During aging, tendons demonstrate substantial disruptions in homeostasis, leading to impairments in structure-function. Impaired tendon function contributes to substantial declines quality of life during aging. Aged tendons are more likely to undergo spontaneous rupture, and the healing response following injury is impaired in aged tendons. Thus, there is a need to develop strategies to maintain tendon homeostasis and healing capacity through the lifespan. Tendon cell density sharply declines by ∼12 months of age in mice, and this low cell density is retained in geriatric tendons. Our data suggests that this decline in cellularity initiates a degenerative cascade due to insufficient production of the extracellular matrix (ECM) components needed to maintain tendon homeostasis. Thus, preventing this decline in tendon cellularity has great potential for maintaining tendon health. Single cell RNA sequencing analysis identifies two changes in the aged tendon cell environment. First, aged tendons primarily lose tenocytes that are associated with ECM biosynthesis functions. Second, the tenocytes that remain in aged tendons have disruptions in proteostasis and an increased pro-inflammatory phenotype, with these changes collectively termed ‘programmatic skewing'. To determine which of these changes drives homeostatic disruption, we developed a model of tenocyte depletion in young animals. This model decreases tendon cellularity to that of an aged tendon, including decreased biosynthetic tenocyte function, while age-related programmatic skewing is absent. Loss of biosynthetic tenocyte function in young tendons was sufficient to induce homeostatic disruption comparable to natural aging, including deficits in ECM organization, composition, and material quality, suggesting loss biosynthetic tenocytes as an initiator of tendon degeneration. In contrast, our data suggest that programmatic skewing underpins impaired healing in aged tendons. Indeed, despite similar declines in the tenocyte environment, middle-aged and young-depleted tendons mount a physiological healing response characterized by robust ECM synthesis and remodeling, while aged tendons heal with insufficient ECM

Matrix-bound vesicles (MBVs) are embedded within osteoid and function as the site of initial mineral formation. However, they remain insufficiently characterised in terms of biogenesis, composition and function while their relationship with secreted culture medium EVs (sEVs) such as exosomes remains debated. We aimed to define the biogenesis and pro-mineralisation capacity of MBVs and sEVs to understand their potential in regenerative orthopaedics. sEVs and MBVs isolated from conditioned medium (differential ultracentrifugation) and ECM (collagenase digestion and differential ultracentrifugation) of mineralising MC3T3 pre-osteoblast and human bone marrow MSC cultures were characterised by nanoparticle tracking analysis, western blotting, nano-flow cytometry, super resolution microscopy (ONI) and TEM. Immunoprecipitated populations positive for alkaline phosphatase (ALP), a putative marker of mineralisation capacity, were also characterised. Collagen binding efficiency was evaluated using MemGlow staining. Results reported were comparative across both cell lines. Western blots indicated MBV fractions were positive for markers of endosomal biogenesis (CD9, CD81, ALIX, TSG101) and pro-mineralising proteins (ALP, Pit1, Annexin II, Annexin V), with Annexin V and CD9 present in immunoprecipitated ALP-positive fractions. MBVs were significantly larger than sEVs (p<0.05) and contained a higher amount of ALP (p<0.05) with a significant increase from day 7 to day 14 of cellular mineralisation (p<0.05). This mirrored the pattern of electron-dense vesicles seen via TEM. Super resolution single vesicle analysis revealed for the first-time co-expression of ALP with markers of endosomal biogenesis (CD9, CD63, CD81, ALIX) and Annexin II in both vesicle types, with higher co-expression percentage in MBVs than sEVs. MBVs also exhibited preferential collagen binding. Advanced imaging methods demonstrated that contrary to opinions in the field, MBVs appear to possess exosomal markers and may arise via endosomal biogenesis. However, it was evident that a higher proportion of MBVs possessed machinery to induce mineralisation and were enriched in mineral-dense material

