Inflammation and Starvation Affect Housekeeping Gene Stability in Adipose Mesenchymal Stromal Cells.

Due to the scientific success of in vitro and in vivo model studies, the interest in using mesenchymal stromal cells (MSCs) for the treatment of orthopaedic conditions is growing. In the context of osteoarthritis (OA), MSCs, and, in particular, those derived from adipose tissues (ASCs), have found broader access to clinical use as active components of minimally manipulated orthobiologics, as well as clinically expanded cell preparations, or to collect their released factors (secretome) for cell-free approaches. In this regard, while both inflammatory priming and starvation are common strategies used to empower cell potency or collect the secretome, respectively, little is known about the possible influence of these approaches on the stability of housekeeping genes (HKGs) for molecular studies able to fingerprint cell phenotype or potency. In this report, the reliability of five commonly used HKGs (ACTB, B2M, GAPDH, HPRT1 and RPLP0) was tested in ASCs cultured under standard protocol after inflammatory priming or starvation. Gene expression data were computed with four different applets able to rank genes depending on their stability in either single or combined conditions. The obtained final ranking suggests that for each treatment, a specific HKG is needed, and that starvation is the condition with the stronger effect on HKGs' stability and, therefore, reliability. The normalization effect of proper HKGs' use was then validated on three genes involved in OA and whose product is released by ASCs. Overall, data presented herein confirm that the choice of the best HKG has to be carefully considered and that each specific condition has to be tested to identify the most reliable candidate.

Men and women undergoing total hip arthroplasty have different clinical presentations before surgery and different outcomes at 1-year follow-up.

PURPOSE: The purpose of this study was to investigate the influence of sex on patients undergoing total hip arthroplasty (THA) for hip osteoarthritis (HOA), aiming to assess the clinical and functional outcomes using patient-reported outcome measures (PROMs). METHODS: A retrospective analysis of patients undergoing THA at Ospedale Galeazzi-Sant'Ambrogio between 2016 and 2022 was conducted. Inclusion criteria encompassed Kellgren-Lawrence grade III or IV HOA, with preoperative and 12-month postoperative PROMs. Enroled patients have been selected from a larger cohort without matching design for confounders. The analyses were performed using R software v4.0.3 (R Core Team) and data distributions were assessed using the Shapiro-Wilk normality test. RESULTS: One hundred ninety patients (72 male and 118 female) who had both preoperative and postoperative PROMs have been analysed from our institutional prosthesis registry (Datareg). Baseline and 12-month post-THA PROMs showed significant improvements overall. VAS score dropped notably from baseline to 3 months postsurgery (7.1 ± 2.1 vs. 0.9 ± 1.7). Functional and mental PROMs, including Harris Hip Score-functional (HHS-F), Harris Hip Score-total (HHS-t), SF-12PS and SF-12MS, exhibited substantial improvements post-THA. Stratifying by sex, males had lower baseline VAS, higher HHS-F, SF-12MS and hip disability and osteoarthritis outcome score-physical function short form (HOOS-PS). At 12 months, males displayed significantly better VAS, HHS-F, SF-12PS and HOOS-PS scores. Complication rates were minimal (1.5%), with stable rates across genders, mostly involving dislocation and periprosthetic fractures. Implant survival at 12 months reached an impressive 99%. CONCLUSION: THA remains an effective treatment for severe HOA. However, females presented with worse baseline conditions and showed relatively less improvement at 1-year postsurgery compared to males. This difference could be attributed to physiological and psychosocial factors associated with sex, including hormonal changes, muscle mass decline and perception of pain. Longer follow-ups and prospective studies are necessary to validate these findings and facilitate personalised approaches in HOA treatment, emphasising the need for careful consideration of sex-related variables in clinical decision-making for THA patients. LEVEL OF EVIDENCE: Level III.

Corticosteroid injections for the treatment of osteoarthritis present a wide spectrum of effects ranging from detrimental to disease-modifying: A systematic review of preclinical evidence by the ESSKA Orthobiologic Initiative.

PURPOSE: Aim of this systematic review of preclinical evidence was to determine the effects of intra-articular corticosteroid (CS) injections in joints affected by osteoarthritis (OA). METHODS: A systematic review was performed on animal studies evaluating intra-articular CS injections for OA joints. The search was performed on PubMed, Cochrane, and Web of Science databases. A synthesis of the results was performed investigating CS effects by evaluating studies comparing CS with control groups. Morphological, histological, immunohistochemistry evaluations, clinical outcomes, biomarkers and imaging results were evaluated. The risk of bias was assessed according to the Systematic Review Centre for Laboratory Animal Experimentation's tool. RESULTS: Thirty-two articles analysing CS effects in OA animal models were included (1079 joints), 18 studies on small and 14 on large animals. CS injections showed overall positive effects in at least one of the outcomes in 68% of the studies, while 16% reported a deleterious effect. CS improved cartilage and synovial outcomes in 68% and 60% of the studies, but detrimental effects were documented in 11% and 20% of the studies, respectively. Clinical parameters evaluated in terms of pain, lameness or joint swelling improved in 63% of the studies but deteriorated in 13%. Evidence is limited on imaging and biomarkers results, as well as on the best CS type, dose, formulation and injection protocol. The risk of bias assessment revealed a 28% low and an 18% high risk of bias. CONCLUSION: Intra-articular CS injections induced a wide range of results on OA joints in experimental animal models, from disease-modifying and positive effects on pain and joint function at short-term evaluation to the lack of benefit or even negative effects. This underlines the need to identify more specific indications and treatment modalities to avoid possible detrimental effects while maximising the anti-inflammatory properties and the benefits of intra-articular CS in OA joints. LEVEL OF EVIDENCE: Level II.

