The Hyaluronidase, TMEM2, Promotes ER Homeostasis and Longevity Independent of the UPRER.

Cells have evolved complex mechanisms to maintain protein homeostasis, such as the UPRER, which are strongly associated with several diseases and the aging process. We performed a whole-genome CRISPR-based knockout (KO) screen to identify genes important for cells to survive ER-based protein misfolding stress. We identified the cell-surface hyaluronidase (HAase), Transmembrane Protein 2 (TMEM2), as a potent modulator of ER stress resistance. The breakdown of the glycosaminoglycan, hyaluronan (HA), by TMEM2 within the extracellular matrix (ECM) altered ER stress resistance independent of canonical UPRER pathways but dependent upon the cell-surface receptor, CD44, a putative HA receptor, and the MAPK cell-signaling components, ERK and p38. Last, and most surprisingly, ectopic expression of human TMEM2 in C. elegans protected animals from ER stress and increased both longevity and pathogen resistance independent of canonical UPRER activation but dependent on the ERK ortholog mpk-1 and the p38 ortholog pmk-1.

Don't Throw Away the Handout: Efficacy of Patient Education in Pediatric Orthopaedic Trauma.

BACKGROUND: Printed educational materials (PEMs) have been used for patient education in various settings. The purpose of this study was to determine the readability, understandability, and actionability of trauma-related educational material from the Pediatric Orthopaedic Society of North America (POSNA, Orthokids), as well as determine its efficacy in educating pediatric orthopaedic trauma patients and caregivers. METHODS: The readability, understandability and actionability of PEMs was assessed using the Patient Education materials Assessment Tool (PEMAT). Five reviewers ranging in experience independently evaluated the educational materials. The efficacy of PEMs was assessed prospectively by randomizing patients into 2 groups. The first group (Education) received the OrthoKids educational material related to the patient's fracture. The second group (No Education) did not receive the educational material. At the first follow-up visit, parents/guardians in both groups completed surveys. Statistical analyses included descriptive and univariate statistics. RESULTS: The understandability of PEMs was similar (68% to 74%); however, the educational materials had varying actionability scores ranging from 20% for femoral shaft fractures to 60% for elbow fractures. In total, 101 patients were randomized to assess the efficacy of educational materials (Education=51, No Education=50). There were no significant differences in sex, age, race/ethnicity, and level of education between caregivers in both groups ( P > 0.05). Only 61% (31/51) participants in the Education group reported using the educational material; however, 67% to 68% of participants in either group reported wanting PEMs. Participants in the group that did not receive PEMs were significantly more likely to use the internet to find more information (74% vs. 51%, P < 0.05). CONCLUSIONS: This study suggests that participants that did not receive PEMs were significantly more likely to search the internet for more information. Improving the quality and actionability of educational resources on electronic platforms is needed to improve patient education. A multi-modal approach using PEMs that includes a list of high-quality online sources would likely be most effective in educating pediatric trauma patients and caregivers. LEVEL OF EVIDENCE: I.

Effect of obesity on short- and long-term complications of shoulder arthroplasty.