To investigate temporal changes in synovial lymphatic system (SLS) drainage function after Anterior cruciate ligament (ACL) injury, a non-invasive ACL rupture model was used to induce the PTOA phenotype without altering the SLS structure. We have created a non-invasive ACL rupture model in the right knee (single overload impact) of 12- week-old C57bl/6 male mice to mimic the ACL rupture-induced PTOA development. 70 kDa-TxRedDextran were injected into the right knee of the mice at 0, 1, 2, and 4 wks post modeling (n=5/group), and the fluorescence signal distribution and intensity were measured by the IVIS system at 1 and 6 hrs post-injection. After 24 hrs, the drainage lymph nodes and whole knee joint were harvested and subjected to ex vivo IVIS imaging and immunofluorescence detection respectively. Manual ACL rupture was induced by 12N overloaded force and validated by a front drawer test. Intraarticular clearance of TxRed-Dextran detected by the IVIS was significantly reduced at 1, and 2 wks at a level of 43% and 55% respectively but was not significantly different from baseline levels at 4 wks (89%). TxRed-Dextran signal in draining lymph nodes was significantly reduced at 1 week at the level of but not for 2 and 4 wks compared to baseline levels (week 1–29%, week 2–50%, week 4–94%). TxRed-Dextran particle was significantly enriched in the synovium at 1, 2 wks but was not significantly different from baseline levels at 4 wks rupture-post ACL rupture (Particle numbers: Sham Ctrl-34 ±14, week 1, 113 ± 17; week 2, 89 ± 13; week 4, 46 ± 18; mean ± SD). We observed the drainage function of SLS significantly decreased at 1 and 2 wks after the ACL rupture, and was slowly restored at 4 wks post-injury in a non-invasive ACL rupture model. Early impairment of SLS drainage function may lead to accumulation of inflammatory factors and promote PTOA progression

Nerve transfer is an emerging treatment to restore upper limb function in people with tetraplegia. The objective of this study is to examine if a flexible collage sheet (FCS) can act as epineurial-like substitute to promote nerve repair in nerve transfer. A preclinical study using FCS was conducted in a rat model of sciatic nerve transection. A prospective case series study of nerve transfer was conducted in patients with C5-C8 tetraplegia who received nerve transfer to restore upper limb function. Motor function in the upper limb was assessed pre-treatment, and at 6-,12-, and 24-months post-treatment. Macroscopic assessment in preclinical model showed nerve healing by FCS without encapsulation or adhesions. Microscopic examination revealed that a new, vascularised epineurium-like layer was observed at the FCS treatment sites, with no evidence of inflammatory reaction or nerve compression. Treatment with FCS resulted in well-organised nerve fibres with dense neurofilaments distal to the coaptation site. Axon counts performed proximal and distal to the coaptation site showed that 97% of proximal axon count of myelinated axons regenerated across the coaptation site after treatment with CND. In the proof of concept clinical study 17 nerve transfers were performed in five patients. Nerve transfers included procedures to restore triceps function (N=4), wrist/finger/thumb extension (N=6) and finger flexion (N=7). Functional motor recovery (MRC ≥3) was achieved in 76% and 88% of transfers at 12 and 24 months, respectively. The preclinical study showed that FCS mimics epineurium and enable to repair nerve resembled to normal nerve tissue. Clinical study showed that patients received nerve transfer with FCS experienced consistent and early return of motor function in target muscles. These results provide proof of concept evidence that CND functions as an epineurial substitute and is promising for use in nerve transfer surgery