Low-density cultured cartilage cells expanded in platelet lysate present distinct features to develop an innovative clinical treatment for diffuse cartilage lesions.

PURPOSE: Chondrocyte-based cell therapies are effective for the treatment of chondral lesions, but remain poorly indicated for diffuse lesions in the context of early osteoarthritis (OA). The aim of this study was to develop a protocol to obtain chondroprogenitor cells suitable for the treatment of diffuse chondral lesions within early OA. METHODS: Cartilage cells were expanded at low density in human platelet lysate (hPL). A test was performed to exclude senescence. The expression of surface cluster of differentiation 146, cluster of differentiation 166, major histocompatibility complex (MHC)-I and MHC-II and of genes of interest were evaluated, as well as the trophic potential of these cells, by the assessment of lubricin and matrix production. The immunomodulatory potential was assessed through their co-culture with macrophages. RESULTS: Cartilage cells expanded at low density in hPL showed higher proliferation rate than standard-density cells, no replicative senescence, low immunogenicity and expression of lubricin. Moreover, they presented an increased expression of chondrogenic and antihypertrophic markers, as well as a superior matrix deposition if compared to cells cultured at standard density. Cartilage cells induced on macrophages an upregulation of CD206, although a higher increase of CD163 expression was observed in the presence of low-density cells. CONCLUSIONS: These findings lay the grounds to explore the clinical usefulness of low-density cultured cartilage cells to treat diffuse lesions in early OA joints for both autologous and allogenic use. LEVEL OF EVIDENCE: Not applicable.

The use of injectable orthobiologics for knee osteoarthritis: A European ESSKA-ORBIT consensus. Part 1-Blood-derived products (platelet-rich plasma).

PURPOSE: The aim of this European Society of Sports Traumatology, Knee Surgery and Arthroscopy (ESSKA) consensus is to provide recommendations based on evidence and expert opinion to improve indications, decision-making and administration-related aspects when using blood-derived orthobiologics (for simplicity indicated as PRP-platelet-rich plasma-with PRP being the most common product) for the management of knee osteoarthritis (OA). METHODS: Leading European expert clinicians and scientists were divided into a steering group, a rating group and a peer review group. The steering group prepared 28 question-statement sets divided into three sections: PRP rationale and indications, PRP preparation and characterisation and PRP protocol. The quality of the statements received grades of recommendation ranging from A (high-level scientific support) to B (scientific presumption), C (low-level scientific support) or D (expert opinion). The question-statement sets were then evaluated by the rating group, and the statements scored from 1 to 9 based on their degree of agreement with the statements produced by the steering group. Once a general consensus was reached between the steering and rating groups, the document was submitted to the peer review group who evaluated the geographic adaptability and approved the document. A final combined meeting of all the members of the consensus was held to produce the official document. RESULTS: The literature review on the use of blood-derived products for knee OA revealed that 9 of 28 questions/statements had the support of high-level scientific literature, while the other 19 were supported by a medium-low scientific quality. Three of the 28 recommendations were grade A recommendations: (1) There is enough preclinical and clinical evidence to support the use of PRP in knee OA. This recommendation was considered appropriate with a strong agreement (mean: 8). (2) Clinical evidence has shown the effectiveness of PRP in patients for mild to moderate degrees of knee OA (KL ≤ 3). This recommendation was considered appropriate with a strong agreement (mean: 8.1). (3) PRP injections have been shown to provide a longer effect in comparison to the short-term effect of CS injections. They also seem to provide a safer use profile with less potential related complications. This recommendation was considered appropriate with a very strong agreement (mean: 8.7). Six statements were grade B recommendations, 7 were grade C and 12 were grade D. The mean rating score was 8.2 ± 0.3. CONCLUSIONS: The consensus group reached a high level of agreement on all the questions/statements despite the lack of clear evidence for some questions. According to the results from this consensus group, given the large body of existing literature and expert opinions, PRP was regarded as a valid treatment option for knee OA and as a possible first-line injectable treatment option for nonoperative management of knee OA, mainly for KL grades 1-3. LEVEL OF EVIDENCE: Level II.

Platelet-rich plasma injections for the management of knee osteoarthritis: The ESSKA-ICRS consensus. Recommendations using the RAND/UCLA appropriateness method for different clinical scenarios.

