Abstract Post-traumatic osteoarthritis (PTOA) is a common long-term consequence of joint injury and a major cause of chronic pain and disability, yet no disease-modifying therapies are currently available. A central barrier to effective intervention is the persistence of maladaptive synovial inflammation, driven in part by macrophage-mediated signaling that sustains tissue degeneration and pain. Here, we developed a scalable, chemically defined platform to generate human induced pluripotent stem cell (iPSC)–derived anti-inflammatory macrophages (iMac-M2) as an off-the-shelf cell therapy designed to restore joint immune homeostasis after injury. These cells maintained a stable anti-inflammatory phenotype and function under osteoarthritis-relevant inflammatory conditions and suppressed inflammatory and catabolic responses in human joint cell co-culture systems. In a preclinical model of PTOA, intra-articular delivery of iMac-M2 after injury improved functional and structural outcomes while modulating synovial inflammatory and pain-associated transcriptional programs. Treatment was well tolerated, with no evidence of systemic immune activation or ectopic tissue formation. Together, these findings support iPSC-derived macrophage therapy as a clinically translatable immunomodulatory strategy to interrupt early inflammatory drivers of PTOA and preserve joint health following injury.

Abstract In the US, 33 million musculoskeletal injuries have been reported per year, with 50% involving tendons and ligaments in both athletic and aging populations. Tendon repair often results in the formation of biomechanically inferior scar tissue rather than functional regeneration. Local cell therapy is garnering significant interest in tendon repair because it provides a targeted means to repopulate defects with potent therapeutic cells. Here, we developed a porcine tendon-derived thermoresponsive extracellular matrix hydrogel (TG) as an injectable, bio-instructive carrier for scleraxis-overexpressing induced mesenchymal stem cell–derived tenocytes (iTenocytes), with the goal of improving cell retention and overall transplantation success. TG was compositionally distinct from purified collagen (PC, from rat tail type 1 collagen) and exhibited favorable material properties for a minimally invasive percutaneous strategy, including thermoresponsive gelation, shear-thinning injectability, retention at the injection site, and controlled biodegradation. In vitro , TG supported three-dimensional cell residence, promoted cell interconnectivity and redistribution at the matrix interface, and increased collagen type I release from iTenocytes compared to those embedded in PC. Transcriptomic and proteomic analyses further showed that TG enhanced programs associated with tenogenic maturation, extracellular matrix assembly, focal adhesion, mechano-transduction, and remodeling. In a rat Achilles tendon partial defect model, both optical imaging and histological analyses demonstrated retention of iTenocytes at the injection site. Additionally, confocal imaging demonstrated retention of iTenocytes within the defect site for up to 10 days. Together, these findings identify TG as a biofunctional injectable carrier that supports the tenogenic characteristics of iTenocytes and enhances their local persistence after transplantation.

Summary Tendon injuries often result in fibrosis, compromising function and predisposing to re-injury. Here, we used a full-width, non-repair Achilles tendon transection model in young (3 weeks old) and adult (18-20 weeks) rats to elucidate the cellular mechanisms governing regenerative versus fibrotic healing. Functional and biomechanical analyses revealed that young tendons recovered motion and load-bearing capacity more rapidly but exhibited more fibrotic early healing. Single-nuclei RNA sequencing identified seven major cell populations within the connective tissue compartment. Adult tendons maintained a “synthetic fibroblast” population marked by upregulated ECM synthesis, reduced stress-related gene expression, whereas young tendons favored expansion of Cxcl12⁺/Lrp6⁺/Gas6⁺ fibrotic fibroblasts linked to oxidative and pro-angiogenic signaling. The young group showed sustained activation of Nox4–Gas6 pathways driving a self-reinforcing fibrotic circuit. These findings define a fibroblast lineage bifurcation that dictates oxidative stress signaling as a key regulator of fibrotic remodeling, highlighting potential therapeutic targets to promote regenerative tendon repair.