Abstract. Objectives. Exploring the association of objective lower limb function pre and post total knee replacement (TKR). Methods. 3D gait analysis was performed on 28 non-pathological participants (NP) and 40 patients with advanced knee osteoarthritis (OA) before and approximately one year after TKR. For NP and OA patients pre/post-TKR, 12 waveforms on kinetic and kinematic variables of the operative side were chosen to perform data reduction through Principal Component (PC) Analysis. The Cardiff Classifier, a classification system based on Dempster-Shafer theory, was trained with the first 3 PCs of each variable. The 18 highest-ranking PCs classifying the biomechanical features of each participant as Belief in Healthy, Belief in OA (BOA) or Belief in Uncertainty were used to quantify biomechanical changes pre- to post-TKR. The correlation between patients’ BOA values (range: 0 to 1, 0 indicates null BOA and 1 high BOA) pre- and post-TKR was tested through Spearman's correlation coefficient. Wilcoxon matched-pair test (α<0.05) determined the significance of the change in BOA. Results. NP (57% women) had a mean age of 38 (SD=18.13), mean height and weight of 1.70 m (SD=0.09) and 68 Kg (SD=15.24), respectively. Their mean BOA was 0.05 (SD=0.08), in line with inclusion criteria. OA patients (38% women) had a mean age of 68.5 (SD=7.59), mean height and weight of 1.68 m (SD=0.10) and 92.6 Kg (SD=21.22), respectively. Their mean pre-TKR BOA was 0.81 (SD=0.18), falling to 0.64 (SD=0.26) post-TKR. The change in BOA (0.16, SD=0.19) pre- to post-TKR was significant (p=0.000). BOAs pre- and post-TKR were non-normally distributed therefore, a Spearman's rank-order correlation was run, revealing a positive, statistically significant (p=0.000), strong correlation (r. S. =0.657) between BOA pre- and post-TKR. Conclusions. This study found that worse objective pre-operative lower limb function in people with advanced knee OA was strongly correlated with poorer one-year post-TKR function (r. S. =0.657, p<0.01). Declaration of Interest. (b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project

Abstract. Objectives. Although hip replacement and resurfacing procedures both aim to restore mobility, improve joint function, and relieve pain, it is unclear how each differ in terms of gait mechanics and if they are affected by varying walking speeds. We compared limb symmetry and ground reaction force (GRF) profiles between bilateral total hip arthroplasty patients (THA), bilateral hip resurfacing arthroplasty patients (HRA), and healthy control participants (CON) during level-treadmill walking at different speeds. Methods. Bilateral THA and bilateral HRA patients (nTHA = 15; nHRA = 15; postoperative 12–18 months), and age-, mass-, and height-matched CON participants (nCON = 20) underwent gait analysis on an instrumented treadmill. Walking trials started at 4 km/h and increased in 0.5 km/h increments until top walking speed (TWS) was achieved. Gait symmetry index (SI = 0% for symmetry) was assessed between limbs during weight-acceptance, mid-stance and push-off phases of gait; and vertical GRFs were captured for the normalised stance phase using statistical parametric mapping (SPM; CI = 95%). Results. THA had a significantly lower TWS (6.51 ± 0.06 km/h) compared to HRA (7.09 ± 0.07 km/h, p = 0.01) and CON (7.15 km/h ± 0.06, p = 0.02). There were no SI differences between groups nor between walking speeds (SI < 5%). There were no GRF differences between groups at slower walking speeds (4.0–5.0 km/h). However, at 5.5 and 6 km/h, THA had lower GRF at the push-off phase (0.88 ± 0.09 N/BW), compared to HRA (1.06 ± 0.08 N/BW, p = 0.01) and CON (1.04 ± 0.02 N/BW, p = 0.01). Conclusions. The main finding was that HRA patients demonstrated restored gait function and similar walking profiles to CON participants at any speed. With a diverging gait profile, walking speeds over 5.5 km/h provided a functional challenge for THA patients

