Gene therapy for fat-1 prevents obesity-induced metabolic dysfunction, cellular senescence, and osteoarthritis.

Obesity is one of the primary risk factors for osteoarthritis (OA), acting through cross talk among altered biomechanics, metabolism, adipokines, and dietary free fatty acid (FA) composition. Obesity and aging have been linked to cellular senescence in various tissues, resulting in increased local and systemic inflammation and immune dysfunction. We hypothesized that obesity and joint injury lead to cellular senescence that is typically associated with increased OA severity or with aging and that the ratio of omega-6 (ω-6) to omega-3 (ω-3) FAs regulates these pathologic effects. Mice were placed on an ω-6-rich high-fat diet or a lean control diet and underwent destabilization of the medial meniscus to induce OA. Obesity and joint injury significantly increased cellular senescence in subcutaneous and visceral fat as well as joint tissues such as synovium and cartilage. Using adeno-associated virus (AAV) gene therapy for fat-1, a fatty acid desaturase that converts ω-6 to ω-3 FAs, decreasing the serum ω-6:ω-3 FA ratio had a strong senomorphic and therapeutic effect, mitigating metabolic dysfunction, cellular senescence, and joint degeneration. In vitro coculture of bone marrow-derived macrophages and chondrocytes from control and AAV8-fat1-treated mice were used to examine the roles of various FA mediators in regulating chondrocyte senescence. Our results suggest that obesity and joint injury result in a premature "aging" of the joint as measured by senescence markers, and these changes can be ameliorated by altering FA composition using fat-1 gene therapy. These findings support the potential for fat-1 gene therapy to treat obesity- and/or injury-induced OA clinically.

Maternal high-fat high-sugar diet impairs bone quality and strength but not cartilage in aging mice.

Osteoarthritis (OA) is a prevalent aging disorder of synovial joints and recent work suggests that a parental high-fat diet increases OA severity following joint injury in offspring. We hypothesized that a maternal high-fat high-sugar (HFHS) diet would promote spontaneous osteoarthritis-related cartilage and bone changes in 1-year-old offspring. Female C57BL/6 J mice were placed on either a chow control or HFHS diet for 6 weeks before mating to a chow-fed C57BL/6 J male and maintained on their assigned diets throughout pregnancy and lactation. Male and female offspring were weaned onto a chow diet, raised to 1 year of age, and evaluated for cartilage and bone changes indicative of OA. However, offspring did not show early signs of OA as measured by histological Mankin scoring, mechanical testing of the pericellular matrix, histological synovitis scoring, or subchondral bone thickening as measured by microcomputed Tomography. On the other hand, male offspring from HFHS-fed dams had reduced trabecular bone quality in the tibial metaphysis and decreased cortical thickness. Although maternal HFHS diet did not impact trabecular or cortical bone quality in tibias of female offspring, the radii of these animals had decreased cortical thickness, increased medullary area, and impaired breaking strength compared to those of control-fed dams. Finally, we evaluated bone quality and strength in male and female F2 offspring and found that the grandmaternal diet modestly impacted radial bone geometry but not strength. Together these results suggest that maternal HFHS diet impairs F1 offspring skeletal integrity in a sex and bone site-specific manner.

A genome-engineered bioartificial implant for autoregulated anticytokine drug delivery.

Biologic drug therapies are increasingly used for inflammatory diseases such as rheumatoid arthritis but may cause significant adverse effects when delivered continuously at high doses. We used CRISPR-Cas9 genome editing of iPSCs to create a synthetic gene circuit that senses changing levels of endogenous inflammatory cytokines to trigger a proportional therapeutic response. Cells were engineered into cartilaginous constructs that showed rapid activation and recovery in response to inflammation in vitro or in vivo. In the murine K/BxN model of inflammatory arthritis, bioengineered implants significantly mitigated disease severity as measured by joint pain, structural damage, and systemic and local inflammation. Therapeutic implants completely prevented increased pain sensitivity and bone erosions, a feat not achievable by current clinically available disease-modifying drugs. Combination tissue engineering and synthetic biology promises a range of potential applications for treating chronic diseases via custom-designed cells that express therapeutic transgenes in response to dynamically changing biological signals.

Immunoengineering the next generation of arthritis therapies.