During vertebrate embryogenesis, axial tendons develop from the paraxial mesoderm and differentiate through specific developmental stages to reach the syndetome stage. While the main roles of signaling pathways in the earlier stages of the differentiation have been well established, pathway nuances in syndetome specification from the sclerotome stage have yet to be explored. Here, we show stepwise differentiation of human iPSCs to the syndetome stage using chemically defined media and small molecules that were modified based on single cell RNA-sequencing and pathway analysis. We identified a significant population of branching off-target cells differentiating towards a neural phenotype overexpressing Wnt. Further transcriptomics post-addition of a WNT inhibitor at the somite stage and onwards revealed not only total removal of the neural off-target cells, but also increased syndetome induction efficiency. Fine-tuning tendon differentiation in vitro is essential to address the current challenges in developing a successful cell-based tendon therapy.

Chronic lower back pain (LBP), a leading cause of disability affecting over half a billion people globally, is often linked to intervertebral disc (IVD) degeneration. Current treatments, like surgery and pain management, fail to address the IVD degeneration that is associated with the deterioration of the nucleus pulposus (NP). We aim to develop an injectable cell therapy for LBP using human nucleus pulposus cells (NPCs) encapsulated in hydrogel microparticles (microgels) as a cell delivery vehicle. The thermoresponsive hydrogels enabled cell encapsulation and preconditioning without the use of ultraviolet irradiation or cytotoxic cross-linkers. The in vitro preconditioning maintained cell viability and promoted type 2 collagen secretion. In a rat IVD degeneration model, NPCs were encapsulated in fibrinogen-based hydrogels at 10 million cells/mL, preconditioned for 7 days, and injected into degenerated discs. Evaluations of disc height using μCT, gene expression using RT-qPCR on the harvested IVD and dorsal root ganglion (DRG), and histology showed that NPC-loaded microgels increased IVD height and reduced pain-related gene expressions compared to controls. Our findings demonstrate that NPC-loaded microgels have the potential to alleviate pain and regenerate IVDs, suggesting minimally invasive treatment for discogenic LBP and future clinical applications.

Background: Current treatments for osteoarthritis (OA) include physical therapy, nonsteroidal anti-inflammatory drugs, and joint arthroplasty, which aim to address symptoms rather than modify disease progression. Autologous platelet-rich plasma (PRP) and other biologics administered in the affected joint are widely used as alternatives that are believed to alleviate symptoms. However, PRP's effectiveness in reducing knee OA pain is inconsistent across the literature, and the specific therapeutic components remain unclear. Purpose: To identify whether specific cellular and protein biomarkers correlate with patient-reported outcomes and can therefore serve as predictive biomarkers for PRP effectiveness. Study Design: Case series; Level of evidence, 4. Methods: PRP samples from 30 patients (30 knees) with knee OA (Kellgren & Lawrence grades 2-3) who were undergoing PRP treatment were analyzed to identify biomarkers associated with response. Blood samples were drawn, and 2 identical PRP samples were prepared, with one set used for therapy and the other for analysis. Component analysis included cell counts, immune cell, and platelet characterization via mass cytometry, along with proteomic analysis using mass spectrometry. Clinical outcomes were assessed using the Knee injury and Osteoarthritis Outcome Score (KOOS) and Patient-Reported Outcomes Measurement Information System patient reported outcomes at baseline and up to 6 months postinjection. Results: PRP therapy improved pain in 19 of 30 patients at 6 weeks postinjection (mean ± SD age, 56.04 ± 10.56 years): Several distinct immune cell clusters (T helper 1 CD4 T cells ( r = 0.44; P < .05), Central memory CD4 T cells ( r = 0.46; P < .05), Effector memory CD4 T cells ( r = 0.52; P < .05), and CD8 T cells ( r = 0.44; P < .05) positively correlated with ΔKOOS ( P < .05). Proteomic analysis showed that differentially expressed proteins integrin-linked kinase ( r = −0.50; P < .05), and glutathione peroxidase 1 ( r = −0.55; P < .01) negatively correlated with ΔKOOS Pain ( P < .05), whereas cathepsin G ( r = 0.51; P < .05), and calprotectin subunit ( r = 0.65; P < .01) were positively correlated with ΔKOOS Pain ( P < .05) (* P < .05, ** P < 0.01). Conclusion: These findings suggest that some cellular and protein biomarkers may potentially be used as therapeutic targets and predictive markers for PRP success, guiding personalized therapeutic strategies. However, these targets must be further validated for knee OA in future studies.

