Exercise to Mend Aged-tissue Crosstalk in Bone Targeting Osteoporosis & Osteoarthritis.

Aging induces alterations in bone structure and strength through a multitude of processes, exacerbating common aging- related diseases like osteoporosis and osteoarthritis. Cellular hallmarks of aging are examined, as related to bone and the marrow microenvironment, and ways in which these might contribute to a variety of age-related perturbations in osteoblasts, osteocytes, marrow adipocytes, chondrocytes, osteoclasts, and their respective progenitors. Cellular senescence, stem cell exhaustion, mitochondrial dysfunction, epigenetic and intracellular communication changes are central pathways and recognized as associated and potentially causal in aging. We focus on these in musculoskeletal system and highlight knowledge gaps in the literature regarding cellular and tissue crosstalk in bone, cartilage, and the bone marrow niche. While senolytics have been utilized to target aging pathways, here we propose non-pharmacologic, exercise-based interventions as prospective "senolytics" against aging effects on the skeleton. Increased bone mass and delayed onset or progression of osteoporosis and osteoarthritis are some of the recognized benefits of regular exercise across the lifespan. Further investigation is needed to delineate how cellular indicators of aging manifest in bone and the marrow niche and how altered cellular and tissue crosstalk impact disease progression, as well as consideration of exercise as a therapeutic modality, as a means to enhance discovery of bone-targeted therapies.

Aging and the emerging role of cellular senescence in osteoarthritis.

OBJECTIVE: The correlation between age and incidence of osteoarthritis (OA) is well known but the causal mechanisms involved are not completely understood. This narrative review summarizes selected key findings from the past 30 years that have elucidated key aspects of the relationship between aging and OA. METHODS: The peer-reviewed English language literature was searched on PubMed using keywords including senescence, aging, cartilage, and osteoarthritis, for original studies and reviews published from 1993 to 2023 with a major focus on more recent studies. Manuscripts most relevant to aging and OA that examined one or more of the hallmarks of aging were selected for further review. RESULTS: All proposed hallmarks of aging have been observed in articular cartilage and some have also been described in other joint tissues. Hallmarks include genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, deregulated nutrient sensing, mitochondrial dysfunction, cellular senescence, stem cell exhaustion, altered intercellular communication, disabled macroautophagy, chronic inflammation, and dysbiosis. There is evidence that these age-related changes contribute to the development of OA in part by promoting cellular senescence. Senescence may therefore serve as a downstream mediator that connects numerous aging hallmarks to OA, likely through the senescence-associated secretory phenotype that is characterized by increased production of proinflammatory cytokines and matrix metalloproteinases. CONCLUSIONS: Progress over the past 30 years has provided the foundation for emerging therapies, such as senolytics and senomorphics, that hold promise for OA disease modification. Mechanistic studies utilizing physiologically-aged animals and cadaveric human joint tissues will be important for continued progress.

Epigenetics as a mediator of genetic risk in osteoarthritis: role during development, homeostasis, aging, and disease progression.

The identification of genomic loci that are associated with osteoarthritis (OA) has provided a starting point for understanding how genetic variation activates catabolic processes in the joint. However, genetic variants can only alter gene expression and cellular function when the epigenetic environment is permissive to these effects. In this review, we provide examples of how epigenetic shifts at distinct life stages can alter the risk for OA, which we posit is critical for the proper interpretation of genome-wide association studies (GWAS). During development, intensive work on the growth and differentiation factor 5 (GDF5) locus has revealed the importance of tissue-specific enhancer activity in controlling both joint development and the subsequent risk for OA. During homeostasis in adults, underlying genetic risk factors may help establish beneficial or catabolic "set points" that dictate tissue function, with a strong cumulative effect on OA risk. During aging, methylation changes and the reorganization of chromatin can "unmask" the effects of genetic variants. The destructive function of variants that alter aging would only mediate effects after reproductive competence and thus avoid any evolutionary selection pressure, as consistent with larger frameworks of biological aging and its relationship to disease. A similar "unmasking" may occur during OA progression, which is supported by the finding of distinct expression quantitative trait loci (eQTLs) in chondrocytes depending on the degree of tissue degradation. Finally, we propose that massively parallel reporter assays (MPRAs) will be a valuable tool to test the function of putative OA GWAS variants in chondrocytes from different life stages.

