Older Adults: Lessons Learned.

Automated Self-Administered 24-hour Dietary Assessment (ASA24) is an economical method of estimating dietary intake as nutrient analysis is automated, but its use in older adults is limited. The purpose of this work was to guide dietitians and future researchers on how to use the ASA24 with older adults, considering potential barriers encountered and strategies used to support completion based on our experience using this tool in a pilot clinical trial. ASA24 was completed by phone interview with 39 older adults. Challenges included: recalling food intake in detail, recording frequent eating occasions and complicated recipes, and general problems with communication. Strategies to support collection included making morning phone calls and suggesting that seniors write down the food consumed. Phone interviews were acceptable to older adults, but sufficient time was required. Dietitians and future researchers can use these findings to obtain dietary intake data from this hard-to-reach group.

The effect of blue dishware versus white dishware on food intake and eating challenges among residents living with dementia: a crossover trial.

OBJECTIVE: Residents living with dementia (RLWD) often experience changes in their visual perception, which could reduce food intake. Inadequate food intake is known to cause malnutrition, which increases the risk of hospitalization, morbidity, and mortality. This study evaluated the effectiveness of using blue dishware compared to white dishware to improve food intake and mitigate eating challenges among 18 RLWD (mean age 84.6 ± 7.9 years, 72.2% female). RESULTS: A within-within person crossover design determined differences in food intake and eating challenges between blue and white dishware conditions. Five participants responded to the blue dishware and increased their average food intake by ≥ 10%. Responders were not different from non-responders in terms of demographic or health characteristics. The proportion of eating challenges experienced was not significantly different between the blue and white dishware conditions. Percent food intake was significantly greater at lunch (83.5 ± 19.0%) compared to dinner (75.8 ± 22.1%; p < 0.0001), regardless of dishware condition. However, there were no significant differences for food intake between the dishware conditions, even after matching food choices. Promoting food intake and reducing eating challenges in RLWD likely needs multi-component interventions targeting meal quality, meal access, and mealtime experience. Trial registration ClincialTrials.gov Identifier: NCT04298788. Retrospectively registered: 6 March 2020, https://clinicaltrials.gov/ct2/show/NCT04298788?term=NCT04298788&draw=2&rank=1 .

Suppression of unwanted CRISPR-Cas9 editing by co-administration of catalytically inactivating truncated guide RNAs.

CRISPR-Cas9 nucleases are powerful genome engineering tools, but unwanted cleavage at off-target and previously edited sites remains a major concern. Numerous strategies to reduce unwanted cleavage have been devised, but all are imperfect. Here, we report that off-target sites can be shielded from the active Cas9•single guide RNA (sgRNA) complex through the co-administration of dead-RNAs (dRNAs), truncated guide RNAs that direct Cas9 binding but not cleavage. dRNAs can effectively suppress a wide-range of off-targets with minimal optimization while preserving on-target editing, and they can be multiplexed to suppress several off-targets simultaneously. dRNAs can be combined with high-specificity Cas9 variants, which often do not eliminate all unwanted editing. Moreover, dRNAs can prevent cleavage of homology-directed repair (HDR)-corrected sites, facilitating scarless editing by eliminating the need for blocking mutations. Thus, we enable precise genome editing by establishing a flexible approach for suppressing unwanted editing of both off-targets and HDR-corrected sites.

Temporal and rheostatic control of genome editing with a chemically-inducible Cas9.

Nuclease-mediated DNA cleavage and subsequent repair lie at the heart of genome editing, and the RNA-guided endonuclease Cas9 has emerged as the most widely-used tool for facilitating this process. Extensive biochemical and biophysical efforts have revealed much regarding the structure, mechanism, and cellular properties of Cas9. This has enabled engineering of Cas9 variants with enhanced activity, specificity, and other features. However, we lack a detailed understanding of the kinetics of Cas9-mediated DNA cleavage and repair in vivo. To study in vivo Cas9 cleavage kinetics and activity dose-dependence, we have engineered a chemically-inducible, single-component Cas9, ciCas9. ciCas9 allows for temporal and rheostatic control of Cas9 activity using a small molecule activator, A115. We have also developed a droplet-digital PCR-based assay (DSB-ddPCR) to directly quantify Cas9-mediated double-stranded breaks (DSBs). The methods in this chapter describe the application of ciCas9 and DSB-ddPCR to study the kinetics and dose-dependence of Cas9 editing in vivo.

Multi-input chemical control of protein dimerization for programming graded cellular responses.

Chemical and optogenetic methods for post-translationally controlling protein function have enabled modulation and engineering of cellular functions. However, most of these methods only confer single-input, single-output control. To increase the diversity of post-translational behaviors that can be programmed, we built a system based on a single protein receiver that can integrate multiple drug inputs, including approved therapeutics. Our system translates drug inputs into diverse outputs using a suite of engineered reader proteins to provide variable dimerization states of the receiver protein. We show that our single receiver protein architecture can be used to program a variety of cellular responses, including graded and proportional dual-output control of transcription and mammalian cell signaling. We apply our tools to titrate the competing activities of the Rac and Rho GTPases to control cell morphology. Our versatile tool set will enable researchers to post-translationally program mammalian cellular processes and to engineer cell therapies.

