Pelvic bone defect in patients with severe congenital dysplasia of the hip (CDH) lead to abnormalities in lumbar spine and lower limb alignment that can determine total hip arthroplasty (THA) patients' outcome. These variables may be different in uni- or bilateral CDH.

We compared the clinical outcome and the spinopelvic and lower limb radiological changes over time in patients undergoing THA due to uni- or bilateral CHD at a minimum follow-up of five years.

Sixty-four patients (77 hips) undergoing THA due to severe CDH between 2006 and 2015 were analyzed: Group 1 consisted of 51 patients with unilateral CDH, and group 2, 113 patients (26 hips) with bilateral CDH. There were 32 females in group 1 and 18 in group 2 (p=0.6). The mean age was 41.6 years in group 1 and 53.6 in group 2 (p<0.001). We compared the hip, spine and knee clinical outcomes. The radiological analysis included the postoperative hip reconstruction, and the evolution of the coronal and sagittal spinopelvic parameters assessing the pelvic obliquity (PO) and the sacro-femoro-pubic (SFP) angles, and the knee mechanical axis evaluating the tibio-femoral angle (TFA).

At latest follow-up, the mean Harris Hip Score was 88.6 in group 1 and 90.7 in group 2 (p=0.025). Postoperative leg length discrepancy of more than 5 mm was more frequent in group 1 (p=0.028). Postoperative lumbar back pain was reported in 23.4% of the cases and knee pain in 20.8%, however, there were no differences between groups. One supracondylar femoral osteotomy and one total knee arthroplasty were required. The radiological reconstruction of the hip was similar in both groups. The PO angle improved more in group 1 (p=0.01) from the preoperative to 6-weeks postoperative and was constant at 5 years. The SFP angle improved in both groups but there were no differences between groups (p=0.5). 30 patients in group 1 showed a TFA less than 10º and 17 in group 2 (p=0.7).

Although the clinical outcome was better in terms of hip function in patients with bilateral CDH than those with unilateral CDH, the improvement in low back and knee pain was similar. Patients with unilateral dysplasia showed a better correction of the PO after THA. All spinopelvic and knee alignment parameters were corrected and maintained over time in most cases five years after THA.

Growth factors produced by inflammatory cells and mesenchymal progenitors are required for proper bone regeneration. Signaling pathways activated downstream of these proteins work in concert and synergistically to drive osteoblast and/or chondrocyte differentiation. While dysregulation can result in abnormal healing, activating these pathways in the correct spatiotemporal context can enhance healing. Bone morphogenetic protein (BMP) signaling is well-recognized as being required for bone regeneration, and BMP is used clinically to enhance bone healing. However, it is imperative to develop new therapeutics that can be used alone or in conjunction with BMP to drive even more robust healing. Notch signaling is another highly conserved signaling pathway involved in tissue development and regeneration. Our work has explored Notch signaling during osteoblastogenesis and bone healing using both in vitro studies with human primary mesenchymal progenitor cells and in vivo studies with genetically modified mouse models. Notch signaling is required and sufficient for osteoblast differentiation, and is required for proper bone regeneration. Indeed, intact Notch signaling through the Jagged-1 ligand is required for BMP induced bone formation. On-going work continues to explore the intersection between BMP and Notch signaling, and determining cell types that express Notch receptors and Notch ligands during bone healing. Our long-term objective is to develop Notch signaling as a clinical therapy to repair bone.

Precision health aims to develop personalised and proactive strategies for predicting, preventing, and treating complex diseases such as osteoarthritis (OA). Due to OA heterogeneity, which makes developing effective treatments challenging, identifying patients at risk for accelerated disease progression is essential for efficient clinical trial design and new treatment target discovery and development.

To create a reliable and interpretable precision health tool that predicts rapid knee OA progression over a 2-year period from baseline patient characteristics using an advanced automated machine learning (autoML) framework, “Autoprognosis 2.0”.

All available 2-year follow-up periods of 600 patients from the FNIH OA Biomarker Consortium were analysed using “Autoprognosis 2.0” in two separate approaches, with distinct definitions of clinical outcomes: multi-class predictions (categorising disease progression into pain and/or radiographic progression) and binary predictions. Models were developed using a training set of 1352 instances and all available variables (including clinical, X-ray, MRI, and biochemical features), and validated through both stratified 10-fold cross-validation and hold-out validation on a testing set of 339 instances. Model performance was assessed using multiple evaluation metrics. Interpretability analyses were carried out to identify important predictors of progression.