Cartilage-specific Sirt6 deficiency represses IGF-1 and enhances osteoarthritis severity in mice.

OBJECTIVES: Prior studies noted that chondrocyte SIRT6 activity is repressed in older chondrocytes rendering cells susceptible to catabolic signalling events implicated in osteoarthritis (OA). This study aimed to define the effect of Sirt6 deficiency on the development of post-traumatic and age-associated OA in mice. METHODS: Male cartilage-specific Sirt6-deficient mice and Sirt6 intact controls underwent destabilisation of the medial meniscus (DMM) or sham surgery at 16 weeks of age and OA severity was analysed at 6 and 10 weeks postsurgery. Age-associated OA was assessed in mice aged 12 and 18 months of age. OA severity was analysed by micro-CT, histomorphometry and scoring of articular cartilage structure, toluidine blue staining and osteophyte formation. SIRT6-regulated pathways were analysed in human chondrocytes by RNA-sequencing, qRT-PCR and immunoblotting. RESULTS: Sirt6-deficient mice displayed enhanced DMM-induced OA severity and accelerated age-associated OA when compared with controls, characterised by increased cartilage damage, osteophyte formation and subchondral bone sclerosis. In chondrocytes, RNA-sequencing revealed that SIRT6 depletion significantly repressed cartilage extracellular matrix (eg, COL2A1) and anabolic growth factor (eg, insulin-like growth factor-1 (IGF-1)) gene expression. Gain-of-function and loss-of-function studies in chondrocytes demonstrated that SIRT6 depletion attenuated, whereas adenoviral overexpression or MDL-800-induced SIRT6 activation promoted IGF-1 signalling by increasing Aktser473 phosphorylation. CONCLUSIONS: SIRT6 deficiency increases post-traumatic and age-associated OA severity in vivo. SIRT6 profoundly regulated the pro-anabolic and pro-survival IGF-1/Akt signalling pathway and suggests that preserving the SIRT6/IGF-1/Akt axis may be necessary to protect cartilage from injury-associated or age-associated OA. Targeted therapies aimed at increasing SIRT6 function could represent a novel strategy to slow or stop OA.

Cells exhibiting strong p16INK4a promoter activation in vivo display features of senescence.

The activation of cellular senescence throughout the lifespan promotes tumor suppression, whereas the persistence of senescent cells contributes to aspects of aging. This theory has been limited, however, by an inability to identify and isolate individual senescent cells within an intact organism. Toward that end, we generated a murine reporter strain by "knocking-in" a fluorochrome, tandem-dimer Tomato (tdTom), into exon 1α of the p16INK4a locus. We used this allele (p16tdTom ) for the enumeration, isolation, and characterization of individual p16INK4a -expressing cells (tdTom+). The half-life of the knocked-in transcript was shorter than that of the endogenous p16INK4a mRNA, and therefore reporter expression better correlated with p16INK4a promoter activation than p16INK4a transcript abundance. The frequency of tdTom+ cells increased with serial passage in cultured murine embryo fibroblasts from p16tdTom/+ mice. In adult mice, tdTom+ cells could be readily detected at low frequency in many tissues, and the frequency of these cells increased with aging. Using an in vivo model of peritoneal inflammation, we compared the phenotype of cells with or without activation of p16INK4a and found that tdTom+ macrophages exhibited some features of senescence, including reduced proliferation, senescence-associated ß-galactosidase (SA-ß-gal) activation, and increased mRNA expression of a subset of transcripts encoding factors involved in SA-secretory phenotype (SASP). These results indicate that cells harboring activation of the p16INK4a promoter accumulate with aging and inflammation in vivo, and display characteristics of senescence.

