Post-traumatic Osteoarthritis Model Research Articles

Comparative transcriptomic analysis of articular cartilage of post-traumatic osteoarthritis mouse models.

Animal models of post-traumatic osteoarthritis (PTOA) recapitulate the pathological changes observed in human PTOA. Here, skeletally mature C57Bl6 mice were subjected to either the rapid-onset, non-surgical, mechanical anterior cruciate ligament (ACL) rupture or surgical destabilisation of the medial meniscus (DMM) models. Transcriptome profiling of micro-dissected cartilage at day 7 and 42 post-ACL and DMM procedure respectively, showed that the two models were comparable and highly correlative (Spearman R =0.82, p<2.2E-16). Gene ontology enrichment analysis identified similarly enriched pathways, which were overrepresented by anabolic terms. To address the transcriptome changes more completely in the ACL model we also performed small RNA-seq, describing the first microRNA profile of this model. miR-199-5p was amongst the most abundant yet differentially expressed microRNAs and its inhibition in primary human chondrocytes led to a comparable transcriptome response to that observed in both human 'OA damaged vs intact cartilage' and murine DMM cartilage datasets. CELSR1, GIT1, ECE1 and SOS2 were all experimentally verified as novel miR-199-5p targets. Together, these data support the use of the ACL rupture model as a non-invasive companion to DMM.

Loss of collagen content is localized near cartilage lesions on the day of injurious loading and intensified on day 12.

Joint injury can lead to articular cartilage damage, excessive inflammation, and post-traumatic osteoarthritis (PTOA). Collagen is an essential component for cartilage function, yet current literature has limited understanding of how biochemical and biomechanical factors contribute to collagen loss in injured cartilage. Our aim was to investigate spatially dependent changes in collagen content and collagen integrity of injured cartilage, with an explant model of early-stage PTOA. We subjected calf knee cartilage explants to combinations of injurious loading (INJ), interleukin-1α-challenge (IL) and physiological cyclic loading (CL). Using Fourier transform infrared microspectroscopy, collagen content (Amide I band) and collagen integrity (Amide II/1338 cm-1 ratio) were estimated on days 0 and 12 post-injury. We found that INJ led to lower collagen content near lesions compared to intact regions on day 0 (p < 0.001). On day 12, near-lesion collagen content was lower compared to day 0 (p < 0.05). Additionally, on day 12, INJ, IL, and INJ + IL groups exhibited lower collagen content along most of tissue depth compared to free-swelling control group (p < 0.05). CL groups showed higher collagen content along most of tissue depth compared to corresponding groups without CL (p < 0.05). Immunohistochemical analysis revealed higher MMP-1 and MMP-3 staining intensities localized within cell lacunae in INJ group compared to CTRL group on day 0. Our results suggest that INJ causes rapid loss of collagen content near lesions, which is intensified on day 12. Additionally, CL could mitigate the loss of collagen content at intact regions after 12 days.

Degradation of lubricating molecules in synovial fluid alters chondrocyte sensitivity to shear strain.

Articular joints facilitate motion and transfer loads to underlying bone through a combination of cartilage tissue and synovial fluid, which together generate a low-friction contact surface. Traumatic injury delivered to cartilage and the surrounding joint capsule causes secretion of proinflammatory cytokines by chondrocytes and the synovium, triggering cartilage matrix breakdown and impairing the ability of synovial fluid to lubricate the joint. Once these inflammatory processes become chronic, posttraumatic osteoarthritis (PTOA) development begins. However, the exact mechanism by which negative alterations to synovial fluid leads to PTOA pathogenesis is not fully understood. We hypothesize that removing the lubricating macromolecules from synovial fluid alters the relationship between mechanical loads and subsequent chondrocyte behavior in injured cartilage. To test this hypothesis, we utilized an ex vivo model of PTOA that involves subjecting cartilage explants to a single rapid impact followed by continuous articulation within a lubricating bath of either healthy synovial fluid, phosphate-buffered saline (PBS), synovial fluid treated with hyaluronidase, or synovial fluid treated with trypsin. These treatments degrade the main macromolecules attributed with providing synovial fluid with its lubricating properties; hyaluronic acid and lubricin. Explants were then bisected and fluorescently stained to assess global and depth-dependent cell death, caspase activity, and mitochondrial depolarization. Explants were tested via confocal elastography to determine the local shear strain profile generated in each lubricant. These results show that degrading hyaluronic acid or lubricin in synovial fluid significantly increases middle zone chondrocyte damage and shear strain loading magnitudes, while also altering chondrocyte sensitivity to loading.