Our final models yielded higher accuracy scores for multi-class predictions (AUC-ROC: 0.858, 95% CI: 0.856-0.860) compared to binary predictions (AUC-ROC: 0.717, 95% CI: 0.712-0.722). Important predictors of rapid disease progression included WOMAC scores and MRI features. Additionally, accurate ML models were developed for predicting OA progression in a subgroup of patients aged 65 or younger.

This study presents a reliable and interpretable precision health tool for predicting rapid knee OA progression. Our models provide accurate predictions and, importantly, allow specific predictors of rapid disease progression to be identified. Furthermore, the transparency and explainability of our methods may facilitate their acceptance by clinicians and patients, enabling effective translation to clinical practice.

After knee replacement, therapy resistant, chronic synovitis is common and leads to effusion and pain.

A cohort of 55 patients with 57 knee replacements and chronic synovitis underwent radiosynoviorthesis. In summary, 101 joints were treated using 182±9 MBq of 90Y-citrate. The number of radiosynoviorthesis ranged from 1 to 4 (53%, 21%, 23%, and 4%). Every patient received a 99mTc-MDP scintigraphy before and three months after every radiosynoviorthesis. Follow-up ranged from 5.7 to 86.7 months. For qualitative analysis, an four steps scoring was used (0 = no response or worsening, 1 = slight, 2 = good, 3 = excellent response). For quantification, the uptake was determined within the 99mTc-MDP scintigraphy soft tissue phase before and after therapy.

At the end of long-term follow-up 27% of patients have an excellent, 24% good, 30% slight and 20% no response. The duration of response was 7.5±8.3 months (maximum 27 months). In repeated treatment, the effect after the first therapy was lesser than in patients who received a single treatment in total. However, three months after the last radiosynoviorthesis, patients with repeated treatment showed a similar effectiveness than single treated patients. At the end of long-term follow-up, patients with repeated radiosynoviorthesis had a higher effectiveness at similar duration response. In the 99mTc-MDP scan 65% of patients showed a reduction of uptake. When comparing subjective and objective response 78% of patients showed a concordance in both, symptoms and scintigraphy. Pilot histological analysis revealed that the synovitis is triggered by small plastic particles.

Radiosynoviorthesis is effective in patients with knee replacement and chronic synovitis. It shows good subjective and objective response rates and long response duration. Repeated treatment leads to a stronger long-time response. The chronic synovitis is caused by plastic particles, which result from the abrasion of the polymeric inlay of endoprothesis.

Despite the clinical relevance of back pain and intervertebral disc herniation, the lack of reliable models has strained their molecular understanding. We characterized the lumbar spinal phenotype of C57BL/6 and SM/J mice during aging. Interestingly, old SM/J lumbar discs evidenced accelerated degeneration, associated with high rates of disc herniation. SM/J AF's and degenerative human's AF transcriptomic profiles showed altered immune cell, inflammation, and p53 pathways. Old SM/J mice presented increased neuronal markers in herniated discs, thicker subchondral bone, and higher sensitization to pain. Dorsal root ganglia transcriptomic studies and spinal cord analysis exhibited increased pain and neuroinflammatory markers associated with altered extracellular matrix regulation. Immune system single-cell and tissue level analysis showed distinctive T-cell and B-cell modulation and negative correlation between mechanical allodynia and INF-α, IL-1β, IL2, and IL4, respectively. This study underscores the multisystemic network behind back pain and highlights the role of genetic background and the immune system in disc herniation disease.

Acknowledgments: This study is supported by grants from the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) R01AR055655, R01AR064733, R01AR074813 to MVR.

Cells typically respond to a variety of geometrical cues in their environment, ranging from nanoscale surface topography to mesoscale surface curvature. The ability to control cellular organisation and fate by engineering the shape of the extracellular milieu offers exciting opportunities within tissue engineering. Despite great progress, however, many questions regarding geometry-driven tissue growth remain unanswered.