Rheostatic Control of Cas9-Mediated DNA Double Strand Break (DSB) Generation and Genome Editing.

We recently reported two novel tools for precisely controlling and quantifying Cas9 activity: a chemically inducible Cas9 variant (ciCas9) that can be rapidly activated by small molecules and a ddPCR assay for time-resolved measurement of DNA double strand breaks (DSB-ddPCR). Here, we further demonstrate the potential of ciCas9 to function as a tunable rheostat for Cas9 function. We show that a new highly potent and selective small molecule activator paired with a more tightly regulated ciCas9 variant expands the range of accessible Cas9 activity levels. We subsequently demonstrate that ciCas9 activity levels can be dose-dependently tuned with a small molecule activator, facilitating rheostatic time-course experiments. These studies provide the first insight into how Cas9-mediated DSB levels correlate with overall editing efficiency. Thus, we demonstrate that ciCas9 and our DSB-ddPCR assay permit the time-resolved study of Cas9 DSB generation and genome editing kinetics at a wide range of Cas9 activity levels.

Psychosocial and Sexual Well-Being Following Nipple-Sparing Mastectomy and Reconstruction.

Nipple-sparing mastectomy (NSM) is considered an oncologically safe option for select patients. As many patients are candidates for nipple-sparing or skin-sparing mastectomy (SSM), reliable patient-reported outcome data are crucial for decision-making. The objective of this study was to determine whether patient satisfaction and/or health-related quality of life (HRQOL) were improved by preservation of the nipple with NSM compared to SSM and nipple reconstruction. Subjects were identified from a prospectively maintained database of patients who completed the BREAST-Q following mastectomy and breast reconstruction between March and October 2011 at Memorial Sloan Kettering Cancer Center. Fifty-two patients underwent NSM followed by immediate expander-implant reconstruction. A comparison group consisted of 202 patients who underwent SSM followed by immediate expander-implant reconstruction and later nipple reconstruction. HRQOL and satisfaction domains as measured by BREAST-Q scores were compared in multivariate linear regression analyzes that controlled for potential confounding factors. NSM patients reported significantly higher scores in the psychosocial (p = 0.01) and sexual well-being (p = 0.02) domains compared to SSM patients. There was no significant difference in the BREAST-Q physical well-being, satisfaction with breast, or satisfaction with outcome domains between the NSM and SSM groups. NSM is associated with higher psychosocial and sexual well-being compared to SSM and nipple reconstruction. Preoperative discussion of such HRQOL outcomes with patients may facilitate informed decision-making and realistic postoperative expectations.

Complications following expander/implant breast reconstruction utilizing acellular dermal matrix: a systematic review and meta-analysis.

BACKGROUND: The recent increase in popularity of acellular dermal matrix assistance in immediate expander/implant breast reconstruction has led to variety of viewpoints. Many studies are published indicating an increase in complications with the use of acellular dermal matrix, while others indicate there is no increase in complications. METHODS: This meta-analysis utilizes information from available studies that directly compare one specific type of acellular dermal matrix with traditional methods of immediate expander/implant breast reconstruction. Eight studies were found through a meticulous literature search that met these criteria. RESULTS: There was more than a 2-fold increase in the number of infections and explanations in the acellular dermal matrix group compared to the control. There was a 3-fold increase in seroma formation in the acellular dermal matrix group compared to the control. There was a significant difference of intraoperative fill volumes between the acellular dermal matrix group compared to the control. CONCLUSIONS: This study illustrates that after pooling all available date regarding the use of acellular dermal matrix in immediate expander/implant breast reconstruction there appears to be an increased rate of complications. However, the increased intraoperative fill volume may lead to ultimately greater patient satisfaction.

Treatment modality affects allograft-derived Schwann cell phenotype and myelinating capacity.

We used peripheral nerve allografts, already employed clinically to reconstruct devastating peripheral nerve injuries, to study Schwann cell (SC) plasticity in adult mice. By modulating the allograft treatment modality we were able to study migratory, denervated, rejecting, and reinnervated phenotypes in transgenic mice whose SCs expressed GFP under regulatory elements of either the S100b (S100-GFP) or nestin (Nestin-GFP) promoters. Well-differentiated SCs strongly expressed S100-GFP, while Nestin-GFP expression was stimulated by denervation, and in some cases, axons were constitutively labeled with CFP to enable in vivo imaging. Serial imaging of these mice demonstrated that untreated allografts were rejected within 20 days. Cold preserved (CP) allografts required an initial phase of SC migration that preceded axonal regeneration thus delaying myelination and maturation of the SC phenotype. Mice immunosuppressed with FK506 demonstrated mild subacute rejection, but the most robust regeneration of myelinated and unmyelinated axons and motor endplate reinnervation. While characterized by fewer regenerating axons, mice treated with the co-stimulatory blockade (CSB) agents anti-CD40L mAb and CTLAIg-4 demonstrated virtually no graft rejection during the 28 day experiment, and had significant increases in myelination, connexin-32 expression, and Akt phosphorylation compared with any other group. These results indicate that even with SC rejection, nerve regeneration can occur to some degree, particularly with FK506 treatment. However, we found that co-stimulatory blockade facilitate optimal myelin formation and maturation of SCs as indicated by protein expression of myelin basic protein (MBP), connexin-32 and phospho-Akt.