Controlled induction and targeted elimination of p16INK4a-expressing chondrocytes in cartilage explant culture.

Cellular senescence is a phenotypic state that contributes to age-related diseases through the secretion of matrix-degrading and inflammatory molecules. An emerging therapeutic strategy for osteoarthritis (OA) is to selectively eliminate senescent cells by initiating apoptosis. This study establishes a cartilage explant model of senescence induction and senolytic clearance using p16Ink4a expression as a biomarker of senescence. Growth-factor stimulation of explants increased the expression of p16Ink4a at both the mRNA and protein levels. Applying this culture system to cartilage from p16tdTom reporter mice (a knockin allele with tdTomato fluorescent protein regulated by the endogenous p16Ink4a promoter) demonstrated the emergence of a p16-high population that was quantified using flow cytometry for tdTomato. Cell sorting was used to separate chondrocytes based on tdTomato fluorescence and p16-high cells showed higher senescence-associated ß-galactosidase activity and increased gene expression of the senescence-associated secretory phenotype as compared with p16-low cells. The potential for effective senolysis within the cartilage extracellular matrix was assessed using navitoclax (ABT-263). Navitoclax treatment reduced the percentage of p16-high cells from 17.9 to 6.1% (mean of 13 matched pairs; P < 0.001) and increased cleaved caspase-3 confirmed apoptotic activity. Together, these findings establish a physiologically relevant cartilage explant model for testing the induction and elimination of senescent chondrocytes, which will support investigations of senolytic therapy for OA.-Sessions, G. A., Copp, M. E., Liu, J.-Y., Sinkler, M. A., D'Costa, S., Diekman, B. O. Controlled induction and targeted elimination of p16INK4a-expressing chondrocytes in cartilage explant culture.

Microfluidics for the study of mechanotransduction.

Mechanical forces regulate a diverse set of biological processes at cellular, tissue, and organismal length scales. Investigating the cellular and molecular mechanisms that underlie the conversion of mechanical forces to biological responses is challenged by limitations of traditional animal models and in vitro cell culture, including poor control over applied force and highly artificial cell culture environments. Recent advances in fabrication methods and material processing have enabled the development of microfluidic platforms that provide precise control over the mechanical microenvironment of cultured cells. These devices and systems have proven to be powerful for uncovering and defining mechanisms of mechanotransduction. In this review, we first give an overview of the main mechanotransduction pathways that function at sites of cell adhesion, many of which have been investigated with microfluidics. We then discuss how distinct microfluidic fabrication methods can be harnessed to gain biological insight, with description of both monolithic and replica molding approaches. Finally, we present examples of how microfluidics can be used to apply both solid forces (substrate mechanics, strain, and compression) and fluid forces (luminal, interstitial) to cells. Throughout the review, we emphasize the advantages and disadvantages of different fabrication methods and applications of force in order to provide perspective to investigators looking to apply forces to cells in their own research.

Targeting aging for disease modification in osteoarthritis.

PURPOSE OF REVIEW: Age is a key risk factor for the development of osteoarthritis and age-related changes within the joint might represent targets for therapy. The recent literature was reviewed to find studies that provide new insight into the role of aging in osteoarthritis, with a focus on the potential for disease modification. RECENT FINDINGS: Preclinical studies using isolated cells and animal models provide evidence that two hallmarks of aging (cellular senescence and mitochondrial dysfunction) contribute to the development of osteoarthritis. Senescent cells secrete pro-inflammatory mediators and matrix degrading enzymes, and killing these cells with 'senolytic' compounds has emerged as a potential disease-modifying therapy. Mitochondrial dysfunction is associated with increased levels of reactive oxygen species (ROS) that can promote osteoarthritis by disrupting homeostatic intracellular signaling. Reducing ROS production in the mitochondria, stimulating antioxidant gene expression through Nrf2 activation, or inhibiting specific redox-sensitive signaling proteins represent additional approaches to disease modification in osteoarthritis that require further investigation. SUMMARY: Although no human clinical trials for osteoarthritis have specifically targeted aging, preclinical studies suggest that targeting cellular senescence and/or mitochondrial dysfunction and the effects of excessive ROS may lead to novel interventions that could slow the progression of osteoarthritis.