Immunoengineering continues to revolutionize healthcare, generating new approaches for treating previously intractable diseases, particularly in regard to cancer immunotherapy. In joint diseases, such as osteoarthritis (OA) and rheumatoid arthritis (RA), biomaterials and anti-cytokine treatments have previously been at that forefront of therapeutic innovation. However, while many of the existing anti-cytokine treatments are successful for a subset of patients, these treatments can also pose severe risks, adverse events and off-target effects due to continuous delivery at high dosages or a lack of disease-specific targets. The inadequacy of these current treatments has motivated the development of new immunoengineering strategies that offer safer and more efficacious alternative therapies through the precise and controlled targeting of specific upstream immune responses, including direct and mechanistically-driven immunoengineering approaches. Advances in the understanding of the immunomodulatory pathways involved in musculoskeletal disease, in combination with the growing emphasis on personalized medicine, stress the need for carefully considering the delivery strategies and therapeutic targets when designing therapeutics to better treat RA and OA. Here, we focus on recent advances in biomaterial and cell-based immunomodulation, in combination with genetic engineering, for therapeutic applications in joint diseases. The application of immunoengineering principles to the study of joint disease will not only help to elucidate the mechanisms of disease pathogenesis but will also generate novel disease-specific therapeutics by harnessing cellular and biomaterial responses. STATEMENT OF SIGNIFICANCE: It is now apparent that joint diseases such as osteoarthritis and rheumatoid arthritis involve the immune system at both local (i.e., within the joint) and systemic levels. In this regard, targeting the immune system using both biomaterial-based or cellular approaches may generate new joint-specific treatment strategies that are well-controlled, safe, and efficacious. In this review, we focus on recent advances in immunoengineering that leverage biomaterials and/or genetically engineered cells for therapeutic applications in joint diseases. The application of such approaches, especially synergistic strategies that target multiple immunoregulatory pathways, has the potential to revolutionize our understanding, treatment, and prevention of joint diseases.

Single-cell RNA sequencing reveals the induction of novel myeloid and myeloid-associated cell populations in visceral fat with long-term obesity.

Macrophages and other immune cells are important contributors to obesity-associated inflammation; however, the cellular identities of these specific populations remain unknown. In this study, we identified individual populations of myeloid cells found in mouse epididymal/visceral adipose tissue by single-cell RNA sequencing, immunofluorescence, and flow cytometry. Multiple canonical correlation analysis identified 11 unique myeloid and myeloid-associate cell populations. In obese mice, we detected an increased percentage of monocyte-derived pro-inflammatory cells expressing Cd9 and Trem2, as well as significantly decreased percentages of multiple cell populations, including tissue-resident cells expressing Lyve1, Mafb, and Mrc1. We have identified and validated a novel myeloid/macrophage population defined by Ly6a expression, exhibiting both myeloid and mesenchymal characteristics, which increased with obesity and showed high pro-fibrotic characteristics in vitro. Our mouse adipose tissue myeloid cell atlas provides an important resource to investigate obesity-associated inflammation and fibrosis.

Is Obesity a Disease of Stem Cells?

Obesity disrupts physiological homeostasis and alters both systemic and local microenvironments that impact stem cell plasticity and impair regenerative capacity. We present growing evidence that reveals the bidirectionality of obesity-induced stem cell dysfunction and how the molecular changes in stem cells residing in obese environments may accelerate disease severity.

Gene therapy for follistatin mitigates systemic metabolic inflammation and post-traumatic arthritis in high-fat diet-induced obesity.

Obesity-associated inflammation and loss of muscle function play critical roles in the development of osteoarthritis (OA); thus, therapies that target muscle tissue may provide novel approaches to restoring metabolic and biomechanical dysfunction associated with obesity. Follistatin (FST), a protein that binds myostatin and activin, may have the potential to enhance muscle formation while inhibiting inflammation. Here, we hypothesized that adeno-associated virus 9 (AAV9) delivery of FST enhances muscle formation and mitigates metabolic inflammation and knee OA caused by a high-fat diet in mice. AAV-mediated FST delivery exhibited decreased obesity-induced inflammatory adipokines and cytokines systemically and in the joint synovial fluid. Regardless of diet, mice receiving FST gene therapy were protected from post-traumatic OA and bone remodeling induced by joint injury. Together, these findings suggest that FST gene therapy may provide a multifactorial therapeutic approach for injury-induced OA and metabolic inflammation in obesity.

Intergenerational Transmission of Diet-Induced Obesity, Metabolic Imbalance, and Osteoarthritis in Mice.

OBJECTIVE: Obesity and osteoarthritis (OA) are 2 major public health issues affecting millions of people worldwide. Whereas parental obesity affects the predisposition to diseases such as cancer or diabetes in children, transgenerational influences on musculoskeletal conditions such as OA are poorly understood. This study was undertaken to assess the intergenerational effects of a parental/grandparental high-fat diet on the metabolic and skeletal phenotype, systemic inflammation, and predisposition to OA in 2 generations of offspring in mice. METHODS: Metabolic phenotype and predisposition to OA were investigated in the first and second (F1 and F2) generations of offspring (n = 10-16 mice per sex per diet) bred from mice fed a high-fat diet (HFD) or a low-fat control diet. OA was induced by destabilizing the medial meniscus. OA, synovitis, and adipose tissue inflammation were determined histologically, while bone changes were measured using micro-computed tomography. Serum and synovial cytokines were measured by multiplex assay. RESULTS: Parental high-fat feeding showed an intergenerational effect, with inheritance of increased weight gain (up to 19% in the F1 generation and 9% in F2), metabolic imbalance, and injury-induced OA in at least 2 generations of mice, despite the fact that the offspring were fed the low-fat diet. Strikingly, both F1 and F2 female mice showed an increased predisposition to injury-induced OA (48% higher predisposition in F1 and 19% in F2 female mice fed the HFD) and developed bone microarchitectural changes that were attributable to parental and grandparental high-fat feeding. CONCLUSION: The results of this study reveal a detrimental effect of parental HFD and obesity on the musculoskeletal integrity of 2 generations of offspring, indicating the importance of further investigation of these effects. An improved understanding of the mechanisms involved in the transmissibility of diet-induced changes through multiple generations may help in the development of future therapies that would target the effects of obesity on OA and related conditions.