Here, we combine mathematical surface design, high-resolution microfabrication, in vitro cell culture, and image-based characterization to study spatiotemporal cell patterning and bone tissue formation in geometrically complex environments. Using concepts from differential geometry, we rationally designed a library of complex mesostructured substrates (10¹-10³ µm). These substrates were accurately fabricated using a combination of two-photon polymerisation and replica moulding, followed by surface functionalisation. Subsequently, different cell types (preosteoblasts, fibroblasts, mesenchymal stromal cells) were cultured on the substrates for varying times and under varying osteogenic conditions. Using imaging-based methods, such as fluorescent confocal microscopy and second harmonic generation imaging, as well as quantitative image processing, we were able to study early-stage spatiotemporal cell patterning and late-stage extracellular matrix organisation. Our results demonstrate clear geometry-dependent cell patterning, with cells generally avoiding convex regions in favour of concave domains. Moreover, the formation of multicellular bridges and collective curvature-dependent cell orientation could be observed. At longer time points, we found clear and robust geometry-driven orientation of the collagenous extracellular matrix, which became apparent with second harmonic generation imaging after ∼2 weeks of culture.

Our results highlight a key role for geometry as a cue to guide spatiotemporal cell and tissue organisation, which is relevant for scaffold design in tissue engineering applications. Our ongoing work aims at understanding the underlying principles of geometry-driven tissue growth, with a focus on the interactions between substrate geometry and mechanical forces.

The advent of modular implants aims to minimise morbidity associated with revision of hemiarthroplasty or total shoulder arthroplasty (TSA) to reverse shoulder arthroplasty (RSR) by allowing retention of the humeral stem. This systematic review aimed to summarise outcomes following its use and reasons why modular humeral stems may be revised.

A systematic review of Pubmed, Medline and EMBASE was performed according to PRISMA guidelines of all patients undergoing revision of a modular hemiarthroplasty or TSA to RSR. Primary implants, glenoid revisions, surgical technique and opinion based reports were excluded. Collected data included demographics, outcomes and incidence of complications.

277 patients were included, with a mean age of 69.8 years (44-91) and 119 being female. Revisions were performed an average of 30 months (6-147) after the index procedure, with the most common reason for revision being cuff failure in 57 patients. 165 patients underwent modular conversion and 112 underwent stem revision. Of those that underwent humeral stem revision, 18 had the stem too proximal, in 15 the stem was loose, 10 was due to infection and 1 stem had significant retroversion. After a mean follow up of 37.6 months (12-91), the Constant score improved from a mean of 21.8 to 48.7. Stem revision was associated with a higher complication rate (OR 3.13, 95% CI 1.82-5.39).

The increased use of modular stems has reduced stem revision, however 40% of these implants still require revision due to intra-operative findings. Further large volume comparative studies between revised and maintained humeral stems post revision of modular implants can adequately inform implant innovation to further improve the stem revision rate.

Critical-sized bone defects remain challenging in the clinical setting. Autologous bone grafting remains preferred by clinicians. However, the use of autologous tissue is associated with donor-site morbidity and limited accessibility to the graft tissue. Advances in the development of synthetic bone substitutes focus on improving their osteoinductive properties. Whereas osteoinductivity has been demonstrated with ceramics, it is still a challenge in case of polymeric composites. One of the approaches to improve the regenerative properties of biomaterials, without changing their synthetic character, is the addition of inorganic ions with known osteogenic and angiogenic properties. We have previously reported that the use of a bioactive composite with high ceramic content composed of poly(ethyleneoxide terephthalate)/poly(butylene terephthalate) (1000PEOT70PBT30, PolyActive, PA) and 50% beta-tricalcium phosphate (β-TCP) with the addition of zinc in a form of a coating of the TCP particles can enhance the osteogenic differentiation of human mesenchymal stromal cells (hMSCs) (3). To further support the regenerative properties of these scaffolds, inorganic ions with known angiogenic properties, copper or cobalt, were added to the coating solution.

β-TCP particles were immersed in a zinc and copper or zinc and cobalt solution with a concentration of 15 or 45 mM. 3D porous scaffolds composed of 1000PEOT70PBT30 and pure or coated β-TCP were additively manufactured by 3D fibre deposition. The osteogenic and angiogenic properties of the fabricated scaffolds were tested in vitro through culture with hMSCs and human umbilical vein endothelial cells, respectively. The materials were further evaluated through ectopic implantation in an in vivo mini-pig model. The early expression of relevant osteogenic gene markers (collagen-1, osteocalcin) of hMSCs was upregulated in the presence of lower concentration of inorganic ions. Further analysis will focus on the evaluation of ectopic bone formation and vascularisation of these scaffolds after implantation in a mini-pig ectopic intramuscular model.