NUMB attenuates posttraumatic osteoarthritis by inhibiting BTRC and inactivating the NF-κB pathway

Posttraumatic osteoarthritis (PTOA) is closely related to the inflammatory response caused by mechanical injury and leads to joint degeneration. Herein, we aimed to evaluate the role and underlying mechanism of NUMB in PTOA progression. Anterior cruciate ligament transection (ACLT)-induced rats and interleukin (IL)-1β-treated chondrocytes were used as in vivo and in vitro models of PTOA, respectively. The NUMB overexpression plasmid (pcDNA-NUMB) was administered by intra-articular injection to PTOA model rats, and safranin O-fast green staining, the Osteoarthritis Research Society International (OARSI) scoring system, and HE staining were used to evaluate the severity of cartilage damage. The secretion of inflammatory cytokines (TNF-α, IL-1β, and IL-6) and chondrocyte-specific markers (MMP13 and COL2A1) was detected via ELISA. Cell viability and apoptosis were evaluated by MTT and TUNEL assays. NUMB was expressed at lower levels in ACLT-induced PTOA rats and in IL-1β-treated chondrocytes than in control rats and cells. NUMB overexpression enhanced cell viability and reduced cell apoptosis, inflammation and cartilage degradation in chondrocytes stimulated by IL-1β. NUMB bound to BTRC to promote p-IκBα expression, resulting in NF-κB pathway inactivation. BTRC overexpression reversed the promoting effect of NUMB overexpression on cell viability and the inhibitory effects of NUMB overexpression on apoptosis, inflammation and cartilage degradation in IL-1β-induced chondrocytes. In addition, overexpression of NUMB alleviated articular cartilage damage by repressing inflammation and cartilage degradation in ACLT-induced PTOA rats. Our data indicated that NUMB regulated PTOA progression through the BTRC/NF-κB pathway, which may be a viable therapeutic target in PTOA.

Mechanism study of BMSC-exosomes combined with hyaluronic acid gel in the treatment of posttraumatic osteoarthritis

ObjectiveTo explore the mechanism and efficacy of gel in the treatment of posttraumatic osteoarthritis (PTOA), combined with hyaluronic acid (HA) and bone marrow mesenchymal stem cell exosomes (BMSC-EXOs), and to explain its role in alleviating oxidative stress damage induced by mitochondrial reactive oxygen species (ROS). MethodsHow is the therapeutic potential of toa influenced by bone marrow mesenchymal stem cells-EXO to be evaluated both in vitro and in vivo. In vitro, BMSC-EXOs were extracted and characterized from rat specimens and labeled with Dil. Rat primary chondrocytes were then isolated to create a cellular PTOA model. BMSC-EXOs + HA group, BMSC-EXOs + HA + 740Y–P group, model group, BMSC-EXOs group, HA group, and control group were included in the cell group, and the function of cartilage matrix and the level of oxidative stress could be evaluated. Cartilage matrix integrity and oxidative stress can be assessed by grouping rats. At the same time, a rat model of ptosis can be established by excision of the anterior cruciate ligament, and joint rehabilitation, with pro-inflammatory and Enzyme-linked immunosorbent assay (ELISA) can be used to determine anti-inflammatory markers. ResultsThe combined use of BMSC-EXOs and HA gel was found to significantly reduce oxidative stress in chondrocytes and PTOA rat models, improving cartilage mechanical properties more effectively than BMSC-EXOs alone. ConclusionBMSC-EXOs combined with HA gel offer a promising treatment for PTOA by modulating damage caused by mitochondrial ROS-induced oxidative stress.

Establishment of a Posttraumatic Osteoarthritis Model in Mice Induced by Noninvasive Anterior Cruciate Ligament Tear

Background: Animal models that use open surgical transection of the anterior cruciate ligament (ACL) do not accurately simulate the clinical condition regarding the pivot-shift mechanism and the associated inflammatory response that occurs before reconstruction. Purpose/Hypothesis: The purpose was to characterize a reproducible manual, nonsurgical method to mimic an isolated ACL tear in a clinically relevant model and to evaluate the development of progressive posttraumatic osteoarthritis due to ACL injury. It was hypothesized that the ACL could be reproducibly torn with minimal damage to other ligaments and that there would be progressive development of degenerative joint disease after ACL injury. Study Design: Controlled laboratory study. Methods: A total of 37 mice (strain C57BL/6) were used to compare the manual procedure with sham surgery (sham group; n = 10) and with the established surgical ACL transection (ACLT) procedure (surgical group; n = 27). In the sham group, a closed manual procedure was performed on the right knee and sham surgery on the left knee. In the surgical group, the closed manual procedure was performed on the right knee and surgical ACLT on the left knee. Dissection using India ink, histological assessment with safranin O and hematoxylin-eosin staining, radiological evaluation through radiographs and microfocus computed tomography scans, and gait analyses were performed to assess cartilage/ligament status. Osteoarthritis Research Society International (OARSI) and synovitis scores, anterior tibial translation, range of motion, bone microstructure, osteophyte volume, and pain were assessed at 2, 4, and 8 weeks postoperatively. Results: The manual procedure successfully resulted in an ACL rupture and associated meniscal injury. The posterior cruciate, lateral collateral, and medial collateral ligaments were intact in all dissected knees. Two weeks after ACL tear, the surgical group showed a significantly higher synovitis score, whereas 8 weeks after ACL tear, the manual group showed a significantly higher volume of osteophytes. No significant differences were found between the groups in terms of OARSI score, anterior tibial translation, range of motion, bone microstructure computed tomography values, and stride distance/irregularity. Conclusion: This procedure can be used to create an ACL tear model without causing grossly evident injuries to other ligaments and avoiding the risk of cartilage damage from surgical instruments. Clinical Relevance: This procedure offers a more clinically relevant ACL tear model and facilitates simple, inexpensive, and reproducible development of posttraumatic osteoarthritis.