PURPOSE: The aim of this consensus was to develop evidence- and expert-based patient-focused recommendations on the appropriateness of intra-articular platelet-rich plasma (PRP) injections in different clinical scenarios of patients with knee osteoarthritis (OA). METHODS: The RAND/UCLA Appropriateness Method was used by the European Society of Sports Traumatology, Knee Surgery, and Arthroscopy (ESSKA), as well as the International Cartilage Regeneration and Joint Preservation Society (ICRS) to reach a consensus and produce recommendations for specific patient categories combining best available scientific evidence with the collective judgement of a panel of experts. RESULTS: Scenarios were defined based on first treatment vs first injective treatment vs second injective treatment, age (<50/50-65/66-80/>80), tibiofemoral vs patellofemoral involvement, OA level (Kellgren-Lawrence/KL 0-I/II-III/IV), and joint effusion (dry knee, minor-mild or major effusion). Out of 216 scenarios, in 84 (38.9%) the indication was considered appropriate, in 9 (4.2%) inappropriate and in 123 (56.9%) uncertain. The parameters associated with the highest consensus were PRP use after failed injective treatments (62.5%), followed by PRP after failed conservative treatments and KL 0-III scenarios (58.3%), while the highest uncertainty was found for PRP use as first treatment and KL IV OA (91.7% and 87.5% of uncertain scenarios, respectively). CONCLUSION: This ESSKA-ICRS consensus established recommendations on the appropriateness or inappropriateness of PRP injections for the treatment of knee OA, providing a useful reference for clinical practice. PRP injections are considered appropriate in patients aged ≤80 years with knee KL 0-III OA grade after failed conservative non-injective or injective treatments, while they are not considered appropriate as first treatment nor in KL IV OA grade. LEVEL OF EVIDENCE: Level I.

A high level of scientific evidence is available to guide treatment of primary shoulder stiffness: The SIAGASCOT consensus.

PURPOSE: Shoulder stiffness (SS) is a condition characterised by active and passive restricted glenohumeral range of motion, which can occur spontaneously in an idiopathic manner or be associated with a known underlying aetiology. Several treatment options are available and currently no consensus has been obtained on which treatment algorithm represents the best choice for the patient. Herein we present the results of a national consensus on the treatment of primary SS. METHODS: The project followed the modified Delphi consensus process, involving a steering, a rating and a peer-review group. Sixteen questions were generated and subsequently answered by the steering group after a thorough literature search. A rating group composed by professionals specialised in the diagnosis and treatment of shoulder pathologies rated the question-answer sets according to the scientific evidence and their clinical experience. RESULTS: Recommendations were rated with an average of 8.4 points out of maximum 9 points. None of the 16 answers received a rating of less than 8 and all the answers were considered as appropriate. The majority of responses were assessed as Grade A, signifying a substantial availability of scientific evidence to guide treatment and support recommendations encompassing diagnostics, physiotherapy, electrophysical agents, oral and injective medical therapies, as well as surgical interventions for primary SS. CONCLUSIONS: A consensus regarding the conservative and surgical treatment of primary SS could be achieved at a national level. This consensus sets basis for evidence-based clinical practice in the management of primary SS and can serve as a model for similar initiatives and adaptable guidelines in other European countries and potentially on a global scale. LEVEL OF EVIDENCE: Level I.

Donor Sites and Harvesting Techniques Affect miRNA Cargos of Extracellular Vesicles Released by Human Adipose-Derived Mesenchymal Stromal Cells.

Osteoarthritis (OA) is a degenerative joint disorder characterized by the progressive deterioration of articular cartilage driven and sustained by catabolic and inflammatory processes that lead to pain and functional impairment. Adipose-derived stem cells (ASCs) have emerged as a promising therapeutic strategy for OA due to their regenerative potential, which mainly relies on the adaptive release of paracrine molecules that are soluble or encapsulated in extracellular vesicles (EVs). The biological effects of EVs specifically depend on their cargo; in particular, microRNAs (miRNAs) can specifically modulate target cell function through gene expression regulation. This study aimed to investigate the impact of collection site (abdominal vs. peri-trochanteric adipose tissue) and collection method (surgical excision vs. lipoaspiration) on the miRNAs profile in ASC-derived EVs and their potential implications for OA therapy. EV-miRNA cargo profiles from ASCs of different origins were compared. An extensive bioinformatics search through experimentally validated and OA-related targets, pathways, and tissues was conducted. Several miRNAs involved in the restoration of cartilage homeostasis and in immunomodulation were identified in all ASC types. However, EV-miRNA expression profiles were affected by both the tissue-harvesting site and procedure, leading to peculiar characteristics for each type. Our results suggest that adipose-tissue-harvesting techniques and the anatomical site of origin influence the therapeutic efficacy of ASC-EVs for tissue-specific regenerative therapies in OA, which warrants further investigation.