Smartphones are often equipped with inertial sensors capable of measuring individuals' physical activities. Their role in monitoring the patients' physical activities in telemedicine, however, needs to be explored. The main objective of this study was to explore the correlation between a participant's daily step counts and the daily step counts reported by their smartphone. This prospective observational study was conducted on patients undergoing lower limb orthopedic surgery and a group of non-patients. The data collection period was from 2 weeks before until four weeks after the surgery for the patients and two weeks for the non-patients. The participants' daily steps were recorded by physical activity trackers employed 24/7, and an application recorded the number of daily steps registered by the participants' smartphones. We compared the cross-correlation between the daily steps time-series taken from the smartphones and physical activity trackers in different groups of participants. We also employed mixed modeling to estimate the total number of steps. Overall, 1067 days of data were collected from 21 patients (11 females) and 10 non-patients (6 females). The cross-correlation coefficient between the smartphone and physical activity tracker was 0.70 [0.53–0.83]. The correlation in the non-patients was slightly higher than in the patients (0.74 [0.60–0.90] and 0.69 [0.52–0.81], respectively). Considering the ubiquity, convenience, and practicality of smartphones, the high correlation between the smartphones and the total daily step time-series highlights the potential usefulness of smartphones in detecting the change in the step counts in remote monitoring of the patient's physical activity.

Tendinopathy is the most common form of chronic tendon disorders, accounting for up 30% of all musculoskeletal clinic visits [1]. In tendon disease, the largely avascular tendon tissue often becomes hypervascularized and fibrotic [2]. As blood vessel growth and angiogenic signaling molecules are often induced by the lack of adequate nutrients and oxygen, hypoxic signaling is speculated to be a root cause of tendon neovascularization and tendinopathy [3,4,5]. However, how the vascular switch is initiated in tendons, and how vascularization contributes to tendon pathology remains unknown. In this talk, we provide evidence that HIF-1α is implicated in tendon disease and HIF-1α stabilization in human tendon cells induces vascular recruitment of endothelial cells via VEGFa secretion. More interesting, HIF-1α stabilization in tendon cells in vivo, seems to recapitulate all main features of fibrotic human tendon disease, including vascular ingrowth, matrix disorganization, changes in tissue mechanics, cell proliferation and innervation. Surprisingly, in vivo knock-out of VEGFa rescued angiogenesis in the tendon core but it did not affect tendon mechanical properties and tissue pathophysiological changes, suggesting that blood vessels ingrowth might not be a primary cause but a consequence of HIF-1α activation.

Several synthetic polymers have been widely investigated for their use in bone tissue engineering applications, but the ideal material is yet to be engineered. Triazine-trione (TATO) based materials and their derivatives are novel in the field of biomedical engineering but have started to draw interest. Different designs of the TATO monomers and introduction of different chemical linkages and end-groups widens the scope of the materials due to a range of mechanical properties.

The aim of our work is to investigate novel TATO based materials, with or without hydroxyapatite filler, for their potential in bone tissue engineering constructs. Initially the biocompatibility of the materials was tested, indirectly and directly, according to ISO standards. Following this the osteoconductive properties were investigated with primary osteoblasts and an osteoblastic cell line. Bone marrow derived mesenchymal stem cells were used to evaluate the osteogenic differentiation and consequently the materials potential in bone tissue engineering applications.

Orthopaedic soft tissues, such as tendons, ligaments, and articular cartilage, rely on their unique collagen fiber architectures for proper functionality. When these structures are disrupted in disease or fail to regenerate in engineered tissues, the tissues transform into dysfunctional fibrous tissues. Unfortunately, collagen synthesis in regenerating tissues is often slow, and in some cases, collagen fibers do not regenerate naturally after injury, limiting repair options. One of the research focuses of my team is to develop functional fiber replacements that can promote in vivo repair of musculoskeletal tissues throughout the body. In this presentation, I will discuss our recent advancements in electrowriting 3D printing of natural polymers for creating functional fiber replacements. This manufacturing process utilizes electrical signals to control the flow of polymeric materials through an extrusion nozzle, enabling precise deposition of polymeric fibers with sizes that cannot be achieved using conventional extrusion printing methods. Furthermore, it allows for the formation of fiber organizations that surpass the capabilities of conventional electrospinning processes. During the presentation, I will showcase examples of electrowritten microfiber scaffolds using various naturally-derived polymers, such as gelatin (a denatured form of collagen) and silk fibroin. I will discuss the functional properties of silk-based scaffolds and highlight how they exhibit restored β-sheet and α-helix structures [1]. This restoration results in an elastic response of up to 20% deformation and the ability to withstand cyclic loading without plastic deformation. Additionally, I will present our latest results on the compatibility of this technique with patterning cell-laden fiber structures [2]. This novel biofabrication process allows for the printing of biomimetic microscale architectures with high cell viability, and offers a promising approach to understanding how shear and elongation forces influence cell development of hierarchical (collagen) fibers.