Exercise following joint distraction inhibits muscle wasting and delays the progression of post-traumatic osteoarthritis in rabbits by activating PGC-1α in skeletal muscle

ObjectiveMuscle wasting frequently occurs following joint trauma. Previous research has demonstrated that joint distraction in combination with treadmill exercise (TRE) can mitigate intra-articular inflammation and cartilage damage, consequently delaying the advancement of post-traumatic osteoarthritis (PTOA). However, the precise mechanism underlying this phenomenon remains unclear. Hence, the purpose of this study was to examine whether the mechanism by which TRE following joint distraction delays the progression of PTOA involves the activation of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), as well as its impact on muscle wasting.MethodsQuadriceps samples were collected from patients with osteoarthritis (OA) and normal patients with distal femoral fractures, and the expression of PGC-1α was measured. The hinged external fixator was implanted in the rabbit PTOA model. One week after surgery, a PGC-1α agonist or inhibitor was administered for 4 weeks prior to TRE. Western blot analysis was performed to detect the expression of PGC-1α and Muscle atrophy gene 1 (Atrogin-1). We employed the enzyme-linked immunosorbent assay (ELISA) technique to examine pro-inflammatory factors. Additionally, we utilized quantitative real-time polymerase chain reaction (qRT-PCR) to analyze genes associated with cartilage regeneration. Synovial inflammation and cartilage damage were evaluated through hematoxylin-eosin staining. Furthermore, we employed Masson’s trichrome staining and Alcian blue staining to analyze cartilage damage.ResultsThe decreased expression of PGC-1α in skeletal muscle in patients with OA is correlated with the severity of OA. In the rabbit PTOA model, TRE following joint distraction inhibited the expressions of muscle wasting genes, including Atrogin-1 and muscle ring finger 1 (MuRF1), as well as inflammatory factors such as interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in skeletal muscle, potentially through the activation of PGC-1α. Concurrently, the production of IL-1β, IL-6, TNF-α, nitric oxide (NO), and malondialdehyde (MDA) in the synovial fluid was down-regulated, while the expression of type II collagen (Col2a1), Aggrecan (AGN), SRY-box 9 (SOX9) in the cartilage, and superoxide dismutase (SOD) in the synovial fluid was up-regulated. Additionally, histological staining results demonstrated that TRE after joint distraction reduced cartilage degeneration, leading to a significant decrease in OARSI scores.TRE following joint distraction could activate PGC-1α, inhibit Atrogin-1 expression in skeletal muscle, and reduce C-telopeptides of type II collagen (CTX-II) in the blood compared to joint distraction alone.ConclusionFollowing joint distraction, TRE might promote the activation of PGC-1α in skeletal muscle during PTOA progression to exert anti-inflammatory effects in skeletal muscle and joint cavity, thereby inhibiting muscle wasting and promoting cartilage regeneration, making it a potential therapeutic intervention for treating PTOA.

Plasma and synovial fluid extracellular vesicles display altered microRNA profiles in horses with naturally occurring post-traumatic osteoarthritis: an exploratory study.

The objective of this study was to characterize extracellular vesicles (EVs) in plasma and synovial fluid obtained from horses with and without naturally occurring post-traumatic osteoarthritis (PTOA). EVs were isolated from plasma and synovial fluid from horses with (n = 6) and without (n = 6) PTOA. Plasma and synovial fluid EVs were characterized with respect to quantity, size, and surface markers. Small RNA sequencing was performed, and differentially expressed microRNAs (miRNAs) underwent bioinformatic analysis to identify putative targets and to explore potential associations with specific biological processes. Plasma and synovial fluid samples from horses with PTOA had a significantly higher proportion of exosomes and a lower proportion of microvesicles compared to horses without PTOA. Small RNA sequencing revealed several differentially expressed miRNAs, including miR-144, miR-219-3p, and miR-199a-3l in plasma and miR-199a-3p, miR-214, and miR-9094 in synovial fluid EVs. Bioinformatics analysis of the differentially expressed miRNAs highlighted their potential role in fibrosis, differentiation of chondrocytes, apoptosis, and inflammation pathways in PTOA. We have identified dynamic molecular changes in the small noncoding signatures of plasma and synovial fluid EVs in horses with naturally occurring PTOA. These findings could serve to identify promising biomarkers in the pathogenesis of PTOA, to facilitate the development of targeted therapies, and to aid in establishing appropriate translational models of PTOA.

