Microgels as Platforms for Antibody-Mediated Cytokine Scavenging.

Therapeutic antibodies are the key treatment option for various cytokine-mediated diseases, such as rheumatoid arthritis, psoriasis, and inflammatory bowel disease. However, systemic injection of these antibodies can cause side effects and suppress the immune system. Moreover, clearance of therapeutic antibodies from the blood is limiting their efficacy. Here, water-swollen microgels are produced with a size of 25 µm using droplet-based microfluidics. The microgels are functionalized with TNFα antibodies to locally scavenge the pro-inflammatory cytokine TNFα. Homogeneous distribution of TNFα-antibodies is shown throughout the microgel network and demonstrates specific antibody-antigen binding using confocal microscopy and FLIM-FRET measurements. Due to the large internal accessibility of the microgel network, its capacity to bind TNFα is extremely high. At a TNFα concentration of 2.5 µg mL-1 , the microgels are able to scavenge 88% of the cytokine. Cell culture experiments reveal the therapeutic potential of these microgels by protecting HT29 colorectal adenocarcinoma cells from TNFα toxicity and resulting in a significant reduction of COX II and IL8 production of the cells. When the microgels are incubated with stimulated human macrophages, to mimic the in vivo situation of inflammatory bowel disease, the microgels scavenge almost all TNFα that is produced by the cells.

Identifying Efficient Clostridium difficile Toxin A Binders with a Multivalent Neo-Glycoprotein Glycan Library.

Clostridium difficile infections cause gastrointestinal disorders and can lead to life-threatening conditions. The symptoms can vary from severe diarrhea to the formation of pseudomembranous colitis and therefore trigger a need for new therapies. The initial step of disease is the binding of the bacterial enterotoxins toxin A and B to the cell surface of epithelial intestinal cells. Scavenging of the toxins is crucial to inhibit their fatal effect in the human body and circumvent the administration of antibiotics. Cell surface glycans are common as ligands for TcdA. Although crucial for carbohydrate-protein interactions, a multivalent presentation of glycans for binding has been hardly considered. Here, we establish a neo-glycoprotein-based glycan library to identify an effective multivalent glycan ligand for TcdA. It comprises 40 different glycan epitopes based on N-acetyllactosamine precursors. Nine structures exhibit strong binding of the receptor domain. Among them, the Lewisy-Lewisx-epitope shows the best performance for binding both the receptor domain and the holotoxin. Therefore, the glycan was synthesized de novo and coupled to BSA as a scaffold for multivalent presentation. The corresponding neo-glycoprotein facilitates the proper scavenging of TcdA in vitro and effectively protects HT29 cells from TcdA-induced cell damage.

Microgels Sopping Up Toxins-GM1a-Functionalized Microgels as Scavengers for Cholera Toxin.

Vibrio cholerae is a Gram-negative bacterium that causes secretory diarrhea and constitutes a major health threat in the industrialized world and even more in developing countries. Its main virulence factor is the cholera toxin, which is internalized by intestinal epithelial cells after binding to the glycosphingolipid receptor GM1a on their apical surface. A potential future solution to dampen complications of cholera infection is by scavenging the cholera toxin by presenting competitive binding motifs to diminish the in vivo toxicity of V. cholerae. Here, we generate GM1a-functionalized and biocompatible microgels with diameters of 20 µm using drop-based microfluidics. The microgels are designed to exhibit a mesoporous and widely meshed network structure, allowing diffusion of the toxin protein deep into the microgel scavengers. Flow cytometry demonstrates strong and multivalent binding at high capacity of these microgels to the binding domain of the cholera toxin. Cell culture-based assays reveal the ability of these microgels to scavenge and retain the cholera toxin in direct binding competition to colorectal cells. This ability is evidenced by suppressed cyclic adenosine monophosphate production as well as reduced vacuole formation in mucus-forming colorectal HT-29 cells. Therefore, glycan-functionalized microgels show great potential as a non-antibiotic treatment for toxin-mediated infectious disorders.

An Injectable Hybrid Hydrogel with Oriented Short Fibers Induces Unidirectional Growth of Functional Nerve Cells.

