Mapping of a hybrid insulin peptide in the inflamed islet β-cells from NOD mice

There is accumulating evidence that pathogenic T cells in T1D recognize epitopes formed by post-translational modifications of β-cell antigens, including hybrid insulin peptides (HIPs). The ligands for several CD4 T-cell clones derived from the NOD mouse are HIPs composed of a fragment of proinsulin joined to peptides from endogenous β-cell granule proteins. The diabetogenic T-cell clone BDC-6.9 reacts to a fragment of C-peptide fused to a cleavage product of pro-islet amyloid polypeptide (6.9HIP). In this study, we used a monoclonal antibody (MAb) to the 6.9HIP to determine when and where HIP antigens are present in NOD islets during disease progression and with which immune cells they associate. Immunogold labeling of the 6.9HIP MAb and organelle-specific markers for electron microscopy were employed to map the subcellular compartment(s) in which the HIP is localized within β-cells. While the insulin B9-23 peptide was present in nearly all islets, the 6.9HIP MAb stained infiltrated islets only in NOD mice at advanced stages of T1D development. Islets co-stained with the 6.9HIP MAb and antibodies to mark insulin, macrophages, and dendritic cells indicate that 6.9HIP co-localizes within insulin-positive β-cells as well as intra-islet antigen-presenting cells (APCs). In electron micrographs, the 6.9HIP co-localized with granule structures containing insulin alone or both insulin and LAMP1 within β-cells. Exposing NOD islets to the endoplasmic reticulum (ER) stress inducer tunicamycin significantly increased levels of 6.9HIP in subcellular fractions containing crinosomes and dense-core granules (DCGs). This work demonstrates that the 6.9HIP can be visualized in the infiltrated islets and suggests that intra-islet APCs may acquire and present HIP antigens within islets

Relationship between subscapularis tears and injuries to the biceps pulley

The purpose of this study was to analyse the relationship between long head of the biceps brachii (LHBT) lesions and subscapularis tears. The hypothesis was that a bicipital pulley might remain intact, even in the case of a subscapularis tear

Engagement Across Professions: A Mixed Methods Study of Debriefing After Interprofessional Team Training

INTRODUCTION: Simulation is an ideal tool for interprofessional (IP) team training. Debriefing after simulation is key to IP learning, although engagement and participation may be adversely influenced by cultural and hierarchical barriers. This mixed-methods study explored factors influencing learner engagement and participation in IP debriefing and the experience of silent but apparently engaged participants. METHODS: Semistructured profession-specific focus groups were conducted with participants from a weekly IP pediatric simulation program. Focus groups were recorded, transcribed, and thematically analyzed. Eligible participants were assigned to silent or verbal groups according to observed behavior and received a questionnaire. Participants\u27 self-rated engagement scores were compared using a t test. RESULTS: Thirty-six of 81 eligible participants were included, 13 completed a questionnaire, and 23 (8 physicians, 10 nursing staff, 4 pharmacists, 1 respiratory therapist) participated in 13 focus groups. Twenty-two subthemes were grouped into 6 themes: psychological safety, realism, distractors, stress, group characteristics, and facilitator behavior, with differences in perspective according to profession. Of the 36 respondents, 18 were silent and 18 verbal. Self-rated engagement scores differed between groups (3.65 vs. 4.17, P = 0.06); however, silent participants described themselves as engaged. CONCLUSIONS: Themes identified that influenced learner engagement in debriefing included aspects of prebriefing and the simulation. Some aligned with general simulation best practices, such as psychological safety, prebriefing, and facilitator behavior. Findings unique to IP simulation included importance of realism to nonphysician professions, protecting time for training, group composition, and direct probing by cofacilitators to decrease physician bias and emphasize IP contributions. Silent participants reported engagement

Image_2_Mapping of a hybrid insulin peptide in the inflamed islet β-cells from NOD mice.tif

