Aims

The localization of necrotic areas has been reported to impact the prognosis and treatment strategy for osteonecrosis of the femoral head (ONFH). Anteroposterior localization of the necrotic area after a femoral neck fracture (FNF) has not been properly investigated. We hypothesize that the change of the weight loading direction on the femoral head due to residual posterior tilt caused by malunited FNF may affect the location of ONFH. We investigate the relationship between the posterior tilt angle (PTA) and anteroposterior localization of osteonecrosis using lateral hip radiographs.

Aims

Pigment epithelium-derived factor (PEDF) is known to induce several types of tissue regeneration by activating tissue-specific stem cells. Here, we investigated the therapeutic potential of PEDF 29-mer peptide in the damaged articular cartilage (AC) in rat osteoarthritis (OA).

Back pain is a leading cause of disability worldwide and it is primarily considered to be triggered by intervertebral disc (IVD) degeneration (IVDD). Current treatments may improve pain and mobility, but carry high costs and fail to address IVD repair or regeneration. As no effective therapeutic approach has been proposed to restore inflamed and degenerated IVDs, there is the urgent need to clarify the key pathomechanism of IVDD, the involvement of inflammation, particularly complement activation in matrix catabolism, and how to target them towards tissue repair/regeneration. Mesenchymal stem cell (MSC)-based therapies have become the focus of several regenerative IVD studies. Although patients in clinical trials reported less pain after cell therapy, the long-term success of cell engraftment is unclear due to the hostile IVD environment. The mechanism-of-action of MSCs is mostly dependent on the secreted soluble factors. Moreover, priming of MSC with interleukin (IL)-1β modulates the secretome content, improving its anti-inflammatory and regenerative effect on IVDD organ culture models. MSC-derived extracellular vesicles (EVs) have also been shown to modulate human IVD cells towards a healthy IVD phenotype in vitro. However, the mechanisms involved in the effect of secretome and EVs, particularly with regard to immunomodulation and matrix metabolism, are not fully understood. Our work investigates the effects of secretome and EVs secreted by IL-1β-primed MSCs to impair IVD matrix degradation and/or improve matrix formation in IVDD

The HIPGEN study funded under EU Horizon 2020 (Grant 7792939) has the aim to investigate the potential of the first regenerative cell therapy for the improvement of recovery after muscle injury in hip fracture patients. For this aim we intramuscularly injected placental derived mesenchymal stromal cells during hip fracture arthroplasty. Despite not having reached the primary endpoint, which was the Short Physical Performance Battery, we could observe an increase in abductor muscle strength and a faster return to balance looking at symmetry in insole measurements during follow up

Osteoarthritis (OA) is the most common joint disease, affecting approximately 16% of the adult population worldwide. The chronic inflammation in the joint leads to the breakdown of cartilage, which leads to permanent pain and limitations in everyday life at an early stage of the disease. To date, there is no therapy that can interrupt the inflammatory state or reverse cartilage damage. The PROTO consortium (funded by the EU Horizon Europe program, Grant 101095635) aims to prevent the development of OA by correcting a pathological biomechanical pattern by a digital training intervention and to treat early stage OA with an innovative allogeneic cell therapy

Mincing cartilage with commercially available shavers is increasingly used for treating focal cartilage defects. This study aimed to compare the impact of mincing bovine articular cartilage using different shaver blades on chondrocyte viability. Bovine articular cartilage was harvested using a scalpel or three different shaver blades (2.5 mm, 3.5 mm, or 4.2 mm) from a commercially available shaver. The cartilage obtained with a scalpel was minced into fragments smaller than 1 mm. 3. All four conditions were cultivated in a culture medium for seven days. After Day 1 and Day 7, metabolic activity, RNA isolation, and gene expression of anabolic (COL2A1, ACAN) and catabolic genes (MMP1, MMP13), Live/Dead staining and visualization using confocal microscopy, and flow cytometric characterization of minced cartilage chondrocytes were measured. The study found that mincing cartilage with shavers significantly reduced metabolic activity after one and seven days compared to scalpel mincing (p<0.001). Gene expression of anabolic genes was reduced, while catabolic genes were increased after day 7 in all shaver conditions. The MMP13/COL2A1 ratio was also increased in all shaver conditions. Confocal microscopy revealed a thin line of dead cells at the lesion site with viable cells below for the scalpel mincing and a higher number of dead cells diffusely distributed in the shaver conditions. After seven days, there was a significant decrease in viable cells in the shaver conditions compared to scalpel mincing (p<0.05). Flow cytometric characterization revealed fewer intact cells and proportionally more dead cells in all shaver conditions compared to the scalpel mincing. Mincing bovine articular cartilage with commercially available shavers reduces the viability of chondrocytes compared to scalpel mincing. This indicates that mincing cartilage with a shaver should be considered a matrix rather than a cell therapy. Further experimental and clinical studies are required to standardize the mincing process with a shaver. Acknowledgements: This study received unrestricted funding from KARL STORZ SE & Co. KG