The BMP7-Derived Peptide p[63-82] Reduces Cartilage Degeneration in the Rat ACLT-pMMx Model for Posttraumatic Osteoarthritis.

Osteoarthritis (OA) is characterized by articular cartilage erosion, pathological subchondral bone changes, and signs of synovial inflammation and pain. We previously identified p[63-82], a bone morphogenetic protein 7 (BMP7)-derived bioactive peptide that attenuates structural cartilage degeneration in the rat medial meniscal tear-model for posttraumatic OA. This study aimed to evaluate the cartilage erosion-attenuating activity of p[63-82] in a different preclinical model for OA (anterior cruciate ligament transection-partial medial meniscectomy [anterior cruciate ligament transection (ACLT)-pMMx]). The disease-modifying action of the p[63-82] was followed-up in this model for 5 and 10 weeks. Skeletally mature male Lewis rats underwent ACLT-pMMx surgery. Rats received weekly intra-articular injections with either saline or 500 ng p[63-82]. Five and 10 weeks postsurgery, rats were sacrificed, and subchondral bone characteristics were determined using microcomputed tomography (µCT). Histopathological evaluation of cartilage degradation and Osteoarthritis Research Society International (OARSI)-scoring was performed following Safranin-O/Fast Green staining. Pain-related behavior was measured by incapacitance testing and footprint analysis. Histopathological evaluation at 5 and 10 weeks postsurgery showed reduced cartilage degeneration and a significantly reduced OARSI score, whereas no significant changes in subchondral bone characteristics were found in the p[63-82]-treated rats compared to the saline-treated rats. ACLT-pMMx-induced imbalance of static weightbearing capacity in the p[63-82] group was significantly improved compared to the saline-treated rats at weeks 5 postsurgery. Footprint analysis scores in the p[63-82]-treated rats demonstrated improvement at week 10 postsurgery. Weekly intra-articular injections of p[63-82] in the rat ACLT-pMMx posttraumatic OA model resulted in reduced degenerative cartilage changes and induced functional improvement in static weightbearing capacity during follow-up.

Low-intensity pulsed ultrasound delays the progression of osteoarthritis by regulating the YAP-RIPK1-NF-κB axis and influencing autophagy.

Osteoarthritis (OA) is a degenerative disease characterized by chronic inflammation of the joint. As the disease progresses, patients will gradually develop symptoms such as pain, physical limitations and even disability. The risk factors for OA include genetics, gender, trauma, obesity, and age. Unfortunately, due to limited understanding of its pathological mechanism, there are currently no effective drugs or treatments to suspend the progression of osteoarthritis. In recent years, some studies found that low-intensity pulsed ultrasound (LIPUS) may have a positive effect on osteoarthritis. Nonetheless, the exact mechanism by which LIPUS affects osteoarthritis remains unknown. It is valuable to explore the specific mechanism of LIPUS in the treatment of OA. In this study, we validated the potential therapeutic effect of LIPUS on osteoarthritis by regulating the YAP-RIPK1-NF-κB axis at both cellular and animal levels. To verify the effect of YAP on OA, the expression of YAP was knocked down or overexpressed by siRNA and plasmid in chondrocytes and adeno-associated virus was injected into the knee joint of rats. The effect of LIPUS was investigated in inflammation chondrocytes induced by IL-1β and in the post-traumatic OA model. In this study, we observed that YAP plays an important role in the development of osteoarthritis and knocking down of YAP significantly inhibited the inflammation and alleviated cartilage degeneration. We also demonstrated that the expression of YAP was increased in osteoarthritis chondrocytes and YAP could interact with RIPK1, thereby regulating the NF-κB signal pathway and influencing inflammation. Moreover, we also discovered that LIPUS decreased the expression of YAP by restoring the impaired autophagy capacity and inhibiting the binding between YAP and RIPK1, thereby delaying the progression of osteoarthritis. Animal experiment showed that LIPUS could inhibit cartilage degeneration and alleviate the progression of OA. These results showed that LIPUS is effective in inhibiting inflammation and cartilage degeneration and alleviate the progression of OA. As a result, our results provide new insight of mechanism by which LIPUS delays the development of osteoarthritis, offering a novel therapeutic regimen for osteoarthritis.