BACKGROUND:Female reproductive hormones are attributed to the higher rate of musculoskeletal (MSK) injuries among female athletes compared with male athletes. PURPOSE:To evaluate association between exposure to hormonal contraceptives (HCs), joint laxity, and number of injuries in female athletes over a 1-year period. STUDY DESIGN:Cohort study. LEVEL OF EVIDENCE:Level 3. METHODS:Professional and Division I collegiate female athletes were recruited into 2 groups based on HC status (HC vs non-HC). Serum relaxin, estrogen and progesterone levels, knee laxity, generalized hypermobility, and lower extremity kinematics during a single-legged jump were examined during preseason in the luteal phase of each athlete's menstrual cycle. Injuries were tracked for 1 year after testing. RESULTS:Circulating levels of progesterone (non-HC, 38.9 ± 15.0 pg/mg vs HC, 28.6 ± 11.4 pg/mg; P < 0.01), estrogen (non-HC, 2.80 ± 3.0 pg/mg vs HC, 2.0 ± 3.0 pg/mg; P = 0.01), and relaxin (non-HC, 0.26 ± 0.08 pg/mg vs HC, 0.22 ± 0.03 pg/mg; P = 0.04) were lower in HC (n = 32) than in non-HC (n = 40) athletes during the luteal phase of the cycle. Non-HC athletes demonstrated significantly greater hip flexion at initial contact (non-HC, 29.42°± 7.64° vs HC, 25.25°± 7.13°; P = 0.02), and greater knee adduction (valgus) at maximum knee flexion (non-HC, -1.90°± 3.25° vs HC, -0.25° ± 3.40°; P = 0.02) during a single-legged drop. Average (±SD) injury count for non-HC athletes was 0.71 (±1.3) compared with 0.35 (±0.70) in the HC group (P = 0.25). Injury count was correlated significantly to circulating levels of relaxin (r = 0.32; P < 0.01). CONCLUSION:A potential relationship between increased circulating relaxin levels and the risk of injury was established. Athletes taking HCs demonstrated significantly reduced peak knee adduction angles when landing from a jump. CLINICAL RELEVANCE:The results advance knowledge of relationships between specific hormones and injury risk, and the potential role of HCs on kinematic patterns when landing from a jump. Future research will determine HC formulations that may protect against MSK injury, and inform broader strategies to reduce sport-related injury risk in female athletes.

Lower back pain (LBP) is one of the most common causes of disability, with up to 40% of LBP cases attributed to intervertebral disc (IVD) degeneration. While small animal models are widely used to study IVD and LBP, the small size of their IVDs limits translational and biological relevance. Large animal models more accurately emulate human disease; however, methods of measuring LBP are not well established. Pigs were also considered unfit for LBP research, due to notochordal cells (NCs) persistence through life, unlike humans. We developed a comprehensive porcine model with quantitative measure of discogenic pain via biobehavioral testing (BBT), MRI, and multi-omics tissue analysis of IVD and DRGs. MRI demonstrated the progression of IVD degeneration beginning at 4 weeks post-injury. BBTs showed the development of significant pain responses by week 4 post-injury, supported by transcriptomics of injury matched DRGs. Single cell transcriptomics, trajectory and cell-cell communication analyses suggest that, with injury, NCs are differentiating to nucleus pulposus cells (NPCs). Furthermore, NPCs showed upregulation of cellular stress, neural outgrowth, and inflammation pathway, consistent with pain-inducing distress signals found in human samples. This study establishes novel MRI and BBT-based methods for quantifying LBP in pigs and supports its translational relevance to human discogenic LBP. The identification of LBP-associated clusters mirrors our previous finding in humans. Moreover, the shift of NC to NPC phenotype further supports that the porcine model is relevant to human pathology, as the injury induced accelerated aging and loss of NCs with IVD degeneration and discogenic pain.