Stable Housekeeping Genes in Bone Marrow, Adipose Tissue, and Amniotic Membrane-Derived Mesenchymal Stromal Cells for Orthopedic Regenerative Medicine Approaches.

The therapeutic effect of mesenchymal stromal cells (MSCs) has been described for a variety of disorders, including those affecting musculoskeletal tissues. In this context, the literature reports several data about the regenerative effectiveness of MSCs derived from bone marrow, adipose tissue, and an amniotic membrane (BMSCs, ASCs, and hAMSCs, respectively), either when expanded or when acting as clinical-grade biologic pillars of products used at the point of care. To date, there is no evidence about the superiority of one source over the others from a clinical perspective. Therefore, a reliable characterization of the tissue-specific MSC types is mandatory to identify the most effective treatment, especially when tailored to the target disease. Because molecular characterization is a crucial parameter for cell definition, the need for reliable normalizers as housekeeping genes (HKGs) is essential. In this report, the stability levels of five commonly used HKGs (ACTB, EF1A, GAPDH, RPLP0, and TBP) were sifted into BMSCs, ASCs, and hAMSCs. Adult and fetal/neonatal MSCs showed opposite HKG stability rankings. Moreover, by analyzing MSC types side-by-side, comparison-specific HKGs emerged. The effect of less performant HKG normalization was also demonstrated in genes coding for factors potentially involved in and predicting MSC therapeutic activity for osteoarthritis as a model musculoskeletal disorder, where the choice of the most appropriate normalizer had a higher impact on the donors rather than cell populations when compared side-by-side. In conclusion, this work confirms HKG source-specificity for MSCs and suggests the need for cell-type specific normalizers for cell source or condition-tailored gene expression studies.

Effects of sleep disturbances and circadian rhythms modifications on cognition in breast cancer women before and after adjuvant chemotherapy: the ICANSLEEP-1 protocol.

BACKGROUND: Many patients treated for breast cancer (BC) complain about cognitive difficulties affecting their daily lives. Recently, sleep disturbances and circadian rhythm disruptions have been brought to the fore as potential contributors to cognitive difficulties in patients with BC. Yet, studies on these factors as well as their neural correlates are scarce. The purpose of the ICANSLEEP-1 (Impact of SLEEP disturbances in CANcer) study is to characterize sleep using polysomnography and its relationship with the evolution of cognitive functioning at both the behavioral and the neuroanatomical levels across treatment in BC patients treated or not with adjuvant chemotherapy. METHODS: ICANSLEEP-1 is a longitudinal study including BC patients treated with adjuvant chemotherapy (n = 25) or not treated with adjuvant chemotherapy (n = 25) and healthy controls with no history of BC (n = 25) matched for age (45-65 years old) and education level. The evaluations will take place within 6 weeks after inclusion, before the initiation of chemotherapy (for BC patients who are candidates for chemotherapy) or before the first fraction of radiotherapy (for BC patients with no indication for chemotherapy) and 6 months later (corresponding to 2 weeks after the end of chemotherapy). Episodic memory, executive functions, psychological factors, and quality of life will be assessed with validated neuropsychological tests and self-questionnaires. Sleep quantity and quality will be assessed with polysomnography and circadian rhythms with both actigraphy and saliva cortisol. Grey and white matter volumes, as well as white matter microstructural integrity, will be compared across time between patients and controls and will serve to further investigate the relationship between sleep disturbances and cognitive decline. DISCUSSION: Our results will help patients and clinicians to better understand sleep disturbances in BC and their relationship with cognitive functioning across treatment. This will aid the identification of more appropriate sleep therapeutic approaches adapted to BC patients. Improving sleep in BC would eventually help limit cognitive deficits and thus improve quality of life during and after treatments. TRIAL REGISTRATION: NCT05414357, registered June 10, 2022. PROTOCOL VERSION: Version 1.2 dated March 23, 2022.

Treatment with CR500® improves algofunctional scores in patients with knee osteoarthritis: a post-market confirmatory interventional, single arm clinical investigation.