Cartilage tissue engineering using differentiated and purified induced pluripotent stem cells.

The development of regenerative therapies for cartilage injury has been greatly aided by recent advances in stem cell biology. Induced pluripotent stem cells (iPSCs) have the potential to provide an abundant cell source for tissue engineering, as well as generating patient-matched in vitro models to study genetic and environmental factors in cartilage repair and osteoarthritis. However, both cell therapy and modeling approaches require a purified and uniformly differentiated cell population to predictably recapitulate the physiological characteristics of cartilage. Here, iPSCs derived from adult mouse fibroblasts were chondrogenically differentiated and purified by type II collagen (Col2)-driven green fluorescent protein (GFP) expression. Col2 and aggrecan gene expression levels were significantly up-regulated in GFP+ cells compared with GFP- cells and decreased with monolayer expansion. An in vitro cartilage defect model was used to demonstrate integrative repair by GFP+ cells seeded in agarose, supporting their potential use in cartilage therapies. In chondrogenic pellet culture, cells synthesized cartilage-specific matrix as indicated by high levels of glycosaminoglycans and type II collagen and low levels of type I and type X collagen. The feasibility of cell expansion after initial differentiation was illustrated by homogenous matrix deposition in pellets from twice-passaged GFP+ cells. Finally, atomic force microscopy analysis showed increased microscale elastic moduli associated with collagen alignment at the periphery of pellets, mimicking zonal variation in native cartilage. This study demonstrates the potential use of iPSCs for cartilage defect repair and for creating tissue models of cartilage that can be matched to specific genetic backgrounds.

Multiplexed single-cell imaging reveals diverging subpopulations with distinct senescence phenotypes during long-term senescence induction.

Cellular senescence is a phenotypic state that contributes to the progression of age-related disease through secretion of pro-inflammatory factors known as the senescence associated secretory phenotype (SASP). Understanding the process by which healthy cells become senescent and develop SASP factors is critical for improving the identification of senescent cells and, ultimately, understanding tissue dysfunction. Here, we reveal how the duration of cellular stress modulates the SASP in distinct subpopulations of senescent cells. We used multiplex, single-cell imaging to build a proteomic map of senescence induction in human epithelial cells induced to senescence over the course of 31 days. We map how the expression of SASP proteins increases alongside other known senescence markers such as p53, p21, and p16 INK4a . The aggregated population of cells responded to etoposide with an accumulation of stress response factors over the first 11 days, followed by a plateau in most proteins. At the single-cell level, however, we identified two distinct senescence cell populations, one defined primarily by larger nuclear area and the second by higher protein concentrations. Trajectory inference suggested that cells took one of two discrete molecular paths from unperturbed healthy cells, through a common transitional subpopulation, and ending at the discrete terminal senescence phenotypes. Our results underscore the importance of using single-cell proteomics to identify the mechanistic pathways governing the transition from senescence induction to a mature state of senescence characterized by the SASP.

Use of a novel magnetically actuated compression system to study the temporal dynamics of axial and lateral strain in human osteochondral plugs.