Effects of dietary fatty acid content on humeral cartilage and bone structure in a mouse model of diet-induced obesity.

Obesity is a primary risk factor for osteoarthritis (OA), and previous studies have shown that dietary content may play an important role in the pathogenesis of cartilage and bone in knee OA. Several previous studies have shown that the ratio of ω-3 polyunsaturated fatty acids (PUFAs), ω-6 PUFAs, and saturated fatty acids can significantly influence bone structure and OA progression. However, the influence of obesity or dietary fatty acid content on shoulder OA is not well understood. The goal of this study was to investigate the role of dietary fatty acid content on bone and cartilage structure in the mouse shoulder in a model of diet-induced obesity. For 24 weeks, mice were fed control or high-fat diets supplemented with ω-3 PUFAs, ω-6 PUFAs, or saturated fatty acids. The humeral heads were analyzed for bone morphometry and mineral density by microCT. Cartilage structure and joint synovitis were determined by histological grading, and microscale mechanical properties of the cartilage extracellular and pericellular matrices were quantified using atomic force microscopy. Diet-induced obesity significantly altered bone morphology and mineral density in a manner that was dependent on dietary free fatty acid content. In general, high-fat diet groups showed decreased bone quality, with the ω-3 diet being partially protective. Cartilage mechanical properties and OA scores showed no changes with obesity or diet. These findings are consistent with clinical literature showing little if any relationship between obesity and shoulder OA (unlike knee OA), but suggest that diet-induced obesity may influence other joint tissues. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.

Physiologic and pathologic effects of dietary free fatty acids on cells of the joint.

Fatty acids (FAs) are potent organic compounds that not only can be used as an energy source during nutrient deprivation but are also involved in several essential signaling cascades in cells. Therefore, a balanced intake of different dietary FAs is critical for the maintenance of cellular functions and tissue homeostasis. A diet with an imbalanced fat composition creates a risk for developing metabolic syndrome and various musculoskeletal diseases, including osteoarthritis (OA). In this review, we summarize the current state of knowledge and mechanistic insights regarding the role of dietary FAs, such as saturated FAs, omega-6 polyunsaturated FAs (PUFAs), and omega-3 PUFAs on joint inflammation and OA pathogeneses. In particular, we review how different types of dietary FAs and their derivatives distinctly affect a variety of cells within the joint, including chondrocytes, osteoblasts, osteoclasts, and synoviocytes. Understanding the molecular mechanisms underlying the effects of FAs on metabolic behavior, anabolic, and catabolic processes, as well as the inflammatory response of joint cells, may help identify therapeutic targets for the prevention of metabolic joint diseases.

Regional Differences Between Perisynovial and Infrapatellar Adipose Tissue Depots and Their Response to Class II and Class III Obesity in Patients With Osteoarthritis.

OBJECTIVE: Obesity is associated with an increased risk of developing osteoarthritis (OA), which is postulated to be secondary to adipose tissue-dependent inflammation. Periarticular adipose tissue depots are present in synovial joints, but the association of this tissue with OA has not been extensively explored. The aim of this study was to investigate differences in local adipose tissue depots in knees with OA and characterize the changes related to class II and class III obesity in patients with end-stage knee OA. METHODS: Synovium and the infrapatellar fat pad (IPFP) were collected during total knee replacement from 69 patients with end-stage OA. Histologic changes, changes in gene and protein expression of adiponectin, peroxisome proliferator-activated receptor γ (PPARγ), and Toll-like receptor 4 (TLR-4), and immune cell infiltration into the adipose tissue were investigated. RESULTS: IPFP and synovium adipose tissue depots differed significantly and were influenced by the patient's body mass index. Compared to adipocytes from the IPFP and synovium of lean patients, adipocytes from the IPFP of obese patients were significantly larger and the synovium of obese patients displayed marked fibrosis, increased macrophage infiltration, and higher levels of TLR4 gene expression. The adipose-related markers PPARγ in the IPFP and adiponectin and PPARγ in the synovium were expressed at lower levels in obese patients compared to lean patients. Furthermore, there were increased numbers of CD45+ hematopoietic cells, CD45+CD14+ total macrophages, and CD14+CD206+ M2-type macrophages in both the IPFP and synovial tissue of obese patients. CONCLUSION: These differences suggest that IPFP and synovium may contain 2 different white adipose tissue depots and support the theory of inflammation-induced OA in patients with class II or III obesity. These findings warrant further investigation as a potentially reversible, or at least suppressible, cause of OA in obese patients.