Background:Low back pain remains the leading cause of disability worldwide, with intervertebral disc degeneration representing a major biological contributor. Although cell-based therapies have shown promise in preclinical models, clinical translation has yielded modest and inconsistent outcomes. Accumulating evidence suggests that therapeutic failure reflects not only limitations in cell source or differentiation potential, but also the hostile biochemical and biomechanical microenvironment of the degenerative disc. Hypoxia, nutrient deprivation, acidity, lactate accumulation, fibrosis, senescence, inflammation, and abnormal mechanical loading collectively impair cell survival, integration, and long-term function. Method:We performed a comprehensive review of the literature using PubMed, Web of Science, and Google Scholar, with emphasis on studies published between 2020 and 2026. Evidence was critically evaluated to examine advances in cell-based therapies for IVDD, including cell sources, mechanisms of repair, biomaterial-assisted delivery systems, microenvironment-targeted strategies, translational studies, and emerging technologies that enhance regenerative efficacy. Discussion:Current evidence indicates that successful disc regeneration depends not only on selecting an appropriate therapeutic cell source but also on overcoming the biological constraints imposed by the degenerative niche. We critically compare the regenerative potential of mesenchymal stromal cells, nucleus pulposus cells, and induced pluripotent stem cell-derived therapies, highlighting their respective advantages and limitations. We further discuss how biomaterial carriers, extracellular vesicles, developmental biology-guided differentiation, genetic engineering, preconditioning approaches, and smart delivery platforms are being integrated to improve cell survival, phenotype stability, extracellular matrix restoration, and functional repair. Conclusion:Future success in intervertebral disc regeneration will require integrated therapeutic strategies that combine optimized cell sources with biomaterial-assisted delivery, microenvironment modulation, and precision bioengineering. Advancing these complementary approaches will be essential for achieving durable biological repair, restoring disc structure and function, and translating regenerative therapies into effective clinical treatments for patients with degenerative disc disease.

To assess lower back pain using quantitative chemical exchange saturation transfer (qCEST) imaging in a porcine model by comparing exchange rate maps obtained from multitasking qCEST with conventional qCEST. Use a permuted random forest (PRF) model trained on CEST-derived magnetization transfer ratio (MTR) and exchange rate (ksw) features to predict Glasgow pain scores. Six Yucatan minipigs were scanned at baseline and at four post-injury time points (weeks 4, 8, 12, and 16) following intervertebral disc injury. Conventional qCEST imaging was performed at four B1 powers using a two-dimensional reduced field of view turbo spin-echo (TSE) sequence, with a total acquisition time of 24 min per slice. Multitasking steady-state (SS) CEST imaging was performed with pulsed saturation to achieve a steady state, acquiring 32 slices at 59 offsets for 4 B1 powers in 36 min. Exchange rate maps were generated using omega plot analysis, and CEST images were analyzed using a multi-pool fitting model to produce MTR and ksw maps. Permuted random forest (PRF) model was trained on MTR and ksw values to predict pain scores. Modic changes were assessed using T2-weighted MR images. The Pearson correlation coefficient between exchange rate maps from multitasking qCEST and conventional qCEST was 0.82, demonstrating strong agreement. The 3D qCEST (SS-CEST) technique effectively differentiated between healthy and injured discs, with injured discs exhibiting significantly higher ksw values. Using MTR and ksw, the PRF model achieved 80% accuracy in predicting pain scores disc-by-disc, outperforming the correlation with Modic changes (r = 0.45, p < 0.05); with a Cohen's Kappa of 0.4. 3D steady-state qCEST with whole-spine coverage can be done at 3T within 32 min using MR Multitasking (acceleration factor of 22), and qCEST-derived biomarkers (MTR and ksw) can predict pain scores with an accuracy of 80%.