BACKGROUND: The proportion of patients undergoing total shoulder arthroplasty (TSA) with obesity continues to grow every year in the United States. Although comorbid obesity is common among TSA patients, the relationship of obesity on medical and surgical complications remains debated. The goal of this study was to evaluate a national database for postoperative medical and surgical complications in patients undergoing TSA with comorbid obesity. METHODS: Patients undergoing anatomic and reverse TSA were studied in the PearlDiver database. Current Procedural Terminology (CPT) and International Classification of Diseases (ICD) codes were used to compare patients with and without preoperative obesity who underwent TSA, and they were stratified based on body mass index (BMI) into nonobese, obese, morbidly obese, and superobese. A matched comparison was performed at a 1:1 ratio based on age, sex, diabetes, smoking, tobacco use, and Charlson Comorbidity Index. RESULTS: From 2010 to 2020, a total of 113,634 patients undergoing anatomic or reverse TSA were identified in a national database. During this time, the percentage of TSA patients with obesity increased every year. Matched cohort analysis demonstrated higher odds of readmission, deep vein thrombosis and pulmonary embolism, superficial infection, and prosthetic joint infection at 90 days postoperatively in the obesity group. There were no increased odds of mechanical complications or revision surgery at 2 years in the obesity group when matched to nonobese patients with similar comorbidities. CONCLUSION: The number of patients undergoing TSA with obesity is rising. Medical complications and infection after TSA are greater in obese patients even when matching for medical comorbidities, age, and sex, and rates of complication increase as BMI increases. Obesity is not an independent risk factor for mechanical surgical complications and revision surgery, and the relatively higher rates are likely due to an increased burden of other comorbidities. Surgeons should counsel obese patients appropriately regarding their perioperative risk of medical complication, but they should not expect higher rates of mechanical complication or revision surgery at 2-year follow-up when compared to a matched control group with similar comorbidities.

Development and Evaluation of Two Integrated Care Models for Children Using a Partnered Formative Evaluation Approach.

Objective: To describe the development and evaluation of two integrated care models using a partnered formative evaluation approach across a private foundation, clinic leaders, providers and staff, and a university-based research center. Design: Retrospective cohort study using multiple data sources. Setting: Two federal qualified health care centers serving low-income children and families in Chicago. Participants: Private foundation, clinic and academic partners. Interventions: Development of two integrated care models and partnered evaluation design. Main Outcome Measures: Accomplishments and early lessons learned. Results: Together, the foundation-clinic-academic partners worked to include best practices in two integrated care models for children while developing the evaluation design. A shared data collection approach, which empowered the clinic partners to collect data using a web-based tool for a prospective longitudinal cohort study, was also created. Conclusion: Across three formative evaluation stages, the foundation, clinic, and academic partners continued to reach beyond their respective traditional roles of project oversight, clinical service, and research as adjustments were collectively made to accommodate barriers and unanticipated events. Together, an innovative shared data collection approach was developed that extends partnered research to include data collection being led by the clinic partners and supported by the technical resources of a university-based research center.

Sex-based Differences in Patient Perspectives and Experiences With Shoulder, Hip, and Knee Arthroplasty.

INTRODUCTION: Sex disparities in presentation of osteoarthritis and utilization of joint replacement surgery (JRS) have been demonstrated. The role of patients' unique perspectives on JRS on their treatment decisions is poorly understood. METHODS: JRS candidates who were offered JRS but declined surgical treatment completed this survey. Survey questions included demographic information, patient experiences and current opinions around JRS, patient experiences with providers, goals and concerns, and barriers to JRS. RESULTS: More women experience barriers to undergoing JRS compared with men (53% versus 16%; P = 0.014). While both men and women indicated pain relief as their primary goal for treatment, women were significantly more likely to prioritize regaining the ability to complete daily tasks and responsibilities when compared with men (P = 0.007). Both men and women indicated that low symptom severity and nonsurgical treatment options were the reasons for not undergoing JRS (P = 0.455). Compared with men, women trended toward feeling that they were not sufficiently educated about JRS (P = 0.051). CONCLUSION: Women have unique perspectives and goals for JRS that may pose sex-specific barriers to care. A better understanding of how patients' gendered experiences affect their decision making is necessary to improve treatment of osteoarthritis and decrease disparities in care.

Distinct human stem cell subpopulations drive adipogenesis and fibrosis in musculoskeletal injury.