To regenerate soft aligned tissues in living organisms, low invasive biomaterials are required to create 3D microenvironments with a structural complexity to mimic the tissue's native architecture. Here, a tunable injectable hydrogel is reported, which allows precise engineering of the construct's anisotropy in situ. This material is defined as an Anisogel, representing a new type of tissue regenerative therapy. The Anisogel comprises a soft hydrogel, surrounding magneto-responsive, cell adhesive, short fibers, which orient in situ in the direction of a low external magnetic field, before complete gelation of the matrix. The magnetic field can be removed after gelation of the biocompatible gel precursor, which fixes the aligned fibers and preserves the anisotropic structure of the Anisogel. Fibroblasts and nerve cells grow and extend unidirectionally within the Anisogels, in comparison to hydrogels without fibers or with randomly oriented fibers. The neurons inside the Anisogel show spontaneous electrical activity with calcium signals propagating along the anisotropy axis of the material. The reported system is simple and elegant and the short magneto-responsive fibers can be produced with an effective high-throughput method, ideal for a minimal invasive route for aligned tissue therapy.

Glycan-Functionalized Microgels for Scavenging and Specific Binding of Lectins.

Lectins are proteins with a well-defined carbohydrate recognition domain. Many microbial proteins such as bacterial toxins possess lectin or lectin-like binding domains to interact with cell membranes that are decorated with glycan recognition motifs. We report a straightforward way to prepare monodisperse and biocompatible polyethylene glycol microgels, which carry glycan motifs for specific binding to lectins. The sugar-functionalized colloids exhibit a wide mesh size and a highly accessible volume. The microgels are prepared via drop-based microfluidics combined with radical polymerization. GSII and ECL are used as model lectins that bind specifically to the corresponding carbohydrates, namely, GlcNAc and LacNAc. LacNAc microgels bind ECL with a high capacity and high affinity (Kd ≈ 0.5 to 1 µM), suggesting multivalent binding of the lectin to the LacNAc-decorated flexible microgel network. Glycan-functionalized microgels present a useful tool for lectin scavenging in biomedical applications.

PUBLIC AND PATIENT INVOLVEMENT IN HEALTH TECHNOLOGY ASSESSMENT: A FRAMEWORK FOR ACTION.

OBJECTIVE: As health technology assessment (HTA) organizations in Canada and around the world seek to involve the public and patients in their activities, frameworks to guide decisions about whom to involve, through which mechanisms, and at what stages of the HTA process have been lacking. The aim of this study was to describe the development and outputs of a comprehensive framework for involving the public and patients in a government agency's HTA process. METHODS: The framework was informed by a synthesis of international practice and published literature, a dialogue with local, national and international stakeholders, and the deliberations of a government agency's public engagement subcommittee in Ontario, Canada. RESULTS: The practice and literature synthesis failed to identify a single, optimal approach to involving the public and patients in HTA. Choice of methods should be considered in the context of each HTA stage, goals for incorporating societal and/or patient perspectives into the process, and relevant societal and/or patient values at stake. The resulting framework is structured around four actionable elements: (i) guiding principles and goals for public and patient involvement (PPI) in HTA, (ii) the establishment of a common language to support PPI efforts, (iii) a flexible array of PPI approaches, and (iv) on-going evaluation of PPI to inform adjustments over time. CONCLUSIONS: A public and patient involvement framework has been developed for implementation in a government agency's HTA process. Core elements of this framework may apply to other organizations responsible for HTA and health system quality improvement.

Nurse practitioner caseload in primary health care: Scoping review.