BACKGROUND: Knee osteoarthritis (OA) is a progressive and degenerative condition. Several pharmacological and non-pharmacological treatments are able to improve the OA symptoms and the structural characteristics of the affected joints. Among these, infiltrative therapy with hyaluronic acid (HA) is the most used and consolidated procedure for the pain management. The addition of skin conditioning peptides to HA promotes the cartilage remodeling processes and a better permeation of the HA-based gel containing a peptide mixture, CR500®. Furthermore, the topic route of administration is convenient over the routinely used intra-articular injective procedures. In this study, the effectiveness of CR500® was evaluated in terms of improvement of the algo-functional symptoms related to unilateral knee OA. METHODS: 38 mild and moderate OA patients were enrolled at a screening visit (V-1), treated at baseline visit (V1), and then continued the topical application of CR500® twice a week for 4 weeks, and followed-up for 3 visits (V2-V4) from week 2 to 4. Lequesne Knee Index (LKI) and Knee injury and Osteoarthritis Outcome Score (KOOS) were collected. Synovial fluid was collected and used for the quantification of neoepitope of type II collagen (C2C), C-terminal telopeptide of type II collagen (CTX-II), type II collagen propeptide (CPII), tumor necrosis factor alpha (TNFα) and HA. The expression of CD11c and CD206 was evaluated on cell pellets. RESULTS: Three patients were excluded, thus 35 patients were included in the analysis. The treatment with CR500® was safe and well tolerated, with 7.9% patients had mild adverse events, not related to the device. The LKI total score showed a significant decrease from V1 to V4. KOOS score also showed a significant improvement of patient condition at V2, V3 and V4 in comparison with V1 for all subscales, except for KOOS sport subscale which improved only from V3. At V1 a negative correlation among KOOS pain subscale values and C2C, CPII and TNFα levels was observed, as well as a positive correlation between KOOS pain subscale and CD11c/CD206 ratio. CONCLUSION: CR500® is safe and appear to be effective in improving pain and function in OA patients during the 4 weeks of treatment. TRIAL REGISTRATION: ClinicalTrials.gov ID: NCT05661162. This trial was registered on 22/12/2022.

Cell-based therapies have disease-modifying effects on osteoarthritis in animal models. A systematic review by the ESSKA Orthobiologic Initiative. Part 2: bone marrow-derived cell-based injectable therapies.

PURPOSE: Aim of this systematic review was to determine if bone marrow-derived cell-based injectable therapies induce disease-modifying effects in joints affected by osteoarthritis (OA) in animal models. METHODS: A systematic review was performed on three electronic databases (PubMed, Web of Science, Embase) according to PRISMA guidelines. A synthesis of the results was performed investigating disease-modifying effects in preclinical animal studies comparing injectable bone marrow-derived products with OA controls or other products, different formulations or injection intervals, and the combination with other products. The risk of bias was assessed according to the SYRCLE's tool. RESULTS: Fifty-three studies were included (1819 animals) with an increasing publication trend over time. Expanded cells were used in 48 studies, point-of-care products in 3 studies, and both approaches were investigated in 2 studies. Among the 47 studies presenting results on the disease-modifying effects, 40 studies (85%) reported better results with bone marrow-derived products compared to OA controls, with positive findings evident in 14 out of 20 studies (70%) in macroscopic assessment, in 30 out of 41 studies (73%) in histological assessment, and in 10 out of 13 studies (77%) in immunohistochemical evaluations. Clinical evaluations showed positive results in 7 studies out of 9 (78%), positive imaging results in 11 studies out of 17 (65%), and positive biomarker results in 5 studies out of 10 (50%). While 36 out of 46 studies (78%) reported positive results at the cartilage level, only 3 out of 10 studies (30%) could detect positive changes at the synovial level. The risk of bias was low in 42% of items, unclear in 50%, and high in 8%. CONCLUSION: This systematic review of preclinical studies demonstrated that intra-articular injections of bone marrow-derived products can induce disease-modifying effects in the treatment of OA, slowing down the progression of cartilage damage with benefits at macroscopic, histological, and immunohistochemical levels. Positive results have been also observed in terms of clinical and imaging findings, as well as in the modulation of inflammatory and cartilage biomarkers, while poor effects have been described on the synovial membrane. These findings are important to understand the potential of bone marrow-derived products and to guide further research to optimise their use in the clinical practice. LEVEL OF EVIDENCE: II.

Bone marrow aspirate concentrate quality is affected by age and harvest site.

PURPOSE: To compare the number and properties of bone marrow stromal cells (BMSCs) collected from bone marrow aspirate concentrate (BMAC) obtained from different harvest sites and from patients of different ages. METHODS: BMAC was obtained from two groups of patients based on age (n = 10 per group): 19.0 ± 2.7 years for the younger and 56.8 ± 12.5 for the older group. In the latter, BMAC was obtained from both iliac crest and proximal tibia for a donor-matched analysis. Mononucleated cell count and CFU-F assay were performed, together with phenotype characterization of BMSCs from iliac crest and proximal tibia, the study of chondrogenic and osteogenic differentiation capacity, histological staining and spectrophotometric quantification, and the analysis of mRNAs expression. RESULTS: Cells derived from iliac crest and proximal tibia showed the same phenotypic pattern at flow cytometry, as well as similar chondrogenic and osteogenic potential. However, a significantly higher number of mononuclear cells per ml was observed in younger patients (3.8 ± 1.8 × 107) compared to older patients (1.2 ± 0.8 × 107) (p < 0.0005). The latter yield, obtained from the iliac crest, was significantly higher than resulting from the BMAC harvested from the proximal tibia in the same group of patients (0.3 ± 0.2 × 107, p < 0.0005). This result was confirmed by the CFU-F analysis at day 10 (15.9 ± 19.4 vs 0.6 ± 1.0, p = 0.001) and day-20 (21.7 ± 23.0 vs 2.9 ± 4.2, p = 0.006). CONCLUSION: Harvest site and age can affect the quality of BMAC. BMSCs obtained from iliac crest and proximal tibia present comparable mesenchymal markers expression as well as osteogenic and chondrogenic differentiation potential, but iliac crest BMAC presents a four times higher number of mononucleated cells with significantly higher clonogenic capacity compared to the tibia. BMAC of younger patients also had a three-time higher number of mononucleated cells. The identification of BMAC characteristics could help to optimize its preparation and to identify the most suitable indications for this orthobiologic treatment in the clinical practice.