Study on the mechanism of cross-linked hyaluronic acid-dexamethasone hydrogelin post-traumatic osteoarthritis

Objective: To explore the mechanism of cross-linked hyaluronic acid-dexamethasone hydrogel (cHA-Dex) in inhibiting chondrocyte apoptosis and alleviating early post-traumatic osteoarthritis (PTOA). Methods: To generate PTOA model, anterior cruciate ligament transection (ACLT)was performed on SD rats (n=70), and the sham surgery group (n=70) was set as control. The changes in inflammatory indicators such as interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), matrix metalloproteinase-3 (MMP-3), and matrix metalloproteinase-13 (MMP-13) in the joint lavage fluid were measured at different time points (1-14 days, 5 rats at each time point) after surgery. The cHA-Dex (0.5 mg/ml) hydrogel (experimental group, n=70) and ordinary low-molecular-weight hyaluronic acid (HA) hydrogel premixed with Dex, that was, HA-Dex (0.5 mg/ml) hydrogel (control group, n=70) were injected into the joint cavity of PTOA rats, and the release amount and cumulative release amount of Dex in the joint fluid of rats at each time point(1-14 days, 5 rats at each time point) were detected to reveal the release mechanism of cHA-Dex hydrogel. The cartilage of knee joint of patients with osteoarthritis (OA) who underwent knee arthroplasty in the Second Hospital of Shanxi Medical University from January 2020 to December 2022 was taken for in vitro tissue block culture (Outbridge score=1 or 2,n=18). After the cartilage tissue block was treated with cHA-Dex hydrogel premixed with 0.1, 0.2, and 0.5 mg/ml Dex, the mRNA expression levels of IL-1β, IL-6, TNF-α, MMP-3, and MMP-13 in the articular cartilage tissue block were detected. OA chondrocytes were isolated from cartilage samples using enzymatic hydrolysis and cultured in vitro (n=18). Chondrocytes were divided into 4 groups: saline, cHA hydrogel, Dex (0.5 mg/ml), and cHA-Dex (0.5 mg/ml) hydrogel group. The effects of different interventions on chondrocyte proliferation and apoptosis were tested. Results: The Osteoarthritis Research Society International (OARSI) score of safranine O-solid green staining in PTOA group was 3.34±0.35, and it was 1.17±0.21 in Sham group(P=0.010). The Meachim score of knee joint osteophytes in PTOA rats was significantly higher than that in the Sham group (2.66±0.41 vs 0.22±0.17, P=0.010), indicating PTOA model in rat was established successfully. The cHA-Dex hydrogel, which corresponded to the peak changes of inflammatory factors in the joints of PTOA rats in the early stage, was also released in the early stage and sustained-released in the late stage. After the OA articular cartilage tissue block was treated with cHA-Dex hydrogel premixed with 0.1, 0.2, and 0.5 mg/ml Dex, the mRNA expression levels of IL-1 β, IL-6, TNF-α, MMP-3, and MMP-13 in the tissue block were reduced significantly (all P<0.05) and in a dose-dependent manner. Compared with Dex (0.5 mg/ml) alone group, the apoptosis rate of cHA-Dex (0.5 mg/ml) hydrogel group was significantly reduced (0.60±0.07 vs 6.63±0.98, P=0.010).Compared with the normal saline or the cHA hydrogel alone group, the cHA-Dex (0.5 mg/ml) hydrogel group had significant cell proliferation, and the difference at each time point were all significant statistically (all P<0.05). Conclusion: For the early inflammation of PTOA, cHA-Dex hydrogel can not only inhibit cartilage inflammation, but also reverse the increased apoptosis and decreased proliferation rate of chondrocytes caused by Dex, and finally alleviate the progress of PTOA by releasing Dex.

Inhibition of Phlpp1 preserves the mechanical integrity of articular cartilage in a murine model of post-traumatic osteoarthritis

ObjectivePhlpp1 inhibition is a potential therapeutic strategy for cartilage regeneration and prevention of post-traumatic osteoarthritis (PTOA). To understand how Phlpp1 loss affects cartilage structure, cartilage elastic modulus was measured with atomic force microscopy (AFM) in male and female mice after injury. MethodsOsteoarthritis was induced in male and female Wildtype (WT) and Phlpp1-/- mice by destabilization of the medial meniscus (DMM). At various timepoints post-injury, activity was measured, and knee joints examined with AFM and histology. In another cohort of WT mice, the PHLPP inhibitor NSC117079 was intraarticularly injected four weeks after injury. ResultsMale WT mice showed decreased activity and histological signs of cartilage damage at 12 but not 6 weeks post-DMM. Female mice showed a less severe response to DMM by comparison, with no histological changes seen at any timepoint. In both sexes the elastic modulus of medial condylar cartilage was decreased in WT mice but not Phlpp1-/- mice after DMM as measured by AFM. By 6 weeks, cartilage modulus had decreased from 2MPa to 1MPa in WT mice. Phlpp1-/- mice showed no change in modulus at 6 weeks and only a 25% decrease at 12 weeks. The PHLPP inhibitor NSC117079 protected cartilage structure and prevented signs of OA 6 weeks post-injury. ConclusionsAFM is a sensitive method for detecting early changes in articular cartilage post-injury. Phlpp1 suppression, either through genetic deletion or pharmacological inhibition, protects cartilage degradation in a model of PTOA, validating Phlpp1 as a therapeutic target for PTOA.