Chronic lower back pain (LBP) is the leading cause of disability worldwide. Due to its close relationship with intervertebral disc (IVD) degeneration (IVDD), research has historically focused more on understanding the mechanism behind IVDD while clinical efforts prioritize pain management. More recently, there has been a shift toward understanding LBP as a distinct pathological entity. This review synthesizes current knowledge on discogenic LBP, combining known pathophysiology, molecular mechanisms, risk factors, diagnostic challenges, and available experimental models. IVDD is a complex, multifactorial process involving biochemical, mechanical, and inflammatory changes within the disc, leading to structural breakdown and potential discogenic pain. Key mechanisms include extracellular matrix degradation, upregulation of inflammatory mediators, immune cell infiltration, and aberrant nerve and vascular ingrowth. However, not all cases of IVDD result in LBP, highlighting the need for further investigation into the cellular, molecular, and biomechanical factors contributing to symptom development. Current diagnostic tools and experimental models for studying discogenic LBP remain limited, impeding the development of targeted treatments. Existing therapies primarily focus on symptom management rather than addressing underlying disease mechanisms.

Introduction:This study investigates the epigenetic landscape underlying painful intervertebral disk (IVD) degeneration in a single subject with a history of low back pain (LBP). Intervertebral disk degeneration is associated with LBP in some individuals; however, there is often a discrepancy between degeneration and pain. We hypothesize that DNA methylation, an epigenetic mechanism previously linked to discogenic LBP, is dysregulated in symptomatic vs asymptomatic IVDs. Objectives:Identify differentially methylated genes and pathways in symptomatic vs asymptomatic IVDs. Methods:Three lumbar IVDs with similar degeneration severity were tested prior to surgery by discography to identify symptomatic IVDs. Methylation analysis was performed on ∼935,000 cytosine guanine dinucleotide sites on nucleus pulposus DNA. We explored differential methylation and pathway enrichment on cytosine guanine dinucleotide sites located within the promoter regions of genes. Results:Two IVDs (L3/L4 and L4/L5) evoked pain ratings of 10/10 and 8/10, one IVD (L5/S1) scored 0/10. DNA methylation differed between symptomatic and asymptomatic IVDs. Several identified genes have roles in extracellular matrix remodeling. Other differentially methylated genes were related to immunomodulation and ion channel function. Finally, several long noncoding RNA genes were identified, encouraging further exploration into these regulatory molecules. Enriched pathways were associated with immune response, hormonal regulation, nervous system development, and musculoskeletal development and remodeling. Conclusion:This case study provides a promising list of candidate genes for therapeutic development for discogenic LBP and suggests a role for DNA methylation in the development of symptomatic vs asymptomatic IVD degeneration, calling for further research to validate and expand these findings.