Cell-based therapies have disease-modifying effects on osteoarthritis in animal models. A systematic review by the ESSKA Orthobiologic Initiative. Part 1: adipose tissue-derived cell-based injectable therapies.

PURPOSE: The aim of this systematic review was to determine if adipose tissue-derived cell-based injectable therapies can induce disease-modifying effects in joints affected by osteoarthritis (OA). METHODS: A systematic review was performed on three electronic databases (PubMed, Web of Science, Embase) according to PRISMA guidelines. A synthesis of the results was performed investigating disease-modifying effects in preclinical studies comparing injectable adipose-derived products with OA controls or other products, different formulations or injection intervals, and the combination with other products. The risk of bias was assessed according to the SYRCLE's tool. RESULTS: Seventy-one studies were included (2,086 animals) with an increasing publication trend over time. Expanded cells were used in 65 studies, 3 studies applied point of care products, and 3 studies investigated both approaches. Overall, 48 out of 51 studies (94%) reported better results with adipose-derived products compared to OA controls, with positive findings in 17 out of 20 studies (85%) in macroscopic, in 37 out of 40 studies (93%) in histological, and in 22 out of 23 studies (96%)  in immunohistochemical evaluations. Clinical and biomarker evaluations showed positive results in 14 studies out of 18 (78%) and 12 studies out of 14 (86%), while only 9 studies out of 17 (53%) of the imaging evaluations were able to detect differences versus controls. The risk of bias was low in 38% of items, unclear in 51%, and high in (11%). CONCLUSION: The current preclinical models document consistent evidence of disease-modifying effects of adipose-derived cell-based therapies for the treatment of OA. The high heterogeneity of the published studies highlights the need for further targeted research to provide recommendations on the optimal methodologies for a more effective application of these injective therapies for the treatment of OA in clinical practice. LEVEL OF EVIDENCE: II.

Biological intra-articular augmentation for osteotomy in knee osteoarthritis: strategies and results : A systematic review of the literature from the ESSKA Orthobiologics Initiative.

PURPOSE: To assess whether there is evidence supporting the use of augmentation strategies, either cartilage surgical procedures or injective orthobiologic options, to improve the results of osteotomies in knees with osteoarthritis (OA). METHODS: A systematic review of the literature was performed on the PubMed, Web of Science and the Cochrane databases in January 2023 on osteotomies around the knee associated with augmentation strategies (either cartilage surgical procedures or injective orthobiologic options), reporting clinical, radiological, or second-look/histological outcomes at any follow-up. The methodological quality of the included studies was assessed with the Coleman Methodology Score (CMS). RESULTS: Out of the 7650 records identified from the databases, 42 articles were included for a total of 3580 patients and 3609 knees treated; 33 articles focused on surgical treatments and 9 on injective treatments performed in association with knee osteotomy. Out of the 17 comparative studies with surgical augmentation, only 1 showed a significant clinical benefit of an augmentation procedure with a regenerative approach. Overall, other studies showed no differences with reparative techniques and even detrimental outcomes with microfractures. Regarding injective procedures, viscosupplementation showed no improvement, while the use of platelet-rich plasma or cell-based products derived from both bone marrow and adipose tissue showed overall positive tissue changes which translated into a clinical benefit. The mean modified CMS score was 60.0 ± 12.1. CONCLUSION: There is no evidence to support the effectiveness of cartilage surgical treatments combined with osteotomies in terms of pain relief and functional recovery of patients affected by OA in misaligned joints. Orthobiologic injective treatments targeting the whole joint environment showed promising findings. However, overall the available literature presents a limited quality with only few heterogeneous studies investigating each treatment option. This ORBIT systematic analysis will help surgeons to choose their therapeutic strategy according to the available evidence, and to plan further and better studies to optimize biologic intra-articular osteotomy augmentation. LEVEL OF EVIDENCE: Level IV.

Micro-fragmented adipose tissue (mFAT) associated with arthroscopic debridement provides functional improvement in knee osteoarthritis: a randomized controlled trial.