Sex-specific effects of injury and beta-adrenergic activation on metabolic and inflammatory mediators in a murine model of post-traumatic osteoarthritis

ObjectiveMetabolic processes are intricately linked to the resolution of innate inflammation and tissue repair, two critical steps for treating post-traumatic osteoarthritis (PTOA). Based on lipolytic and immunoregulatory actions of norepinephrine, we hypothesized that intra-articular β-adrenergic receptor (βAR) stimulation would suppress PTOA-associated inflammation in the infrapatellar fat pad (IFP) and synovium. DesignWe used the βAR agonist isoproterenol to perturb intra-articular metabolism 3.5 weeks after applying a non-invasive single-load compression injury to knees of 12-week-old male and female mice. We examined the acute effects of intra-articular isoproterenol treatment relative to saline on IFP histology, multiplex gene expression of synovium-IFP tissue, synovial fluid metabolomics, and mechanical allodynia. ResultsInjured knees developed PTOA pathology characterized by heterotopic ossification, articular cartilage loss, and IFP atrophy and fibrosis. Isoproterenol suppressed the upregulation of pro-fibrotic genes and downregulated the expression of adipose genes and pro-inflammatory genes (Adam17, Cd14, Icam1, Csf1r, and Casp1) in injured joints of female (but not male) mice. Analysis of published single-cell RNA-seq data identified elevated catecholamine-associated gene expression in resident-like synovial-IFP macrophages after injury. Injury substantially altered synovial fluid metabolites by increasing amino acids, peptides, sphingolipids, phospholipids, bile acids, and dicarboxylic acids, but these changes were not appreciably altered by isoproterenol. Intra-articular injection of either isoproterenol or saline increased mechanical allodynia in female mice, whereas neither substance affected male mice. ConclusionsAcute βAR activation altered synovial-IFP transcription in a sex and injury-dependent manner, suggesting that women with PTOA may be more sensitive than men to treatments targeting sympathetic neural signaling pathways.

Transcript-dependent effects of the CALCA gene on the progression of post-traumatic osteoarthritis in mice

Osteoarthritis represents a chronic degenerative joint disease with exceptional clinical relevance. Polymorphisms of the CALCA gene, giving rise to either a procalcitonin/calcitonin (PCT/CT) or a calcitonin gene-related peptide alpha (αCGRP) transcript by alternative splicing, were reported to be associated with the development of osteoarthritis. The objective of this study was to investigate the role of both PCT/CT and αCGRP transcripts in a mouse model of post-traumatic osteoarthritis (ptOA). WT, αCGRP−/− and CALCA−/− mice were subjected to anterior cruciate ligament transection (ACLT) to induce ptOA of the knee. Mice were sacrificed 4 and 8 weeks post-surgery, followed by micro-CT and histological evaluation. Here we show that the expression of both PCT/CT and αCGRP transcripts is induced in ptOA knees. CALCA−/− mice show increased cartilage degeneration and subchondral bone loss with elevated osteoclast numbers compared to αCGRP−/− and WT mice. Osteophyte formation is reduced to the same extent in CALCA−/− and αCGRP−/− mice compared to WT controls, while a reduced synovitis score is noticed exclusively in mice lacking CALCA. Our data show that expression of the PCT/CT transcript protects from the progression of ptOA, while αCGRP promotes osteophyte formation, suggesting that CALCA-encoded peptides may represent novel targets for the treatment of ptOA.

Collagen type II solution extracted from supercritical carbon dioxide decellularized porcine cartilage: regenerative efficacy on post-traumatic osteoarthritis model