Intervertebral disc (IVD) degeneration is a leading cause of lower back pain (LBP). Current treatments primarily address symptoms without halting the degenerative process. Cell transplantation offers a promising approach for early-stage IVD degeneration, but challenges such as cell viability, retention, and harsh host environments limit its efficacy. This study aimed to compare the injectability and biocompatibility of human nucleus pulposus cells (hNPC) attached to two types of microscaffolds designed for minimally invasive delivery to IVD. Microscaffolds are developed from poly(lactic-co-glycolic acid) (PLGA) using electrospinning and femtosecond laser structuration. These microscaffolds are tested for their physical properties, injectability, and biocompatibility. This study evaluates cell adhesion, proliferation, and survival in vitro and ex vivo within a hydrogel-based nucleus pulposus model. The microscaffolds demonstrate enhanced surface architecture, facilitating cell adhesion and proliferation. Laser structuration improved porosity, supporting cell attachment and extracellular matrix deposition. Injectability tests show that microscaffolds can be delivered through small-gauge needles with minimal force, maintaining high cell viability. The findings suggest that laser-structured PLGA microscaffolds are viable for minimally invasive cell delivery. These microscaffolds enhance cell viability and retention, offering potential improvements in the therapeutic efficiency of cell-based treatments for discogenic LBP. Laser-induced microporosity and structuration of electrospun mats enables the efficient production of cell-carriers. Nucleus pulposus cells attach and proliferate forming cell-populated agglomerations. Injectability studies with hyaluronic acid show high injectability rate through a 23G and 26G needles, with improved cell viability compared to cell suspension. The required ejection force remain below 10 N, suitable for manual injection. image

IntroductionTendon injuries represent an ongoing challenge in clinical practice due to poor regenerative capacity, structure, and biomechanical function recovery of ruptured tendons. This study is focused on the assessment of a novel strategy to repair ruptured Achilles tendons in a Nude rat model using stem cell-seeded biomaterial.MethodsSpecifically, we have used induced pluripotent stem cell (iPSC)-derived mesenchymal stem cells (iMSCs) overexpressing the early tendon marker Scleraxis (SCX, iMSCSCX+, iTenocytes) in combination with an elastic collagen scaffold. Achilles tendon defects in Nude rat models were created by isolating the tendon and excising 3 mm of the midsection. The Achilles tendon defects were then repaired with iTenocyte-seeded scaffolds, unseeded scaffolds, or suture only and compared to native Nude rat tendon tissue using gait analyses, biomechanical testing, histology, and immunohistochemistry.ResultsThe results show faster functional recovery of gait in iTenocyte-seeded scaffold group comparing to scaffold only and suture only groups. Both iTenocyte-seeded scaffold and scaffold only treatment groups had improved biomechanical properties when compared to suture only treatment group, however no statistically significant difference was found in comparing the cell seeding scaffold an scaffold only group in terms of biomechanical properties. Immunohistochemistry staining further demonstrated that iTenocytes successfully populated the collagen scaffolds and survived 9 weeks after implantation in vivo. Additionally, the repaired tissue of iTenocyte-treated injuries exhibited a more organized structure when compared to tendon defects that were repaired only with suturing or unseeded scaffolds.ConclusionWe suggest that iTenocyte-seeded DuRepair™ collagen scaffold can be used as potential treatment to regenerate the tendon tissue biomechanically and functionally.

Secreted protein acidic and rich in cysteine (SPARC) is the most abundant glycoprotein in bone and is thought to play a critical role in bone remodeling and homeostasis. However, the effect of SPARC in relation to gender and exercise on bone quality is not well understood. The purpose of this study was to quantify differences in the structural and biomechanical properties between calvarial and femoral bone from male and female wild-type (WT) and SPARC null (SPARC((-/-))) mice as well as the ability of exercise to rescue bone health. Male and female WT and transgenic SPARC((-/-)) mice were given either a fixed or rotating running wheel for exercise. Bone structural, biomechanical, and morphological parameters were quantified using micro computed tomography, push out testing for the calvaria, three-point flexural testing for the femurs, histological and immunofluorescent staining. Similar reductions in structural and biomechanical strength were observed in both male and female SPARC((-/-)) calvaria, most of which were not significantly affected by exercise. In femurs, SPARC((-/-)) had a significant effect on structural parameters in both sexes, but was more pronounced in females with some properties being rescued with running. Interestingly, the effect of SPARC((-/-)) on bone mineral density was only detected in female SPARC((-/-)) mice, not males, and was subsequently rescued with exercise. This study emphasizes the differences between sexes in WT and SPARC((-/-)) mice in regard to structural parameters and biomechanical properties. Research into gender differences can help inform and personalize treatment options to more accurately meet patient needs.