PURPOSE: Current conservative treatments for knee OA provide limited benefits, with symptoms relief for a short amount of time. Regenerative medicine approaches such as the use of microfragmented adipose tissue (mFAT) showed promising results in terms of durable effects and the possibility to enhance tissue healing and counteract the progression of the pathology. Nevertheless, up to today, the large part of clinical data about mFAT use refers to uncontrolled studies, especially in the surgical setting. The purpose of this study was to evaluate the effectiveness of mFAT applied in association with arthroscopic debridement (AD) for the treatment of knee OA, in terms of symptoms relief and tissue healing. METHODS: This study is a prospective, randomized controlled clinical trial. 78 patients affected by knee OA grade 3-4 according to KL classification were randomly assigned to AD or AD + mFAT treatment groups. Clinical, radiological and serological assessments were performed at 6 months after treatment. Additional clinical evaluation was performed at the end of the study with an average follow-up of 26.1 ± 9.5 months. VAS, KOOS, WOMAC and SF-12 were also collected at both timepoints, KSS only at 6 months. RESULTS: Treatment with AD + mFAT improved functional scores at both 6 months (KOOS-PS: + 11.7 ± 20.2 vs + 24.4 ± 22.5, in AD and AD + mFAT, respectively, p = 0.024; KSS: + 14.9 ± 15.9 vs + 24.8 ± 23.5, in AD and AD + mFAT, respectively, p = 0.046) and 24-month follow-ups (KOOS-PS Functional subscale: - 2.0 ± 3.5 vs - 4.7 ± 4.2, in AD and AD + mFAT, respectively, p = 0.012). Lower T2-mapping scores were obtained in AD + mFAT-treated group in medial and lateral condyle compartments (p < 0.001). Slight increase was observed in the levels of a serum biomarker of cartilage deposition (PIIINP) in both groups at 6-month follow-up (p = 0.037). CONCLUSION: mFAT improves functional outcome and MRI appearance when used in association with AD, therefore supporting its use in the treatment of knee OA in an arthroscopic setting.

Inflammatory Treatment Used to Mimic Osteoarthritis and Patients' Synovial Fluid Have Divergent Molecular Impact on Chondrocytes In Vitro.

Osteoarthritis (OA) is a chronic disease characterized by joint tissue disruption and inflammation with a paucity of therapeutic options. Chondrocyte in vitro models are commonly used as the first step in evaluating new approaches and rely on the stimulation of an OA-like phenotype with inflammation often the method of choice. Inflammatory priming is frequently based on cytokines used at concentrations very far from the reality in the patients' synovial fluid (SF). The aim of this work was to compare the transcriptional response of chondrocytes to different inflammatory conditions: the high levels of IL1ß that are used for standardized inflammation protocols, OA-SF, IL1ß, IL6 and IFNγ at SF-like concentrations both individually and simultaneously to mimic a simplified "in vitro" SF. Both high IL1ß and OA-SF strongly influenced chondrocytes, while SF-like concentrations of cytokines gave weak (IL1ß alone or in combination) or no (IL6 and IFNγ alone) outcomes. Chondrocytes under the two most powerful polarizing conditions had a clearly distinct fingerprint, with only a shared albeit molecularly divergent effect on ECM stability, with IL1ß mainly acting on ECM degrading enzymes and OA-SF accounting for a higher turnover in favor of fibrous collagens. Moreover, OA-SF did not induce the inflammatory response observed with IL1ß. In conclusion, although partially similar in the endpoint phenotype, this work intends to encourage reflection on the robustness of inflammation-based in vitro OA models for molecular studies on chondrocytes.

Italian version of the anterior cruciate ligament-return to sport after injury scale (IT ACL-RSI): translation, cross-cultural adaptation, validation and ability to predict the return to sport at medium-term follow-up in a population of sport patients.

PURPOSE: The timing of psychological and physical recovery after anterior cruciate ligament reconstruction represents an open issue in current orthopedic practice. Several tools have been developed to evaluate these factors, with the most recent being represented by the anterior cruciate ligament (ACL) return to sport injury scale (ACL-RSI). The aims of this study were to provide a validated Italian translation of the ACL-RSI in a population of sport patients, and to identify a possible correlation of the ACL-RSI score with the return to sport (RTS) time and the level of sport participation in comparison to the pre-injury one. METHODS: The Italian translation and cultural adaptation of the scale were completed using a using the "translation-back translation" method. A total of 130 patients were enrolled and completed the study questionnaires 6 months after ACL reconstruction. Randomly, 65 of them were re-tested for the ACL-RSI within 2 weeks. The internal consistency, reliability, feasibility, and construct validity of the Italian version of ACL-RSI were assessed and compared to Italian version of the KOOS, the Lysholm Score, the AKPS and the IKDC subjective score. Responsiveness was tested comparing patients returning to sport at 6 and 12 months. The Tegner activity scale was collected at baseline, 6 and 12 months to identify the level of activity after return to sport, in relation to the ACL-RSI score. RESULTS: The Italian adaptation of the ACL-RSI demonstrated excellent internal consistency (Cronbach's alpha = 0.953), reliability (test-retest ICC = 0.916) and feasibility, with no ceiling or floor effect. Construct validity was confirmed by the moderate to strong correlation with all the other scales (p < 0.0001). Slight and non-significant higher ACL-RSI score was shown by patients returned to sport at 6 or 12 months after surgery. Nevertheless, the ACL-RSI score at 6 months was significantly different between patients who returned and those who did not returned to the same level of sport activity 12 months after the procedure. CONCLUSIONS: This study demonstrated that the Italian ACL-RSI is a reliable tool for evaluating the psychological readiness for return to sports of athletes who underwent ACL reconstruction, especially when collected at the end of the rehabilitation process. Since the IT ACL-RSI used in this study is a faithful translation of the original English version, this finding can be generalized to other cultural contexts and languages too. LEVEL OF EVIDENCE: Level II.