Osteoarthritis (OA) of the knee is a common degenerative articular disorder and is one of the main causes of pain and functional disability. Cartilage damage is frequently linked to elevated osteoarthritis incidence. Supercritical carbon dioxide (scCO2) decellularized cartilage graft produced from the porcine cartilage is an ideal candidate for cartilage tissue engineering. In the present study, we derived collagen type II (Col II) solution from the scCO2 decellularized porcine cartilage graft (dPCG) and compared its efficacy with hyaluronic acid (HA) in the surgical medial meniscectomy (MNX) induced post-traumatic osteoarthritis (PTOA) model. Dose-dependent attenuation of the OA (12.3 ± 0.8) progression was observed in the intra‐articular administration of Col II solution (7.3 ± 1.2) which significantly decreased the MNX-induced OA symptoms similar to HA. The pain of the OA group (37.4 ± 2.7) was attenuated dose-dependently by Col II solution (45.9 ± 4.1) similar to HA (43.1 ± 3.5) as evaluated by a capacitance meter. Micro‐CT depicted a dose-dependent attenuation of articular cartilage damage by the Col II solution similar to HA treatment. A significant (p < 0.001) dose-dependent elevation in the bone volume was also observed in Col II solution-treated OA animals. The protective competence of Col II solution on articular cartilage damage is due to its significant (p < 0.001) increase in the expression of type II collagen, aggrecan and SOX‐9 similar to HA. To conclude, intra‐articular administration of type II collagen solution and HA reestablished the injured cartilage and decreased osteoarthritis progression in the experimental PTOA model.Graphical

Exercise-induced modulation of myokine irisin on muscle-bone unit in the rat model of post-traumatic osteoarthritis

Background and aimPost-traumatic osteoarthritis (PTOA) is a subtype of osteoarthritis (OA). Exercise may produce and release the myokine irisin through muscle fiber contraction. However, the effect of exercise-promoted irisin production on the internal interactions of the muscle–bone unit in PTOA studies remains unclear.MethodsEighteen 8-week-old Sprague–Dawley (SD) rats were randomly divided into three groups: Sham/sedentary (Sham/Sed), PTOA/sedentary (PTOA/Sed), and PTOA/treadmill-walking (PTOA/TW). The PTOA model was established by transection of anterior cruciate ligament (ACLT) and destabilization of medial meniscus (DMM). After 4 weeks of modeling, the PTOA/TW group underwent treadmill exercise (15 m/min, 30 min/d, 5 d/ week, 8 weeks), and the other two groups were free to move in the cage. Evaluation and correlation analysis of muscle, cartilage, subchondral bone and serological indexes were performed after euthanasia.ResultsEight weeks of treadmill exercise effectively alleviated the trauma-induced OA phenotype, thereby maintaining cartilage and subchondral bone integrity in PTOA, and reducing quadriceps atrophy and myofibril degradation. Exercise reversed the down-regulated expression of peroxisome proliferator-activated receptor-gamma coactivator-1α (PGC-1α) and fibronectin type III structural domain protein 5 (FNDC5) in muscle tissue of PTOA rats, and increased the blood irisin level, and the irisin level was positively correlated with the expression of PGC-1α and FNDC5. In addition, correlation analysis showed that irisin metabolism level was strongly negatively correlated with Osteoarthritis Research Society International (OARSI) and subchondral bone loss, indicating that irisin may be involved in cartilage biology and PTOA-related changes in cartilage and subchondral bone. Moreover, the metabolic level of irisin was strongly negatively correlated with muscle fiber cross-sectional area (CSA), Atrogin-1 and muscle ring-finger protein-1(MuRF-1) expression, suggesting that irisin may alleviate muscle atrophy through autocrine action.ConclusionTreadmill exercise can alleviate the atrophy and degeneration of muscle fibers in PTOA rats, reduce the degradation of muscle fibrin, promote the expression of serum irisin, and alleviate the degeneration of articular cartilage and subchondral bone loss in PTOA rats. These results indicate that treadmill exercise can affect the process of PTOA by promoting the expression of myokine irisin in rat muscle–bone unit.

Diverse Response to Local Pharmacological Blockade of Sirt1 Cleavage in Age-Induced versus Trauma-Induced Osteoarthritis Female Mice.

Objective: Previous studies have shown that the cleavage of Sirt1 contributes to the development of osteoarthritis (OA). In fact, OA was effectively abrogated by the intra-articular (IA) administration of two compounds, one blocking Sirt1 cleavage (CA074me) and the other activating Sirt1 (SRT1720), using a post-traumatically induced model (PTOA) in young female mice. In this study, we attempted to understand if this local treatment is effective in preventing age-associated OA (AOA) progression and symptoms. Design: A group of 17-month-old female C57BL/6J mice were IA administered with CA074me and/or SRT1720 or their combination. Joint histopathological analysis and bone histomorphometry were carried out, with an assessment of knee mechanical hyperalgesia. A serum analysis for NT/CT Sirt1 was carried out along with immunohistochemistry for articular cartilage to detect p16INK4A or γH2A.X. Similarly, meniscal cartilage was monitored for Lef1 and Col1a1 deposition. The data were compared for young female mice subjected to post-traumatic OA (PTOA). Results: Similar to PTOA, combination-treated AOA exhibited improved knee hyperalgesia, yet structural improvements were undetected, corresponding to unchanged NT/CT Sirt1 serum levels. Both AOA and PTOA exhibited unchanged staining for nuclear p16INK4A or γH2A.X and lacked a correlation with OA severity. Contrarily to PTOA, the combination treatment with AOA did not exhibit a local reduction in the Lef1 and Col1 targets. Conclusions: When targeting Sirt1 cleavage, the PTOA and AOA models exhibited a similar pain response to the combination treatment; however, they displayed diverse structural outcomes for joint-related damage, related to Lef1-dependent signaling. Interestingly, nuclear p16INK4A was unaffected in both models, regardless of the treatment's effectiveness. Finally, these findings highlight the variations in the responses between two highly researched OA preclinical models, reflecting OA pathophysiology heterogeneity and variations in gender-related drug-response mechanisms.