The high water content of articular cartilage allows this biphasic tissue to withstand large compressive loads through fluid pressurization. The system presented here, termed the "MagnaSquish", provides new capabilities for quantifying the effect of rehydration on cartilage behavior during cyclic loading. An imbalanced rate of fluid exudation during load and fluid re-entry during recovery can lead to the accumulation of strain during successive loading cycles - a phenomenon known as ratcheting. Typical experimental systems for cartilage biomechanics use continuous contact between the platen and sample, which may affect tissue rehydration by compressing the top layer of cartilage and slowing fluid re-entry. To address this limitation, we developed a magnetically actuated device that provides full lift-off of the platen in between loading cycles. We investigated strain accumulation in cadaveric human osteochondral plugs during 750 loading cycles, with two dimensional profiles of the cartilage captured at 30 frames per second throughout loading and 10 min of additional free swelling recovery. Axial and lateral strain measurements were extracted from the tissue profiles using a UNet-based deep learning algorithm to circumvent manual tracing. We observed increased axial strain accumulation with shorter inter-cycle recovery, with static loading serving as the extreme case of zero recovery. The loading waveform during the 750 cycles dictated the pace of the recovery during the extended free swelling period, as shorter inter-cycle recovery led to more persistent axial strain accumulation for up to five minutes. This work showcases the importance of fluid re-entry in resisting strain accumulation during cyclical compression.

Response eQTLs, chromatin accessibility, and 3D chromatin structure in chondrocytes provide mechanistic insight into osteoarthritis risk.

Osteoarthritis (OA) poses a significant healthcare burden with limited treatment options. While genome-wide association studies (GWAS) have identified over 100 OA-associated loci, translating these findings into therapeutic targets remains challenging. Integrating expression quantitative trait loci (eQTL), 3D chromatin structure, and other genomic approaches with OA GWAS data offers a promising approach to elucidate disease mechanisms; however, comprehensive eQTL maps in OA-relevant tissues and conditions remain scarce. We mapped gene expression, chromatin accessibility, and 3D chromatin structure in primary human articular chondrocytes in both resting and OA-mimicking conditions. We identified thousands of differentially expressed genes, including those associated with differences in sex and age. RNA-seq in chondrocytes from 101 donors across two conditions uncovered 3782 unique eGenes, including 420 that exhibited strong and significant condition-specific effects. Colocalization with OA GWAS signals revealed 13 putative OA risk genes, 10 of which have not been previously identified. Chromatin accessibility and 3D chromatin structure provided insights into the mechanisms and conditional specificity of these variants. Our findings shed light on OA pathogenesis and highlight potential targets for therapeutic development. Highlights: ∘ Comprehensive analysis of sex- and age-related global gene expression in human chondrocytes revealed differences that correlate with osteoarthritis ∘ First response eQTLs in chondrocytes treated with an OA-related stimulus ∘ Deeply sequenced Hi-C in resting and activated chondrocytes helps connect OA risk variants to their putative causal genes ∘ Colocalization analysis reveals 13 (including 10 novel) putative OA risk genes.

Stem cell-based therapies for osteoarthritis: challenges and opportunities.

PURPOSE OF REVIEW: Regenerative medicine offers the exciting potential of developing alternatives to total joint replacement for treating osteoarthritis. In this article, we highlight recent work that addresses key challenges of stem cell-based therapies for osteoarthritis and provide examples of innovative ways in which stem cells can aid in the treatment of osteoarthritis. RECENT FINDINGS: Significant progress has been made in understanding the challenges to successful stem cell therapy, such as the effects of age or disease on stem cell properties, altered stem cell function due to an inflammatory joint environment and phenotypic instability in vivo. Novel scaffold designs have been shown to enhance the mechanical properties of tissue-engineered cartilage and have also improved the integration of newly formed tissue within the joint. Emerging strategies such as injecting stem cells directly into the joint, manipulating endogenous stem cells to enhance regenerative capacity and utilizing stem cells for drug discovery have expanded the potential uses of stem cells in treating osteoarthritis. SUMMARY: Several recent studies have greatly advanced the development and preclinical evaluation of potential stem cell-based treatments for osteoarthritis through novel approaches focused on cell therapy, tissue engineering and drug discovery.

SIRT6 activation rescues the age-related decline in DNA damage repair in primary human chondrocytes.