Mesenchymal stromal cells (MSC) have been proposed as an emerging cell therapy for bone tissue engineering applications. However, the healing capacity of the bone tissue is often compromised by patient's age and comorbidities, such as osteoporosis. In this context, it is important to understand the impact of donor age on the therapeutic potential of MSC. Importantly, the impact on donor age is not restricted to cells themselves but also to their microenvironment that is known to affect cell function. The extracellular matrix (ECM) has an important role in stem cell microenvironment, being able to modulate cell proliferation, self-renewal and differentiation. Decellularized cell-derived ECM (dECM) has been explored for regenerative medicine applications due to its bioactivity and its resemblance to the in vivo microenvironment. Thus, dECM offers the opportunity not only to develop microenvironments with customizable properties for improvement of cellular functions but also as a platform to study cellular niches in health and disease. In this study, we investigated the capacity of the microenvironment to rescue the impaired proliferative and osteogenic potential of aged MSC. The goal of this work was to understand if the osteogenic capacity of MSC could be modulated by exposure to a dECM derived from cells obtained from young donors. When aged MSC were cultured on dECM derived from young MSC, their in vitro proliferative and osteogenic capacities were enhanced. Our results suggest that the microenvironment, specifically the ECM, plays a crucial role in the osteogenic differentiation capacity of MSC. dECM might be a valuable clinical strategy to overcome the age-related decline in the osteogenic potential of MSC by recapitulating a younger microenvironment, attenuating the effects of aging on the stem cell niche. Overall, this study opens new possibilities for developing clinical strategies for elderly patients with limited bone formation capacity who currently lack effective treatments. Acknowledgements: The authors thank FCT for funding through the project DentalBioMatrix (PTDC/BTM-MAT/3538/2020) and to the research institutions iBB (UIDB/04565/2020 and UIDP/04565/2020) and Associate Laboratory i4HB (LA/P/0140/2020)

Despite osteoarthritis (OA) representing a large burden for healthcare systems, there remains no effective intervention capable of regenerating the damaged cartilage in OA. Mesenchymal stromal cells (MSCs) are adult-derived, multipotent cells which are a candidate for musculoskeletal cell therapy. However, their precise mechanism of action remains poorly understood. The effects of an intra-articular injection of human bone-marrow derived MSCs into a knee osteochondral injury model were investigated in C57Bl/6 mice. The cell therapy was retrieved at different time points and single cell RNA sequencing was performed to elucidate the transcriptomic changes relevant to driving tissue repair. Mass cytometry was also used to study changes in the mouse immune cell populations during repair. Histological assessment reveals that MSC treatment is associated with improved tissue repair in C57Bl/6 mice. Single cell analysis of retrieved human MSCs showed spatial and temporal transcriptional heterogeneity between the repair tissue (in the epiphysis) and synovial tissue. A transcriptomic map has emerged of some of the distinct genes and pathways enriched in human MSCs isolated from different tissues following osteochondral injury. Several MSC subpopulations have been identified, including proliferative and reparative subpopulations at both 7 days and 28 days after injury. Supported by the mass cytometry results, the immunomodulatory role of MSCs was further emphasised, as MSC therapy was associated with the induction of increased numbers of regulatory T cells correlating with enhanced repair in the mouse knee. The transcriptomes of a retrieved MSC therapy were studied for the first time. An important barrier to the translation of MSC therapies is a lack of understanding of their heterogeneity, and the consequent lack of precision in its use. MSC subpopulations with different functional roles may be implicated in the different phases of tissue repair and this work offers further insights into repair process