Surgical management of chronic wounds T.

The management of chronic wounds poses a challenge to internists and surgeons alike. Chronic wounds are trapped in a nonadvancing phase of healing and are unable to progress through the sequential stages of tissue repair. The primary roles of surgery in managing these wounds are debridement and wound closure. Surgical debridement releases the chronic wound from its arrested state by removing nonviable tissue, bacteria, and other inhibitory factors, effectively converting it into an acute wound that can undergo healing more effectively. Debridement can be accomplished with a number of techniques, and recent innovations in the wound care industry have added invaluable tools to the wound healing armamentarium. While surgical debridement and closure provide definitive treatment, identifying and treating the underlying cause of a chronic wound is critical to promoting timely healing and preventing wound recurrence.

Binary imaging analysis for comprehensive quantitative histomorphometry of peripheral nerve.

Quantitative histomorphometry is the current gold standard for objective measurement of nerve architecture and its components. Many methods still in use rely heavily upon manual techniques that are prohibitively time consuming, predisposing to operator fatigue, sampling error, and overall limited reproducibility. More recently, investigators have attempted to combine the speed of automated morphometry with the accuracy of manual and semi-automated methods. Systematic refinements in binary imaging analysis techniques combined with an algorithmic approach allow for more exhaustive characterization of nerve parameters in the surgically relevant injury paradigms of regeneration following crush, transection, and nerve gap injuries. The binary imaging method introduced here uses multiple bitplanes to achieve reproducible, high throughput quantitative assessment of peripheral nerve. Number of myelinated axons, myelinated fiber diameter, myelin thickness, fiber distributions, myelinated fiber density, and neural debris can be quantitatively evaluated with stratification of raw data by nerve component. Results of this semi-automated method are validated by comparing values against those obtained with manual techniques. The use of this approach results in more rapid, accurate, and complete assessment of myelinated axons than manual techniques.

Self-organization and branching morphogenesis of primary salivary epithelial cells.

Embryonic tissues may provide clues about mechanisms required for tissue reassembly and regeneration, but few studies have utilized primary embryonic tissue to study tissue assembly. To test the capacity of tissue fragments to regenerate, we cultured fragments of embryonic day 13 (E13) mouse submandibular gland (SMG) epithelium and found that fragments as small as a quarter-bud retain the ability to branch. Further, we found that completely dissociated SMG epithelial cells self-organize into structures that undergo significant branching. Investigation into the mechanisms involved in tissue self-assembly demonstrated that inhibition of beta(1) integrin prevents cell aggregation, while inhibition of E-cadherin hinders aggregate compaction. Immunostaining showed that the cellular architecture and expression patterns of E-cadherin, beta-catenin, and actin in the reassembled aggregates mirror those seen in intact glands. Adding SMG mesenchymal cells to the epithelial cell cultures facilitates branching and morphological differentiation. Quantitative real-time RT-PCR indicated that the aggregates express the differentiation markers aquaporin-5 (AQP5), prolactin-inducible protein (PIP), and SMG protein C (SMGC). Together, these data show that dissociated SMG epithelial cells self-organize and undergo branching morphogenesis to form tissues with structural features and differentiation markers characteristic of the intact gland. These findings provide insights into self-assembly and branching that will facilitate future regeneration strategies in the salivary gland and other organs.

Cell and fibronectin dynamics during branching morphogenesis.

Branching morphogenesis is a dynamic developmental process shared by many organs, but the mechanisms that reorganize cells during branching morphogenesis are not well understood. We hypothesized that extensive cell rearrangements are involved, and investigated cell migration using two-color confocal time-lapse microscopy to image cell and extracellular-matrix dynamics in developing salivary glands. We labeled submandibular salivary gland (SMG) epithelial cells with green fluorescent protein and matrix with fluorescent fibronectin. Surprisingly, we observed substantial, rapid and relatively random migration of individual epithelial cells during branching morphogenesis. We predicted that cell migration would decrease after formation of acini and, indeed, found that rapid cell movements do not occur in SMG from newborn mice. However, in embryonic SMG epithelial cells, we observed an absence of choreographed cell migration, indicating that patterned cell migration alone cannot explain the highly ordered process of branching morphogenesis. We therefore hypothesized a role for directional fibronection assembly in branching. Washout and pulse-chase experiments revealed that older fibronectin accumulates at the base of the clefts and translocates inwards as a wedge, with newer fibronectin assembling behind it. These findings identify a new mechanism for branching morphogenesis involving directional fibronectin translocation superimposed on individual cell dynamics.