Fibroadipogenic progenitors (FAPs) maintain healthy skeletal muscle in homeostasis but drive muscle degeneration in chronic injuries by promoting adipogenesis and fibrosis. To uncover how these stem cells switch from a pro-regenerative to pro-degenerative role we perform single-cell mRNA sequencing of human FAPs from healthy and injured human muscles across a spectrum of injury, focusing on rotator cuff tears. We identify multiple subpopulations with progenitor, adipogenic, or fibrogenic gene signatures. We utilize full spectrum flow cytometry to identify distinct FAP subpopulations based on highly multiplexed protein expression. Injury severity increases adipogenic commitment of FAP subpopulations and is driven by the downregulation of DLK1. Treatment of FAPs both in vitro and in vivo with DLK1 reduces adipogenesis and fatty infiltration, suggesting that during injury, reduced DLK1 within a subpopulation of FAPs may drive degeneration. This work highlights how stem cells perform varied functions depending on tissue context, by dynamically regulating subpopulation fate commitment, which can be targeted improve patient outcomes after injury.

Defining Endogenous Mitochondrial Transfer in Muscle After Rotator Cuff Injury.

BACKGROUND: Rotator cuff muscle degeneration leads to poor clinical outcomes for patients with rotator cuff tears. Fibroadipogenic progenitors (FAPs) are resident muscle stem cells with the ability to differentiate into fibroblasts as well as white and beige adipose tissue. Induction of the beige adipose phenotype in FAPs has been shown to improve muscle quality after rotator cuff tears, but the mechanisms of how FAPs exert their beneficial effects have not been fully elucidated. PURPOSE: To study the horizontal transfer of mitochondria from FAPs to myogenic cells and examine the effects of ß-agonism on this novel process. STUDY DESIGN: Controlled laboratory study. METHODS: In mice that had undergone a massive rotator cuff tear, single-cell RNA sequencing was performed on isolated FAPs for genes associated with mitochondrial biogenesis and transfer. Murine FAPs were isolated by fluorescence-activated cell sorting and treated with a ß-agonist versus control. FAPs were stained with mitochondrial dyes and cocultured with recipient C2C12 myoblasts, and the rate of transfer was measured after 24 hours by flow cytometry. PdgfraCreERT/MitoTag mice were generated to study the effects of a rotator cuff injury on mitochondrial transfer. PdgfraCreERT/tdTomato mice were likewise generated to perform lineage tracing of PDGFRA+ cells in this injury model. Both populations of transgenic mice underwent tendon transection and denervation surgery, and MitoTag-labeled mitochondria from Pdgfra+ FAPs were visualized by fluorescent microscopy, spinning disk confocal microscopy, and 2-photon microscopy; overall mitochondrial quantity was compared between mice treated with ß-agonists and dimethyl sulfoxide. RESULTS: Single-cell RNA sequencing in mice that underwent rotator cuff tear demonstrated an association between transcriptional markers of adipogenic differentiation and genes associated with mitochondrial biogenesis. In vitro cocultures of murine FAPs with C2C12 cells revealed that treatment of cells with a ß-agonist increased mitochondrial transfer compared to control conditions (17.8% ± 9.9% to 99.6% ± 0.13% P < .0001). Rotator cuff injury in PdgfraCreERT/MitoTag mice resulted in a robust increase in MitoTag signal in adjacent myofibers compared with uninjured mice. No accumulation of tdTomato signal from PDGFRA+ cells was seen in injured fibers at 6 weeks after injury, suggesting that FAPs do not fuse with injured muscle fibers but rather contribute their mitochondria. CONCLUSION: The authors have described a novel process of endogenous mitochondrial transfer that can occur within the injured rotator cuff between FAPs and myogenic cells. This process may be leveraged therapeutically with ß-agonist treatment and represents an exciting target for improving translational therapies available for rotator cuff muscle degeneration. CLINICAL RELEVANCE: Promoting endogenous mitochondrial transfer may represent a novel translational strategy to address muscle degeneration after rotator cuff tears.

Multimodal prerehabilitation for elderly patients with sarcopenia in colorectal surgery.