Abstract. Objectives. Bone marrow aspirate concentrate (BMAC), together with fibrin glue (Tisseel, Baxter, UK) and Hyaluronic acid (HA) were used as a one-step cell therapy treating patients with ankle cartilage defects in our hospital. This therapy was proven to be safe, with patients demonstrating a significant improvement 12 months post-treatment. Enriched mesenchymal stem cells (MSCs) in BMAC are suggested inducers of cartilage regeneration, however, currently there is no point-of-care assessment for BMAC quality; especially regarding the proportion of MSCs within. This study aims to characterise the cellular component of CCR-generated BMAC using a point-of-care device, and to investigate if the total nucleated cell (TNC) count and patient age are predictive of MSC concentration. Methods. During surgery, 35ml of bone marrow aspirate (BMA) was collected from each patients’ iliac crest under anaesthesia, and BMAC was obtained via a commercial kit (Cartilage Regeneration kit, CCR, Innotec. ®. , UK). BMAC was then mixed with thrombin (B+T) for injection with HA and fibrinogen. In our study, donor-matched BMA, BMAC and B+T were obtained from consented patients (n=12, age 41 ± 16years) undergoing surgery with BMAC therapy. TNC, red blood cell (RBC) and platelet (PLT) counts were measured via a haematology analyser (ABX Micros ES 60, Horiba, UK), and the proportion of MSCs in BMA, BMAC and B+T were assessed via colony forming unit-fibroblast (CFU-F) assays. Significant differences data in matched donors were tested using Friedman test. All data were shown as mean ± SD. Results. Mean TNC counts in BMA and BMAC were not significantly different (14.0 ± 4.4 million/ml and 19.4 ± 32.9 million/ml, respectively, P>0.9999). However, TNC counts were significantly lower in B+T compared to BMAC (9.7 ± 24.5 million/ml and 19.4 ± 32.9 million/ml, respectively, P=0.0167). Similarly, PLT counts were decreased in B+T compared to BMAC (40.7 ± 30.7 million/ml and 417.5 ± 365.5 million/ml, respectively, P<0.0001), however, PLTs were significantly concentrated in BMAC compared to BMA (417.5 ± 365.5 million/ml and 114.8 ± 61.6 million/ml, respectively, P=0.0429). RBC counts were significantly decreased in BMAC and B+T compared to BMA (P=0.0322 and P<0.0001, respectively). Higher concentration of MSCs were observed in BMAC compared to BMA (0.006% ± 0.01% and 0.00007% ± 0.0001%, respectively, P=0.0176). Similar to TNCs and PLTs, the proportion of MSCs significantly decreased in B+T compared to BMAC (0.0004% ± 0.001% and 0.006% ± 0.01%, respectively, P=0.0023). Furthermore, patient age and TNC counts did not correlate with MSC concentration (Spearman's Rank test, P=0.3266 and P=0.4880, respectively). Conclusions. BMAC successfully concentrated PLTs, but BMAC preparations were highly variable. Mixing BMAC and thrombin however, as described in the CCR protocol, resulted in a dramatic reduction in TNCs, PLTs and MSCs. TNC counts and patient age could not be used to predict the MSC proportion in the BMAC based on current data. Future work aims to look at the biomolecule profile of BMAC plasma, and to correlate them to patient clinical outcomes. Declaration of Interest. (a) fully declare any financial or other potential conflict of interest

Aims

Astragalus polysaccharide (APS) participates in various processes, such as the enhancement of immunity and inhibition of tumours. APS can affect osteoporosis (OP) by regulating the osteogenic differentiation of human bone mesenchymal stem cells (hBMSCs). This study was designed to elucidate the mechanism of APS in hBMSC proliferation and osteoblast differentiation.

Aims

Implantation of ultra-purified alginate (UPAL) gel is safe and effective in animal osteochondral defect models. This study aimed to examine the applicability of UPAL gel implantation to acellular therapy in humans with cartilage injury.

Due to the presence of megakaryocytes, platelets and clotting factors, bone marrow aspirate (BMA) tends to coagulate. For the first time, starting from our previous studies on mesenchymal vertebral stem cells, it has been hypothesized that coagulated BMA represents a safe and effective autologous biological scaffold for bone regeneration in spinal surgery. The present research involved advanced preclinical in vitro models and the execution of a pilot clinical study.