Acknowledgements: The author would like to thank the Reprint project (OCENW.XS5.161) and the program “Materials Driven Regeneration” (024.003.013) by the Netherlands Organization for Scientific Research for the financial support.

The hypoxic nucleus pulposus cells were thought to contain few, functionally redundant mitochondria. However in contrast to this widely held notion, new evidence shows presence of functional mitochondrial networks in disc cells. The lecture will discuss this evidence and provide insights into how microenvironmental cues govern mitochondrial function. The lecture will also discuss emerging evidence on how mitochondrial dysfunction of nucleus pulposus cells results in metabolic dysregulation and acquisition of a state that promotes inflammation and degeneration.

Monomeric C reactive protein (mCRP) presents important proinflammatory effects in endothelial cells, leukocytes, or chondrocytes. However, CRP in its pentameric form exhibits weak anti-inflammatory activity. It is used as a biomarker to follow severity and progression in infectious or inflammatory diseases, such as intervertebral disc degeneration (IVDD). This work assesses for the first time the mCRP effects in human intervertebral disc cells, trying to verify the pathophysiological relevance and mechanism of action of mCRP in the etiology and progression of IVD degeneration.

We demonstrated that mCRP induces the expression of multiple proinflammatory and catabolic factors, like nitric oxide synthase 2 (NOS2), cyclooxygenase 2 (COX2), matrix metalloproteinase 13 (MMP13), vascular cell adhesion molecule 1 (VCAM1), interleukin (IL)-6, IL-8, and lipocalin 2 (LCN2), in human annulus fibrosus (AF) and nucleus pulposus (NP) cells. We also showed that nuclear factor-κβ (NF-κβ), extracellular signal-regulated kinase 1/2 (ERK1/2), and phosphoinositide 3-kinase (PI3K) are at play in the intracellular signaling of mCRP.

Our results indicate that the effect of mCRP is persistent and sustained, regardless of the proinflammatory environment, as it was similar in healthy and degenerative human primary AF cells. This is the first article that demonstrates the localization of mCRP in intravertebral disc cells of the AF and NP and that provides evidence for the functional activity of mCRP in healthy and degenerative human AF and NP disc cells.

A major obstacle in biofabrication is replicating the organization of the extracellular matrix and cellular patterns found in anisotropic tissues within bioengineered constructs. While magnetically-assisted 3D bioprinting techniques have the potential to create scaffolds that mimic natural biological structures, they currently lack the ability to accurately control the dispersion of magnetic substances within the bioinks without compromising the fidelity of the intended composite. To overcome this dichotomy, the concepts of magnetically- and matrix-assisted 3D bioprinting are combined here. This method preserves the resolution of printed structures by keeping low viscosity bioinks uncrosslinked during printing, which allows for the arrangement of magnetically-responsive microfibers without compromising the structural integrity of the design. Solidification is induced after the microfibers are arranged in the desired pattern. Furthermore, the precise design of these magnetic microfillers permits the utilization of low levels of inorganic materials and weak magnetic field strengths, which reduces the potential risks that may be associated with their use. The effectiveness of this approach is evaluated in the context of tendon tissue engineering, and the results demonstrate that combining the tendons like anisotropic fibrous microstructure with remote magneto-mechanical stimulation during in vitro maturation provides both biochemical and biophysical cues that effectively guide human adipose-derived stem cells towards a tenogenic phenotype In summary, the developed strategy allows the fabrication of anisotropic high-resolution magnetic composites with remote stimulation functionalities, opening new horizons for tissue engineering applications.

Acknowledgments: ERC Grant CoG MagTendon nr 772817, BioChips PoC project nr 10106930, (PD/BD/129403/2017), (CEECIND/01375/2017), (2020.03410.CEECIND), (2022.05526.PTDC), (ED481B2019/025).