Sarcopenia, which is characterized by progressive and generalized loss of skeletal muscle mass and strength, has been well described to be associated with numerous poor postoperative outcomes, such as increased perioperative mortality, postoperative sepsis, prolonged length of stay, increased cost of care, decreased functional outcome, and poorer oncological outcomes in cancer surgery. Multimodal prehabilitation, as a concept that involves boosting and optimizing the preoperative condition of a patient prior to the upcoming stressors of a surgical procedure, has the purported benefits of reversing the effects of sarcopenia, shortening hospitalization, improving the rate of return to bowel activity, reducing the costs of hospitalization, and improving quality of life. This review aims to present the current literature surrounding the concept of sarcopenia, its implications pertaining to colorectal cancer and surgery, a summary of studied multimodal prehabilitation interventions, and potential future advances in the management of sarcopenia.

Sex-Based Differences in the Utilization of Shoulder, Hip, and Knee Arthroplasty.

INTRODUCTION: Studies show that females have a higher prevalence of osteoarthritis, worse symptoms, but lower rates of joint replacement surgery (JRS). The reason for this remains unknown. METHODS: A database of JRS candidates was created for patients seen in 2019 at an academic center. Demographics, Kellgren-Lawrence grades, symptom duration, visual analogue pain score, Charlson Comorbidity Index, and nonsurgical treatments were collected. Patients who were offered but declined surgery were invited to focus groups. Two independent sample t-tests, Mann-Whitney U tests, and chi-square tests were used for continuous, scored, and categorical variables, respectively, with two-tailed significance <0.05. Qualitative, code-based analysis was performed for the focus groups. RESULTS: The cohort included 321 patients (81 shoulder, 59 hip, and 181 knee) including 199 females (62.0%). There were no differences in proportions of females versus males who underwent JRS or in nonsurgical treatments. Female shoulder arthritis patients were older, had a higher visual analogue pain score, and had a higher Charlson Comorbidity Index. In focus groups, males prioritized waiting for technology advancements to return to an active lifestyle, whereas females experienced negative provider interactions, self-advocated for treatment, concerned about pain, and believed that their sex affected their treatment. DISCUSSION: We found equal utilization of JRS at our institution. However, female patients experienced unique barriers to surgery.

Muscle-Derived Beige Adipose Precursors Secrete Promyogenic Exosomes That Treat Rotator Cuff Muscle Degeneration in Mice and Are Identified in Humans by Single-Cell RNA Sequencing.