Evaluation of cell morphology, growth kinetics, immunophenotyping, clonogenicity, trilineage-differentiation, growth-factors and HOX and TALE gene expression were analyzed on clotted- and un-clotted human V-BMA. In parallel, a pilot clinical study on ten patients with degenerative spine diseases submitted to instrumented posterior arthrodesis, is ongoing to assess the ability of clotted-V-BMA to improve spinal fusion at 6- and 12-months follow-up.

Results demonstrated that clotted-V-BMA have significantly higher growth-factor expression and mesenchymal stem cell (MSCs) viability, homogeneity, clonogenicity, and ability to differentiate towards the osteogenic phenotype than un-clotted-V-BMA. Clotted-V-BMA also highlighted significant reduced expression of PBX1 and of MEIS3 genes negatively involved in osteoblast maturation and differentiation. From December 2020, eight patients have already been enrolled with first promising results that will be finally evaluated in the next two months.

The application of V-BMA-clot as carrier of progenitors and cytokines and as natural scaffold with a structural texture represents a point-of-care orthobiologic product to improve spinal fusion. Clinical application seems to be efficacy, and we will confirm and strengthen these data with the final results of the pilot clinical study.

Osteochondral injuries are a recognised factor in the development of osteoarthritis (OA). Mesenchymal stromal cells (MSCs) represent a promising biological therapeutic option as an OA-modifying treatment, and they also secrete factors that may have an anti-catabolic effect and/or encourage endogenous repair. We aim to study the effects of (i) intra-articular injection of human bone-marrow-derived MSCs and (ii) their secretome on recovery in a murine knee osteochondral injury model. The MSC secretome was generated by stimulating human bone-marrow-derived MSCs with tumour necrosis factor alpha (TNFα). Mice (n=48) were injected with i) MSC secretome, ii) MSCs or iii) cell culture medium (control). Pain was assessed by activity monitoring, and cartilage repair, subchondral bone volume and synovial inflammation were evaluated using histology and microCT. Both MSC- and MSC-secretome-injected mice showed significant pain reduction at day 7 when compared to control mice, but only the MSC-injected mice maintained a significant improvement over the controls at day 28. Cartilage repair was significantly improved in MSC-injected mice. No significant effects were observed with regards to synovial inflammation or subchondral bone volume. The MSC secretome demonstrates regenerative effects but this does not appear to be as sustained as a MSC cell therapy. Further studies are required to investigate if this can be overcome using different dosing regiments for injection of the MSC secretome. As we further understand the regenerative properties of the MSC secretome, we may be able to enhance the clinical translatability of these therapies. Direct intra-articular injection of MSCs for the treatment of OA also appears promising as a potential future strategy for OA management. Acknowledgements: MS is supported by a grant from the Wellcome Trust (PhD Programme for Clinicians)

Autologous tendon cell injection (ATI) is a promising non-surgical treatment for tendinopathies and tendon tear that address its underlying pathology. The procedure involves harvesting autologous tendon tissue, the isolation of the tendon cells, expansion under quality assured GMP cell laboratory and the injection of the tendon cells via U/S into the degenerative tendon tissue. In clinical practice, the patella (PT) and palmaris longus (PL) tendons are common sites used for tendon tissue biopsy. The objective of this study is to compare the tendon cell quality, identity, purity, doubling time and yield of cells between PT and PL tendons for ATI. Tendon tissue biopsies were harvested from PT via U/S using a 14-gauge needle or resected surgically from the PL tendon. The biopsies were transported to a GMP cell laboratory, where tendon cells were isolated, cultured and expanded for 4 to 6 weeks, and analysed for viability, cell doubling time, cellular characteristics including cell purity, potency and identity (PPI). Tendon samples from 149 patients were analysed (63 PT). Average biopsy weight was 62mg for PT and 119mg for PI (p<0.001). Average cell doubling time (83.9 vs 82.7 hours), cellular yield (16.2 vs 15.2x106), viability (98.7 vs 99.0%) and passage number (3 vs 3) were not significantly different between tendons. Additionally, ddPCR analyses showed no differences of PPI including tendon cell markers of collagen type I, scleraxis and tenomodulin. No post-biopsy complications or contamination were reported for either group. Assessing tendon tissue from palmaris tendon is relatively easier. Tendon tissue biopsy tissue for autologous tendon cell therapy can be obtained from either the PT or PL tendons. Tendon cells isolated from PT and PL were equal in growth characteristics and PPI. There are no differences in the quality of tendon cells isolated from the PT or PL

Cite this article: Bone Joint Res 2023;12(1):5–8.