Sufficient MRI graft structural integrity at 9 months after anterior cruciate ligament reconstruction with hamstring tendon autograft.

PURPOSE: To assess the MRI-measured structural integrity of hamstring autograft at 9 months after anterior cruciate ligament reconstruction (ACLR) surgery. METHODS: A prospective cohort study was conducted including 34 patients (mean age 36.8 ± 11.8 years; 73.5% males) who underwent ACLR with hamstring autograft. In addition, 36 patients (mean age 39.2 ± 10.5 years; 69.4% males) without any ACL injury were also reviewed and served as control group. The primary outcome used for examining graft maturity at 9 months after ACLR was the MRI-based signal-to-noise quotient (SNQ) of reconstructed ACL. SNQ values were stratified into 3 different categories: excellent: < 0.1; good: ≥ 0.1 and ≤ 0.19; fair: ≥ 0.2. The KT-1000 knee arthrometer was used to measure the side-to-side difference in the anterior tibial translation between the ACLR knee and the contralateral healthy knee in the ACLR group. RESULTS: Reconstructed ACLs were found with a mean SNQ of 0.078 ± 0.061, while almost all ACL-reconstructed patients (97%; 33 out of 34) were found with excellent or good SNQ values (< 0.019). The mean KT-1000 in the ACLR group was 0.071 mm ± 0.926 mm, while there were no patients in the ACLR cohort with a KT-1000 value > 3 mm. The mean 9-months MRI-based SNQ of ACLR group was significantly higher compared to the mean MRI-based SNQ of the control group (p < 0.001). Multiple regression analysis showed no correlation between SNQ and age, gender, time from injury to ACLR, graft size, or simultaneous treatment of additional intra-articular knee lesions. CONCLUSIONS: In this cohort of 34 ACL-reconstructed patients, 97% of hamstring tendon autografts demonstrated excellent/good MRI signal intensity and excellent functional results (KT-1000 < 3 mm) at 9 months after surgery. Based on this finding, it is suggested that return to sports after ACLR with hamstring autograft can be considered safe at 9 months post surgery. Furthermore, while structural integrity of the graft has been achieved at this time point, statistical differences found in SNQ values of ACL-reconstructed patients compared to ACLs of healthy individuals highlight the continuing process of graft maturation and remodelling. LEVEL OF EVIDENCE: Level III.

Different timing in allograft and autograft maturation after primary anterior cruciate ligament reconstruction does not influence the clinical outcome at mid-long-term follow-up.

PURPOSE: The use of allografts in primary anterior cruciate ligament reconstruction (ACLR) is increasing although they are still supposed to be associated to greater risk of re-rupture due to a slower and less efficient graft maturation. The aim of this prospective randomized controlled study was to compare the graft maturation after ACLR with allograft and autograft by MRI at 6- and 12-month follow-up and integrate these data with the functional and clinical results observed at 6-, 12- and 60-month follow-up. METHODS: Fifty patients with indication to primary ACLR were randomly and equally divided into hamstring autograft or allograft tendon groups. The graft maturation was measured at 6- and 12-month follow-up by the SNQ score and other radiological parameters on MRI scans. Clinical and functional recovery was evaluated by Lysholm score, Visual Analogues Scale, Tegner activity scale and modified Cincinnati knee rating system at 6, 12 and 60 months after surgery to estimate the predictive value of the radiological parameters for clinical outcomes. Return-to-sport (ACL-RSI) was measured 60 months after surgery. RESULTS: Three patients had retear of the neo-ligament (two from Auto group and one from Allo group). All the clinical/functional parameters significantly improved over time, with no statistically significant difference between the groups. At 6 months, the SNQ value was significantly higher in the Auto than in the Allo group (12.9 vs 7.9, p = 0.038), but at 12 months they were comparable (9.8 vs 10.4). The 6-month SNQ values did not correlate with the clinical scores, whereas the 12-month SNQ values significantly correlated with the Cincinnati score, Lysholm score and Tegner activity scale collected at 60-month follow-up. CONCLUSION: No clinical or functional differences have been found between the two treatment groups, supporting the suitability of using allograft in primary ACLR, when available. The results at MRI scans showed a different graft maturation trend in the two groups, with allografts being more reactive in the first 6 months. MRI together with the subjective evaluation allows to evaluate objectively the status of the neo-ligamentization process and therefore helps the surgeon to dictate the individual time for return-to-sport. LEVEL OF EVIDENCE: Level I.