BACKGROUND: Muscle atrophy, fibrosis, and fatty infiltration are common to a variety of sports-related and degenerative conditions and are thought to be irreversible. Fibroadipogenic progenitors (FAPs) are multipotent resident muscle stem cells with the capacity to differentiate into fibrogenic as well as white and beige adipose tissue (BAT). FAPs that have assumed a BAT differentiation state (FAP-BAT) have proven efficacious in treating muscle degeneration in numerous injury models. PURPOSE: To characterize the subpopulation of murine FAPs with FAP-BAT activity, determine whether their promyogenic effect is mediated via exosomes, and analyze human FAPs for an analogous promyogenic exosome-rich subpopulation. STUDY DESIGN: Controlled laboratory study. METHODS: FAPs from UCP1 reporter mice were isolated via fluorescence-activated cell sorting and sorted according to the differential intensity of the UCP1 signal observed: negative for UCP1 (UCP1-), intermediate intensity (UCP1+), and high intensity (UCP1++). Bulk RNA sequencing was performed on UCP1-, UCP1+, and UCP1++ FAPs to evaluate distinct characteristics of each population. Exosomes were harvested from UCP1++ FAP-BAT exosomes (Exo-FB) as well as UCP1- non-FAP-BAT exosomes (Exo-nFB) cells using cushioned-density gradient ultracentrifugation and used to treat C2C12 cells and mouse embryonic fibroblasts in vitro, and the myotube fusion index was assessed. Exo-FB and Exo-nFB were then used to treat wild type C57B/L6J mice that had undergone a massive rotator cuff tear. At 6 weeks mice were sacrificed, and supraspinatus muscles were harvested and analyzed for muscle atrophy, fibrosis, fatty infiltration, and UCP1 expression. Single-cell RNA sequencing was then performed on FAPs isolated from human muscle that were treated with the beta-agonist formoterol or standard media to assess for the presence of a parallel promyogenic subpopulation of FAP-BAT cells in humans. RESULTS: Flow cytometry analysis of sorted UCP1 reporter mouse FAPs revealed a trimodal distribution of UCP1 signal intensity, which correlated with 3 distinct transcriptomic profiles characterized with bulk RNA sequencing. UCP1++ cells were marked by high mitochondrial gene expression, BAT markers, and exosome surface makers; UCP1- cells were marked by fibrogenic markers; and UCP1+ cells were characterized differential enrichment of white adipose tissue markers. Exo-FB treatment of C2C12 cells resulted in robust myotube fusion, while treatment of mouse embryonic fibroblasts resulted in differentiation into myotubes. Treatment of cells with Exo-nFB resulted in poor myotube formation. Mice that were treated with Exo-FB at the time of rotator cuff injury demonstrated markedly reduced muscle atrophy and fatty infiltration as compared with treatment with Exo-nFB or phosphate-buffered saline. Single-cell RNA sequencing of human FAPs from the rotator cuff revealed 6 distinct subpopulations of human FAPs, with one subpopulation demonstrating the presence of UCP1+ beige adipocytes with a distinct profile of BAT, mitochondrial, and extracellular vesicle-associated markers. CONCLUSION: FAP-BAT cells form a subpopulation of FAPs with upregulated beige gene expression and exosome production that mediate promyogenic effects in vitro and in vivo, and they are present as a transcriptomically similar subpopulation of FAPs in humans. CLINICAL RELEVANCE: FAP-BAT cells and their exosomes represent a potential therapeutic avenue for treating rotator cuff muscle degeneration.

Rotator Cuff Tear Size Regulates Fibroadipogenic Progenitor Number and Gene Expression Profile in the Supraspinatus Independent of Patient Age.

BACKGROUND: Fatty infiltration of rotator cuff muscle is a limiting factor in the success of repairs. Fibroadipogenic progenitors (FAPs) are a population of stem cells within the rotator cuff that can differentiate into white adipocytes, fibroblasts, and beige adipocytes. The effects of patient age and rotator cuff tendon tear size on the number, differentiation patterns, and gene expression profiles of FAPs have not yet been analyzed. PURPOSE: To determine if patient age and rotator cuff tear size independently regulate FAP number, differentiation patterns, and gene expression profiles. STUDY DESIGN: Controlled laboratory study. METHODS: Supraspinatus muscle samples were collected from 26 patients between the ages of 42 and 76 years with partial- or full-thickness rotator cuff tears. FAPs were quantified using fluorescence-activated cell sorting. Gene expression analysis was performed across a custom 96-gene panel using NanoString. In vitro differentiation assays of FAPs were conducted using adipogenic, fibrogenic, and beige-inducing (amibegron-treated) media, and quantitative polymerase chain reaction was used to assess gene expression differences between adipogenic and amibegron media conditions. Multivariable linear regressions were performed using Stata to independently analyze the effects of age and rotator cuff tear size on FAP number, differentiation, and gene expression. RESULTS: Increasing age and tear size were independently correlated with increased FAP number (ßage = 0.21, P = .03; ßtear size = 3.86, P = .05). There was no clear association between age and gene expression of freshly sorted FAPs. Under adipogenic and fibrogenic media conditions, increasing age and tear size were independently associated with increased adipogenic and fibrogenic differentiation of FAPs. Under amibegron treatment conditions, age positively correlated with increased beige differentiation (ß = 1.03; P < .0001), while increasing tear size showed a trend toward decreased beige differentiation (ß = -4.87; P = .1). When gene expression patterns between adipogenic and amibegron media conditions were compared, larger tear size strongly inhibited beige gene expression, while advanced age did not. CONCLUSION: Patient age and rotator cuff tear size independently regulated FAP number, differentiation, and gene expression. Age and tear size were positively correlated with increased FAP number and fibrogenic/adipogenic differentiation. Advancing patient age did not limit FAP beige differentiation and gene expression, while increasing rotator cuff tear size strongly inhibited these processes.