Aims. Successful cell therapy in hip osteonecrosis (ON) may help to avoid ON progression or total hip arthroplasty (THA), but the achieved bone regeneration is unclear. The aim of this study was to evaluate amount and location of bone regeneration obtained after surgical injection of expanded autologous mesenchymal stromal cells from the bone marrow (BM-hMSCs). Methods. A total of 20 patients with small and medium-size symptomatic stage II femoral head ON treated with 140 million BM-hMSCs through percutaneous forage in the EudraCT 2012-002010-39 clinical trial were retrospectively evaluated through preoperative and postoperative (three and 12 months) MRI. Then, 3D reconstruction of the original lesion and the observed postoperative residual damage after bone regeneration were analyzed and compared per group based on treatment efficacy. Results. The mean preoperative lesion volume was 18.7% (SD 10.2%) of the femoral head. This reduced to 11.6% (SD 7.5%) after three months (p = 0.015) and 3.7% (SD 3%) after one year (p < 0.001). Bone regeneration in healed cases represented a mean 81.2% (SD 13.8%) of the initial lesion volume at one year. Non-healed cases (n = 1 stage progression; n = 3 THAs) still showed bone regeneration but this did not effectively decrease the ON volume. A lesion size under mean 10% (SD 6%) of the femoral head at three months predicted no ON stage progression at one year. Regeneration in the lateral femoral head (C2 under Japanese Investigation Committee (JCI) classification) and in the central and posterior regions of the head was predominant in cases without ON progression. Conclusion. Bone regeneration was observed in osteonecrotic femoral heads three months after expanded autologous BM-hMSC injection, and the volume and location of regeneration indicated the success of the therapy. Cite this article: Bone Joint Res 2022;11(12):881–889

Intervertebral disc degeneration (IDD) affects more than 80% of the population all over the world. Current strategies for the treatment of IDD are based on conservative or surgical procedures with the aim of relieving pain. Mesenchymal stem cell (MSC) transplantation has emerged as a promising therapy in recent decades, but studies showed that the particularly hostile microenvironment in the intervertebral disc (IVD) can compromise cells survival rate. The use of exosomes, extracellular vesicles released by various cell types, possess considerable economic advantages including low immunogenicity and toxicity. Exosomes allow intercellular communication by conveying functional proteins, RNA, miRNA and lipids between cells. The purpose of this study is to assess the therapeutic effects of exosomes derived from Wharton Jelly mesenchymal stromal cells (WJ-MSC) on human nucleuspulposus cells (hNPC) in an in vitro 3D culture model. Exosomes (exos) were isolated by tangential flow filtration of WJ-MSC conditioned media and characterized by: quantification with BCA test; morphological observation with TEM, surface marker expression by WB and size evaluation by NTA. Confocal microscopy has been used to identify exosomes marked with PKH26 and monitor fusion and/or incorporation in hNPC. hNPC were isolated from waste surgical material from patients undergoing discectomy (n = 5), expanded, encapsulated in alginate beads and treated with: culture medium (control group); WJ-MSC exos (WJ-exos) at different concentrations (10 μg/ml, 50 μg/ml and 100 μg/ml). They were then analysed for: cell proliferation (Trypan Blu); viability (Live/Dead Assay); quantification of nitrites (Griess) and glycosaminoglycans, GAG (DMBB). The hNPC in alginate beads treated for 7 days were included in paraffin and histologically analysed to determine the presence of extracellular matrix (ECM) components. Finally, the expression levels of catabolic and anabolic genes were evaluated through real-time polymerase chain reaction (qPCR). All concentrations of WJ-exos under exam were capable to induce a significant increase in cell proliferation after 10 and 14 days of treatment (p < 0.01 and p < 0.001, respectively). Live/Dead assay showed a decrease in cell death at 50 μg/ml of WJ-exos (p < 0.05). While cellular oxidative stress indicator, nitrite production, was reduced in a dose-dependent way and statistically significant only with 100 μg/ml of WJ-exos (p < 0.05). WJ-exos at 10 and 100 μg/ml induced a significant increase in GAG content (p < 0.05; p < 0.01, respectively) confirmed by Alcian Blu staining. Exos derived from WJ-MSC modulated gene expression levels by increasing expression of ACAN and SOX-9 genes and reducing significantly of IL-6, MMP-1, MMP-13 and ADAMTS-5 levels (p < 0.05; p < 0.01) compared to the control group. Our results supported the potential use of exosomes from WJ-MSC for the treatment of IDD. Exosomes improved hNPC growth, attenuated ECM degradation and reduced oxidative stress and inflammation. This study offers a new scenario in IVD regeneration, promoting the potential use of extracellular vesicles as an alternative strategy to cell therapy