OBJECTIVES: To identify recommendations for determining patient panel/caseload size for nurse practitioners in community-based primary health care settings. DESIGN: Scoping review of the international published and grey literature. DATA SOURCES: The search included electronic databases, international professional and governmental websites, contact with experts, and hand searches of reference lists. Eligible papers had to (a) address caseload or patient panels for nurse practitioners in community-based primary health care settings serving an all-ages population; and (b) be published in English or French between January 2000 and July 2014. Level one testing included title and abstract screening by two team members. Relevant papers were retained for full text review in level two testing, and reviewed by two team members. A third reviewer acted as a tiebreaker. Data were extracted using a structured extraction form by one team member and verified by a second member. Descriptive statistics were estimated. Content analysis was used for qualitative data. RESULTS: We identified 111 peer-reviewed articles and grey literature documents. Most of the papers were published in Canada and the United States after 2010. Current methods to determine panel/caseload size use large administrative databases, provider work hours and the average number of patient visits. Most of the papers addressing the topic of patient panel/caseload size in community-based primary health care were descriptive. The average number of patients seen by nurse practitioners per day varied considerably within and between countries; an average of 9-15 patients per day was common. Patient characteristics (e.g., age, gender) and health conditions (e.g., multiple chronic conditions) appear to influence patient panel/caseload size. Very few studies used validated tools to classify patient acuity levels or disease burden scores. DISCUSSION: The measurement of productivity and the determination of panel/caseload size is complex. Current metrics may not capture activities relevant to community-based primary health care nurse practitioners. Tools to measure all the components of these role are needed when determining panel/caseload size. Outcomes research is absent in the determination of panel/caseload size. CONCLUSION: There are few systems in place to track and measure community-based primary health care nurse practitioner activities. The development of such mechanisms is an important next step to assess community-based primary health care nurse practitioner productivity and determine patient panel/caseload size. Decisions about panel/caseload size must take into account the effects of nurse practitioner activities on outcomes of care.

Supporting quality public and patient engagement in health system organizations: development and usability testing of the Public and Patient Engagement Evaluation Tool.

OBJECTIVES: Only rudimentary tools exist to support health system organizations to evaluate their public and patient engagement (PPE) activities. This study responds to this gap by developing a generic evaluation tool for use in a wide range of organizations. METHODS: The evaluation tool was developed through an iterative, collaborative process informed by a review of published and grey literature and with the input of Canadian PPE researchers and practitioners. Over a 3-year period, structured e-mail, telephone and face-to-face exchanges, including a modified Delphi process, were used to produce an evaluation tool that includes core principles of high-quality engagement, expected outcomes for each principle and three unique evaluation questionnaires that were tested and revised with input from 65 end users. RESULTS: The tool is structured around four core principles of 'quality engagement': (i) integrity of design and process; (ii) influence and impact; (iii) participatory culture; and (iv) collaboration and common purpose. Three unique questionnaires were developed to assess each of these four evaluation domains from the following perspectives: (i) those who participate in PPE activities; (ii) those who plan, execute or sponsor PPE activities within organizations; and (iii) those who provide the leadership and capacity for PPE within their organizations. CONCLUSIONS: This is the first known collaboration of researchers and practitioners in the co-design of a comprehensive PPE evaluation tool aimed at three distinct respondent groups and for use in a wide range of health system organization settings.

Assessing the impacts of citizen deliberations on the health technology process.

OBJECTIVES: We assessed the impacts of a Citizens' Reference Panel on the deliberations of a provincial health technology advisory committee and its secretariat, which produce, recommendations for the use of health technologies in Ontario, Canada. METHODS: A fourteen-member citizens' reference panel was convened five times between February 2009 and May 2010 to participate in informed, facilitated discussions to inform the assessment of individual technologies and provincial health technology assessment processes more generally. Qualitative data collection methods were used to document observed and perceived impacts of the citizens' panel on the health technology assessment (HTA) process. RESULTS: Panel impacts were observed for all technologies reviewed, at two different stages in the HTA process, and represented macro- (raising awareness) and micro-level (informing recommendations) impacts. These impacts were shaped by periodic opportunities for direct and brokered exchange between the Panel and the expert advisory committee to clarify roles, foster accountability, and build trust. Our findings offer new insights about one of the main considerations in the design of deliberative participatory structures-how to maintain the independence of a citizens' panel while ensuring that their input is considered at key junctures in the HTA process. CONCLUSIONS: Citizens' panels can exert various impacts on the HTA process. Ensuring these types of structures include opportunities for direct exchange between citizens and experts, to clarify roles, promote accountability, and build trust will facilitate their impacts in a variety of settings.