While advanced age has long been recognized as the greatest risk factor for osteoarthritis (OA), the biological mechanisms behind this connection remain unclear. Previous work has demonstrated that chondrocytes from older cadaveric donors have elevated levels of DNA damage as compared to chondrocytes from younger donors. The purpose of this study was to determine whether a decline in DNA repair efficiency is one explanation for the accumulation of DNA damage with age, and to quantify the improvement in repair with activation of Sirtuin 6 (SIRT6). Using an acute irradiation model to bring the baseline level of all donors to the same starting point, this study demonstrates a decline in repair efficiency during aging when comparing chondrocytes from young (≤45 years old), middle-aged (50-65 years old), or older (>70 years old) cadaveric donors with no known history of OA or macroscopic cartilage degradation at isolation. Activation of SIRT6 in middle-aged chondrocytes with MDL-800 (20 µM) improved the repair efficiency, while inhibition with EX-527 (10 µM) inhibited the rate of repair and the increased the percentage of cells that retained high levels of damage. Treating chondrocytes from older donors with MDL-800 for 48 hours significantly reduced the amount of DNA damage, despite this damage having accumulated over decades. Lastly, chondrocytes isolated from the proximal femurs of mice between 4 months and 22 months of age revealed both an increase in DNA damage with aging, and a decrease in DNA damage following MDL-800 treatment.

Sirtuin 6 activation rescues the age-related decline in DNA damage repair in primary human chondrocytes.

While advanced age is widely recognized as the greatest risk factor for osteoarthritis (OA), the biological mechanisms behind this connection remain unclear. Previous work has demonstrated that chondrocytes from older cadaveric donors have elevated levels of DNA damage as compared to chondrocytes from younger donors. The purpose of this study was to determine whether a decline in DNA repair efficiency is one explanation for the accumulation of DNA damage with age, and to quantify the improvement in repair with activation of Sirtuin 6 (SIRT6). After acute damage with irradiation, DNA repair was shown to be more efficient in chondrocytes from young (≤45 years old) as compared to middle-aged (50-65 years old) or older (>70 years old) cadaveric donors. Activation of SIRT6 with MDL-800 improved the repair efficiency, while inhibition with EX-527 reduced the rate of repair and increased the percentage of cells that retain high levels of damage. In addition to affecting repair after acute damage, treating chondrocytes from older donors with MDL-800 for 48 hours significantly reduced the amount of baseline DNA damage. Chondrocytes isolated from the knees of mice between 4 months and 22 months of age revealed both an increase in DNA damage with aging, and a decrease in DNA damage following MDL-800 treatment. Lastly, treating murine cartilage explants with MDL-800 lowered the percentage of chondrocytes with high p16 promoter activity, which supports the concept that using SIRT6 activation to maintain low levels of DNA damage may prevent the initiation of senescence.

Redox-active endosomes mediate α5ß1 integrin signaling and promote chondrocyte matrix metalloproteinase production in osteoarthritis.

Mechanical cues sensed by integrins induce cells to produce proteases to remodel the extracellular matrix. Excessive protease production occurs in many degenerative diseases, including osteoarthritis, in which articular cartilage degradation is associated with the genesis of matrix protein fragments that can activate integrins. We investigated the mechanisms by which integrin signals may promote protease production in response to matrix changes in osteoarthritis. Using a fragment of the matrix protein fibronectin (FN) to activate the α5ß1 integrin in primary human chondrocytes, we found that endocytosis of the integrin and FN fragment complex drove the production of the matrix metalloproteinase MMP-13. Activation of α5ß1 by the FN fragment, but not by intact FN, was accompanied by reactive oxygen species (ROS) production initially at the cell surface, then in early endosomes. These ROS-producing endosomes (called redoxosomes) contained the integrin-FN fragment complex, the ROS-producing enzyme NADPH oxidase 2 (NOX2), and SRC, a redox-regulated kinase that promotes MMP-13 production. In contrast, intact FN was endocytosed and trafficked to recycling endosomes without inducing ROS production. Articular cartilage from patients with osteoarthritis showed increased amounts of SRC and the NOX2 complex component p67phox. Furthermore, we observed enhanced localization of SRC and p67phox at early endosomes, suggesting that redoxosomes could transmit and sustain integrin signaling in response to matrix damage. This signaling mechanism not only amplifies the production of matrix-degrading proteases but also establishes a self-perpetuating cycle that contributes to the ongoing degradation of cartilage matrix in osteoarthritis.