Aims

Autologous chondrocyte implantation (ACI) is a promising treatment for articular cartilage degeneration and injury; however, it requires a large number of human hyaline chondrocytes, which often undergo dedifferentiation during in vitro expansion. This study aimed to investigate the effect of suramin on chondrocyte differentiation and its underlying mechanism.

Aims. The aim of the HIPGEN consortium is to develop the first cell therapy product for hip fracture patients using PLacental-eXpanded (PLX-PAD) stromal cells. Methods. HIPGEN is a multicentre, multinational, randomized, double-blind, placebo-controlled trial. A total of 240 patients aged 60 to 90 years with low-energy femoral neck fractures (FNF) will be allocated to two arms and receive an intramuscular injection of either 150 × 10. 6. PLX-PAD cells or placebo into the medial gluteal muscle after direct lateral implantation of total or hemi hip arthroplasty. Patients will be followed for two years. The primary endpoint is the Short Physical Performance Battery (SPPB) at week 26. Secondary and exploratory endpoints include morphological parameters (lean body mass), functional parameters (abduction and handgrip strength, symmetry in gait, weightbearing), all-cause mortality rate and patient-reported outcome measures (Lower Limb Measure, EuroQol five-dimension questionnaire). Immunological biomarker and in vitro studies will be performed to analyze the PLX-PAD mechanism of action. A sample size of 240 subjects was calculated providing 88% power for the detection of a 1 SPPB point treatment effect for a two-sided test with an α level of 5%. Conclusion. The HIPGEN study assesses the efficacy, safety, and tolerability of intramuscular PLX-PAD administration for the treatment of muscle injury following arthroplasty for hip fracture. It is the first phase III study to investigate the effect of an allogeneic cell therapy on improved mobilization after hip fracture, an aspect which is in sore need of addressing for the improvement in standard of care treatment for patients with FNF. Cite this article: Bone Jt Open 2022;3(4):340–347

Abstract. Objectives. Mesenchymal stromal/stem cells (MSCs) are increasingly recognized as regulators of immune cells during disease or tissue repair. During these situations, the extracellular matrix (ECM) is very dynamic and therefore, our studies aim to understand how ECM influences the activity of MSCs. Methods. Human MSCs cultured on tissue culture plastic (TCP) and encapsulated within collagen type I, fibrin, or mixed Collagen-Fibrin were exposed to low dose TNFα and IFNɣ. Transcription profiles were examined using bulk RNA sequencing (RNAseq) after 24h of treatment. ELISA, Western blot, qPCR and immunofluorescence were employed to validate RNAseq results and to investigate the significance of transcriptional changes. Flow cytometry evaluated monocyte/macrophage phenotype. Results. Previously, we showed that human MSC expression of TNFAIP6 and CXCL10 in 3D environments is significantly upregulated in response to pro-inflammatory stimuli. Here, RNAseq revealed that there were 2,085 highly significant upregulated genes in 3D matrices compared to TCP. Notably, >90% of highly expressed genes (including FOSB, FOS and TNFAIP6) were shared in all hydrogels. Gene ontology confirmed the TNF signalling pathway among the most significantly represented. Protein-protein interaction predictions identified TNF-alpha/NF-kappa B and AP1 pathways as differentially influenced by the hydrogel environment. Using inhibitors to these pathways, NFkB, but not AP1, impacted on the upregulation of TNFAIP6 and CXCL10 in 3D culture. Conditioned media from these studies was added to cultures of human monocytes with distinct changes in the resulting macrophage phenotype. MSCs in a 3D environment promoted a greater acquisition of the M2 repair macrophage phenotype and impacted on the numbers of pro-inflammatory M1 macrophages. Conclusion. These data provide further evidence that the immunomodulatory action of human MSCs can be influenced by the surrounding structural environment. These observations have significance for understanding the events that following skeletal injury and the potential to be exploited in preconditioning MSCs for cell therapy