Wnt5a deficiency in osteocalcin-expressing cells could not alleviate the osteoarthritic phenotype in a mouse model of post-traumatic osteoarthritis.

Wnt5a, which regulates the activities of osteoblasts and osteoclasts, is reportedly overexpressed in osteoarthritis (OA) tissues. The purpose of this study was to elucidate its role in the development of OA by deleting Wnt5a in osteocalcin (OCN)-expressing cells. Knee OA was induced by anterior cruciate ligament transection (ACLT) in OCN-Cre;Wnt5afl/fl knockout (Wnt5a-cKO) mice and control littermates. Eight weeks after surgery, histological changes, cell apoptosis, and matrix metabolism of cartilage were evaluated by toluidine blue, TUNEL staining, and im-immunohistochemistry analyses, respectively. In addition, the subchondral bone microarchitecture of mice was examined by micro-computed tomography (micro-CT). Histological scores show substantial cartilage degeneration occurred in ACLT knees, coupled with decreased collagen type II expression and enhanced matrix metalloproteinase 13 expression, as well as higher proportions of apoptotic cells. Micro-CT results show that ACLT resulted in decreased bone mineral density, bone volume/trabecular volume, trabecular number, and structure model index of subchondral bones in both Wnt5a-cKO and control littermates; although Wnt5a-cKO mice display lower BMD and BV/TV values, no significant difference was observed between Wnt5a-cKO and control mice for any of these values. Our findings indicate that Wnt5a deficiency in OCN-expressing cells could not prevent an osteoarthritic phenotype in a mouse model of post-traumatic OA.

GDF8 inhibition enhances musculoskeletal recovery and mitigates posttraumatic osteoarthritis following joint injury.

Musculoskeletal disorders contribute substantially to worldwide disability. Anterior cruciate ligament (ACL) tears result in unresolved muscle weakness and posttraumatic osteoarthritis (PTOA). Growth differentiation factor 8 (GDF8) has been implicated in the pathogenesis of musculoskeletal degeneration following ACL injury. We investigated GDF8 levels in ACL-injured human skeletal muscle and serum and tested a humanized monoclonal GDF8 antibody against a placebo in a mouse model of PTOA (surgically induced ACL tear). In patients, muscle GDF8 was predictive of atrophy, weakness, and periarticular bone loss 6 months following surgical ACL reconstruction. In mice, GDF8 antibody administration substantially mitigated muscle atrophy, weakness, and fibrosis. GDF8 antibody treatment rescued the skeletal muscle and articular cartilage transcriptomic response to ACL injury and attenuated PTOA severity and deficits in periarticular bone microarchitecture. Furthermore, GDF8 genetic deletion neutralized musculoskeletal deficits in response to ACL injury. Our findings support an opportunity for rapid targeting of GDF8 to enhance functional musculoskeletal recovery and mitigate the severity of PTOA after injury.

A Reproducible Cartilage Impact Model to Generate Post-Traumatic Osteoarthritis in the Rabbit.

Post-traumatic osteoarthritis (PTOA) is responsible for 12% of all osteoarthritis cases in the United States. PTOA can be initiated by a single traumatic event, such as a high-impact load acting on articular cartilage, or by joint instability, as occurs with anterior cruciate ligament rupture. There are no effective therapeutics to prevent PTOA currently. Developing a reliable animal model of PTOA is necessary to better understand the mechanisms by which cartilage damage proceeds and to investigate novel treatment strategies to alleviate or prevent the progression of PTOA. This protocol describes an open, drop tower-based rabbit femoral condyle impact model to induce cartilage damage. This model delivered peak loads of 579.1 ± 71.1 N, and peak stresses of 81.9 ± 10.1 MPa with a time-to-peak load of 2.4 ± 0.5 ms. Articular cartilage from impacted medial femoral condyles (MFCs) had higher rates of apoptotic cells (p = 0.0058) and possessed higher Osteoarthritis Research Society International (OARSI) scores of 3.38 ± 1.43 compared to the non-impacted contralateral MFCs (0.56 ± 0.42), and other cartilage surfaces of the impacted knee (p < 0.0001). No differences in OARSI scores were detected among the non-impacted articular surfaces (p > 0.05).