Intra-articular injection is a common way to deliver biologics to joints, but their effectiveness is limited by rapid clearance from the joint space. This barrier can be overcome by genetically modifying cells within the joint such that they produce anti-arthritic gene products endogenously, thereby achieving sustained, therapeutic, intra-articular concentrations of the transgene products without re-dosing. A variety of non-viral and viral vectors have been subjected to preclinical testing to evaluate their suitability for delivering genes to joints. The first transfer of a gene to a human joint used an ex vivo protocol involving retrovirally transduced, autologous, synovial fibroblasts. Recent advances in vector technology allow in vivo delivery using adeno-associated virus (AAV). We have developed an AAV vector encoding the interleukin-1 receptor antagonist (AAV.IL-1Ra) for injection into joints with osteoarthritis (OA). It showed efficacy and safety in equine and rat models of OA, leading to a recently-completed, investigator-initiated, Phase I, dose-escalation clinical trial in 9 subjects with mid-stage OA of the knee (ClinicalTrials.gov Identifier: NCT02790723). Three cohorts of three subjects with mild to moderate OA in the index knee were injected intra-articularly under ultrasound guidance with a low (10e11 viral genomes) medium (10e12 viral genomes) or high (10e13 viral genomes) dose of AAV.IL-1Ra and followed for one year. The data confirm safety, with evidence of sustained intra-articular expression of IL-1Ra and a clinical response in certain subjects. Funding for a subsequent Phase Ib trial involving 50 subjects (ClinicalTrials.gov Identifier: NCT05835895), expected to start later this year, has been acquired. Progress in this area has stimulated commercial activity and there are now at least seven different companies developing gene therapies for OA and a number of clinical trials are in progress.

Acknowledgement: Clinical trial funded by US Department of Defense Clinical Trial Award W81XWH-16-1-0540.

To date, few studies have investigated the feasibility of the loop-mediated isothermal amplification (LAMP) assay for identifying pathogens in tissue samples. This study aimed to investigate the feasibility of LAMP for the rapid detection of methicillin-susceptible or methicillin-resistant Staphylococcus aureus (MSSA or MRSA) in tissue samples, using a bead-beating DNA extraction method. Twenty tissue samples infected with either MSSA (n = 10) or MRSA (n = 10) were obtained from patients who underwent orthopedic surgery for suspected musculoskeletal infection between December 2019 and September 2020. DNA was extracted from the infected tissue samples using the bead-beating method. A multiplex LAMP assay was conducted to identify MSSA and MRSA infections. To recognize the Staphylococcus genus, S. aureus, and methicillin resistance, 3 sets of 6 primers for the 16S ribosomal ribonucleic acid (rRNA) and the femA and mecA genes were used, respectively. The limit of detection and sensitivity (detection rate) of the LAMP assay for diagnosing MSSA and MRSA infection were analyzed. The results of this study suggest that the LAMP assay performed with tissue DNA samples can be a useful diagnostic method for the rapid detection of musculoskeletal infections caused by MSSA and MRSA.

Deriving autologous mesenchymal stem cells (MSCs) from adipose tissues without using enzymes requires sophisticated biomedical instruments. Applied pressure on tissues and cells are adjusted manually although centrifugation and filtration systems are frequently used. The number of derived MSCs therefore could differ between instruments. We compared the number of MSCs obtained from four commercially available devices and our newly designed and produced instrument (A2, B3, L3, M2 and T3). Three-hundred mL of adipose tissue was obtained from a female patient undergoing liposuction using the transillumination solution. Obtained tissue was equally distributed to each device and handled according to the producers' guides. After handling, 3 mL stromal vascular fraction (SVF) was obtained from each device. Freshly isolated SVF was characterized using multi-color flow cytometry (Navios Flow Cytometer, Beckman Coulter, USA). Cell surface antigens were chosen according to IFATS and ISCT. CD31-FITC, CD34-PC5,5, CD73-PE, CD90-PB and CD45-A750 (Backman Coulter, USA) fluorochrome-labeled monoclonal antibodies were assessed. Markers were combined with ViaKrome (Beckman Coulter, USA) to determine cell viability. At least 10⁵ cells were acquired from each sample. A software (Navios EX, Beckman Coulter, USA) was used to create dot plots and to calculate the cell composition percentages. The data was analyzed in the Kaluza 2.1 software package (Beckman Coulter, USA). Graphs were prepared in GraphPad Prism. CD105 PC7/CD31 FITC cell percentages were 23,9%, 13,5%, 24,6%, 11,4% and 28,8% for the A2, B3, L3, M2 and T3 devices, respectively. We conclude that the isolated MSC percentage ranged from 11,4% to 28,8% between devices. The number of MSCs in SVF are key determinants of success in orthobiological treatments. Developing a device should focus on increasing the number of MSCs in the SVF while preserving its metabolic activity.