Direct Tie2 Agonists Stabilize Vasculature for the Treatment of Diabetic Macular Edema.

Purpose: Diabetic macular edema (DME) is the leading cause of vision loss and blindness among working-age adults. Although current intravitreal anti-vascular endothelial growth factor (VEGF) therapies improve vision for many patients with DME, approximately half do not achieve the visual acuity required to drive. We therefore sought additional approaches to resolve edema and improve vision for these patients. Methods: We explored direct agonists of Tie2, a receptor known to stabilize vasculature and prevent leakage. We identified a multivalent PEG-Fab conjugate, Tie2.1-hexamer, that oligomerizes Tie2 and drives receptor activation and characterized its activities in vitro and in vivo. Results: Tie2.1-hexamer normalized and stabilized intercellular junctions of stressed endothelial cell monolayers in vitro, suppressed vascular leak in mice under conditions where anti-VEGF alone was ineffective, and demonstrated extended ocular exposure and robust pharmacodynamic responses in non-human primates. Conclusions: Tie2.1-hexamer directly activates the Tie2 pathway, reduces vascular leak, and is persistent within the vitreal humor. Translational Relevance: Our study presents a promising potential therapeutic for the treatment of DME.

Proximal Humerus Fractures in the Pediatric Population.

PURPOSE OF REVIEW: The purpose of this review is to review recent literature focusing on proximal humerus anatomy, epidemiology of these fractures, diagnosis and treatment options, and clinical outcomes. RECENT FINDINGS: Non- or minimally displaced proximal humerus fractures treated nonoperatively do not lead to short- or long-term complication and do not cross over to operative treatment. There is a higher rate of operative management with older age, increased injury severity score, treatment at an adult hospital, and private insurance. Operative management is preferred with closed or open reduction and percutaneous pinning, but elastic nailing and plate fixation are other options with good postoperative outcomes. Pediatric proximal humerus fractures occur after fall onto the affected shoulder or arm. Diagnosis is usually made with radiographs. Understanding the proximal humerus anatomy is critical to the proper management of these injuries to aid reduction and predict remodeling potential. There is considerable debate around the management of proximal humerus fractures in the pediatric population. Treatment is based on patient age, fracture displacement, and remodeling capacity. Nonoperative management is successful in younger patients or less displaced fractures, and operative management is usually considered in older patients with more displaced fractures.

Cross-species screening platforms identify EPS-8 as a critical link for mitochondrial stress and actin stabilization.

The dysfunction of mitochondria is associated with the physiological consequences of aging and many age-related diseases. Therefore, critical quality control mechanisms exist to protect mitochondrial functions, including the unfolded protein response of the mitochondria (UPRMT). However, it is still unclear how UPRMT is regulated in mammals with mechanistic discrepancies between previous studies. Here, we reasoned that a study of conserved mechanisms could provide a uniquely powerful way to reveal previously uncharacterized components of the mammalian UPRMT. We performed cross-species comparison of genetic requirements for survival under­and in response to­mitochondrial stress between karyotypically normal human stem cells and the nematode Caenorhabditis elegans. We identified a role for EPS-8/EPS8 (epidermal growth factor receptor pathway substrate 8), a signaling protein adaptor, in general mitochondrial homeostasis and UPRMT regulation through integrin-mediated remodeling of the actin cytoskeleton. This study also highlights the use of cross-species comparisons in genetic screens to interrogate cellular pathways.