There is accumulating evidence that pathogenic T cells in T1D recognize epitopes formed by post-translational modifications of β-cell antigens, including hybrid insulin peptides (HIPs). The ligands for several CD4 T-cell clones derived from the NOD mouse are HIPs composed of a fragment of proinsulin joined to peptides from endogenous β-cell granule proteins. The diabetogenic T-cell clone BDC-6.9 reacts to a fragment of C-peptide fused to a cleavage product of pro-islet amyloid polypeptide (6.9HIP). In this study, we used a monoclonal antibody (MAb) to the 6.9HIP to determine when and where HIP antigens are present in NOD islets during disease progression and with which immune cells they associate. Immunogold labeling of the 6.9HIP MAb and organelle-specific markers for electron microscopy were employed to map the subcellular compartment(s) in which the HIP is localized within β-cells. While the insulin B9-23 peptide was present in nearly all islets, the 6.9HIP MAb stained infiltrated islets only in NOD mice at advanced stages of T1D development. Islets co-stained with the 6.9HIP MAb and antibodies to mark insulin, macrophages, and dendritic cells indicate that 6.9HIP co-localizes within insulin-positive β-cells as well as intra-islet antigen-presenting cells (APCs). In electron micrographs, the 6.9HIP co-localized with granule structures containing insulin alone or both insulin and LAMP1 within β-cells. Exposing NOD islets to the endoplasmic reticulum (ER) stress inducer tunicamycin significantly increased levels of 6.9HIP in subcellular fractions containing crinosomes and dense-core granules (DCGs). This work demonstrates that the 6.9HIP can be visualized in the infiltrated islets and suggests that intra-islet APCs may acquire and present HIP antigens within islets.</p

ECM Production of Primary Human and Bovine Chondrocytes in Hybrid PEG Hydrogels Containing Type I Collagen and Hyaluronic Acid

The development of advanced materials that facilitate hyaline cartilage formation and regeneration in aging populations is imperative. Critical to the success of this endeavor is the optimization of ECM production from clinically relevant cells. However, much of the current literature focuses on the investigation of primary bovine chondrocytes from young calves, which differ significantly than osteoarthritic cells from human sources. This study examines the levels of extracellular matrix (ECM) production using various levels of type I collagen and hyaluronic acid in poly(ethylene glycol) dimethacrylate (PEGDM) hydrogels in total knee arthroplasties, compared with the results from bovine chondrocytes. The addition of type 1 collagen in both the presence and absence of low levels of hyaluronic acid increased ECM production and/or retention in scaffolds containing either bovine or human chondrocytes. These findings are supported consistently with colorimetric quantification, whole mount extracellular matrix staining for both cell types, and histological staining for glycoaminoglycans and collagen of human chondrocyte containing samples. While exhibiting similar trends, the relative ECM productions levels for the primary human chondrocytes are significantly less than the bovine chondrocytes which reinforces the need for additional optimization

Image_1_Mapping of a hybrid insulin peptide in the inflamed islet β-cells from NOD mice.tif

There is accumulating evidence that pathogenic T cells in T1D recognize epitopes formed by post-translational modifications of β-cell antigens, including hybrid insulin peptides (HIPs). The ligands for several CD4 T-cell clones derived from the NOD mouse are HIPs composed of a fragment of proinsulin joined to peptides from endogenous β-cell granule proteins. The diabetogenic T-cell clone BDC-6.9 reacts to a fragment of C-peptide fused to a cleavage product of pro-islet amyloid polypeptide (6.9HIP). In this study, we used a monoclonal antibody (MAb) to the 6.9HIP to determine when and where HIP antigens are present in NOD islets during disease progression and with which immune cells they associate. Immunogold labeling of the 6.9HIP MAb and organelle-specific markers for electron microscopy were employed to map the subcellular compartment(s) in which the HIP is localized within β-cells. While the insulin B9-23 peptide was present in nearly all islets, the 6.9HIP MAb stained infiltrated islets only in NOD mice at advanced stages of T1D development. Islets co-stained with the 6.9HIP MAb and antibodies to mark insulin, macrophages, and dendritic cells indicate that 6.9HIP co-localizes within insulin-positive β-cells as well as intra-islet antigen-presenting cells (APCs). In electron micrographs, the 6.9HIP co-localized with granule structures containing insulin alone or both insulin and LAMP1 within β-cells. Exposing NOD islets to the endoplasmic reticulum (ER) stress inducer tunicamycin significantly increased levels of 6.9HIP in subcellular fractions containing crinosomes and dense-core granules (DCGs). This work demonstrates that the 6.9HIP can be visualized in the infiltrated islets and suggests that intra-islet APCs may acquire and present HIP antigens within islets.</p