Acknowledgments: Scientific and Technological Research Council of Türkiye (TÜBİTAK)- Technology and Innovation Funding Program Directorate (TEYDEB) funded this project (#321893). Servet Kürümoğlu and Bariscan Önder of Disposet Ltd., Ankara, Türkiye (www.disposet.com) contributed to the industrial design and research studies. Ali Tuncel and Feza Korkusuz are members of the Turkish Academy of Sciences (TÜBA). Nilsu Baysal was funded by the STAR Program of TÜBITAK Grant # 3210893.

Osteoarthritis (OA) and diabetis mellitus type 2 (DMT2) are pathogenetically linked. Complement dysregulation contributes to OA and could be involved in DMT2. The inflammatory anaphylatoxin C5a is released during complement activation. This study aims to understand the specific responses of chondrocytes isolated from diabetic and non-diabetic rats exposed to C5a and/or the proinflammatory cytokine TNFα in vitro dependent on the glucose supply. Articular chondrocytes of adult Zucker Diabetic Fatty (ZDF) rats (homozygous: fa/fa, diabetic, heterozygous: fa/+, lean controls) were exposed to 10 ng/mL TNFα and 25 ng/mL C5a alone or in combination, both, under normo- (NG, 1 g/L glucose) and hyperglycemic (HG, 4.5 g/L glucose) conditions (4 or 24 h). Chondrocyte survival, metabolic activity and gene expression of collagen type 2, suppressors of cytokine signaling (SOCS)1, −3 and anti-oxidative hemoxygenase-1 (HMOX1) were assessed. The complement regulatory protein CD46 and cell nuclei sizes were analyzed. Chondrocyte vitality remained unaffected by the treatment. Metabolic activity was impaired in chondrocytes of non-diabetic rats under HG conditions. Collagen type 2 transcription was suppressed by TNFα under HG condition in chondrocytes from nondiabetic donors and under both conditions in those of DMT2 rats (24 h)

Except for DMT2 chondrocytes under HG (4 h), HMOX1 was generally induced by TNFα +/- C5a (NG, HG). C5a elevated HMOX1 only in chondrocytes of controls. The SOCS1/3 genes were increased by TNFα (NG, diabetic, non diabetic, 4 and 24 h). This could also be observed in chondrocytes of diabetic, but not of lean rats (24 h, HG). At 4 h, C5a induced SOCS1 only in non diabetic chondrocytes (NG, HG). Cytoprotective CD46 protein was suppressed by TNFα under NG condition. Nuclear volumes of chondrocyte were lower in chondrocytes from DMT2 rats compared to those from controls. The differential response suggests that chondrocytes are irreversibly compromised by DMT2.

Achnowledgement: The authors are grateful for the support by the “Stiftung Edoprothetik (S 04/21)”

Stem cell therapy for the intervertebral disc (IVD) is highly debated but holds great promises. From previous studies, it is known that notochordal cells are highly regenerative and may stimulate other differentiated cells to produce more matrix. Lately, a particular tissue-specific progenitor cell population has been identified in the centre of the intervertebral disc (IVD. The current hope is that these nucleus pulposus progenitor cells (NPPC) could play a particular role in IVD regeneration.

Current evidence confirms the presence of these cells in murine, canine, bovine and in the human fetal/surgical samples. Noteworthy, one of the main markers to identify these cells, i.e., Tie2, is a typical marker for endothelial cells. Thus, it is not very clear what their origin and their role might be in the context of developmental biology. In human surgical specimens, their presence is, even more, obscured depending on the donor's age and the condition of the IVD and other yet unknown factors.

Here, I revisit the recent literature on regenerative cells identified for the IVD in the past decades. Current evidence how these NPPC can be isolated and detected in various species and tissues will be recapitulated. Future directions will be provided on how these progenitor cells could be used for regenerative medicine and tissue engineering.