Patient-derived extracellular matrix demonstrates role of COL3A1 in blood vessel mechanics.

Vascular Ehlers-Danlos Syndrome (vEDS) is a rare autosomal dominant disease caused by mutations in the COL3A1 gene, which renders patients susceptible to aneurysm and arterial dissection and rupture. To determine the role of COL3A1 variants in the biochemical and biophysical properties of human arterial ECM, we developed a method for synthesizing ECM directly from vEDS donor fibroblasts. We found that the protein content of the ECM generated from vEDS donor fibroblasts differed significantly from ECM from healthy donors, including upregulation of collagen subtypes and other proteins related to ECM structural integrity. We further found that ECM generated from a donor with a glycine substitution mutation was characterized by increased glycosaminoglycan content and unique viscoelastic mechanical properties, including increased time constant for stress relaxation, resulting in a decrease in migratory speed of human aortic endothelial cells when seeded on the ECM. Collectively, these results demonstrate that vEDS patient-derived fibroblasts harboring COL3A1 mutations synthesize ECM that differs in composition, structure, and mechanical properties from healthy donors. These results further suggest that ECM mechanical properties could serve as a prognostic indicator for patients with vEDS, and the insights provided by the approach demonstrate the broader utility of cell-derived ECM in disease modeling. STATEMENT OF SIGNIFICANCE: The role of collagen III ECM mechanics remains unclear, despite reported roles in diseases including fibrosis and cancer. Here, we generate fibrous, collagen-rich ECM from primary donor cells from patients with vascular Ehlers-Danlos syndrome (vEDS), a disease caused by mutations in the gene that encodes collagen III. We observe that ECM grown from vEDS patients is characterized by unique mechanical signatures, including altered viscoelastic properties. By quantifying the structural, biochemical, and mechanical properties of patient-derived ECM, we identify potential drug targets for vEDS, while defining a role for collagen III in ECM mechanics more broadly. Furthermore, the structure/function relationships of collagen III in ECM assembly and mechanics will inform the design of substrates for tissue engineering and regenerative medicine.

Pilot comparison of outcome measures across chemical and surgical experimental models of chronic osteoarthritis in the rat (Rattus norvegicus).

Relatively little work has evaluated both the disease of osteoarthritis (OA) and clinically-relevant pain outcome measures across different OA models in rats. The objective of this study was to compare sensitivity, pain, and histological disease severity across chemical and surgical models of OA in the rat. Stifle OA was induced in Sprague-Dawley rats via intraarticular injection of monoiodoacetate (MIA) or surgical transection of anterior cruciate ligament and/or destabilization of medial meniscus (ACL+DMM or DMM alone). Reflexive (e.g., mechanical and thermal stimuli) measures of sensitivity and non-reflexive assays (e.g., lameness, static hindlimb weight-bearing asymmetry, dynamic gait analysis) of pain were measured over time. Joint degeneration was assessed histologically. Six-weeks post OA-induction, the ACL+DMM animals had significantly greater visually observed lameness than MIA animals; however, both ACL+DMM and MIA animals showed equal pain as measured by limb use during ambulation and standing. The MIA animals showed increased thermal, but not mechanical, sensitivity compared to ACL+DMM animals. Joint degeneration was significantly more severe in the MIA model at 6 weeks. Our pilot data suggest both the ACL+DMM and MIA models are equal in terms of clinically relevant pain behaviors, but the MIA model is associated with more severe histological changes over time potentially making it more suitable for screening disease modifying agents. Future work should further characterize each model in terms of complex pain behaviors and biochemical, molecular, and imaging analysis of the sensory system and joint tissues, which will allow for more informed decisions associated with model selection and investigative outcomes.