Abstract. Objectives. Exploring the relationship of gait function pre and post total knee replacement (TKR) in two groups of patients. Methods. Three-dimensional gait analysis was performed at Cardiff University, UK, and Karolinska University Hospital, Sweden, on 29 and 25 non-pathological (NP) volunteers, and 39 and 28 patients with end-stage knee osteoarthritis (OA), respectively. Patients were assessed pre and one-year post-TKR. Data reduction was performed via Principal Component (PC) analysis on twenty-four kinematic and kinetic waveforms in both NP and pre/post-TKR. Cardiff's and Karolinska's cohorts were analysed separately. The Cardiff Classifier, a classification system based on the Dempster-Shafer theory, was trained with the first 3 PCs of each variable for each cohort. The Classifier classifies each participant by assigning them a belief in NP, belief in OA (BOA) and belief in uncertainty, based on their biomechanical features. The correlation between patient's BOA values (range: 0–1, 0 indicates null BOA and 1 high BOA) pre and post-TKR was tested through Spearman's correlation coefficient in each cohort. The related-samples Wilcoxon signed-rank test (α=0.05) determined the significant changes in BOA in each cohort of patients. The Mann-Whitney U test (α=0.05) was run to explore differences between the patients’ cohorts. Results. There were no significant differences between patients’ cohorts in median age (p=0.096), height (p=0.673), weight (p=0.064) or KOOS sub-scores pre or post-TKR (p-value ranged 0.069 to 0.955) but Cardiff's patients had a significantly higher BMI (p=0.047). There was a significant, median decrease of 0.12 and 0.19 in the BOA pre to post TKR (p<0.001) in Cardiff's and Karolinska's patients, respectively. There was a statistically significant, strong positive correlation between the BOA pre and post-TKR (Cardiff:r. s. =0.706, p<0.001; Karolinska:r. s. =0.669, p<0.001). Conclusions. In two distinct cohorts of patients, having a more compromised gait function in end-stage knee OA was correlated with poorer gait function post-TKR

The knee joint has also a periarticular adipose tissue, which is known as Hoffa's fat pad (IPFP). IPFP has a dual function in the joint it reduces the concentration of Nitric Oxide, the release of glycosaminoglycans and the expression of MMP1 in the cartilage, but it also contains MSC and macrophages. Our hypothesis is that synovial fluid contains elements, not all of which are understood, which act as messengers and alter the “homeostasis” of the knee and the metabolism of all the cellular components of the joint, including the MSC of Hoffa's fat pad, thus making them another piece in the puzzle as far as OA of the knee is concerned. The IPFP of 37 patients with OA and 36 patients with ACL rupture were analyzed. Isolation, primary culture, and a functional and proteomic study of MSCs from IPFP were performed. Our results show that OA of the knee, in its more severe phases, also affects the MSC's of IPFP, which is a new actor in the OA degenerative process and which can contribute to the origin, onset and progression of the disease. A differential protein profile between OA and ACL patients were identified. Infrapatellar pad should be regarded as an adipose tissue with its own characteristics and it´s also able to produce and excrete important inflammatory mediators directly into the knee joint

To investigate changes in quadriceps and hamstrings muscle groups during sustained isokinetic knee flexion and extension. 125 paediatric participants (45 males and 80 females, mean age 14.2 years) were divided into two groups: participants with a confirmed ACL tear (ACLi, n = 64), and puberty- and activity-level matched control participants with no prior history of knee injuries (CON, n = 61). Participants completed a series of 44 repetitions of isokinetic knee flexion and extension at 90 deg/ sec using a Biodex dynamometer (Biodex Medical Systems Inc, Shirley, New York). Surface EMG sensors (Delsys Incorporated, Natick, MA) simultaneously recorded the quadriceps and hamstring activations. Muscle function was assessed as the change in quadriceps activation and extension torque were calculated using the percent difference between the mean of the first five trials, and the mean of the last five trials. ACLi participants had significantly higher percent change in quadriceps activation for both healthy and injured legs, in comparison to CON dominant leg. As such, the healthy leg of the ACLi participants is activating significantly more than their health matched controls, while also demonstrating reduced muscular endurance (less torque in later repetitions). Therefore, we conclude that the non-injured limb of the ACLi participant is not performing as a healthy limb. Since return to activity clearance following ACLi implies return to sport against age- and activity matched opponents, clearing young athletes based on the non-injured contralateral limb may put them at greater risk of reinjury

The hip's capsular ligaments (CL) passively restrain extreme range of motion (ROM) by wrapping around the native femoral head/neck, and protect against impingement and instability. We compared how CL function was affected by device (hip resurfacing arthroplasty, HRA; dual mobility total hip arthroplasty, DM-THA; and conventional THA, C-THA), and surgical approach (anterior and posterior), with and without CL surgical-repair. We hypothesized that CL function would only be preserved when native head-size (HRA/DM-THA) was restored. CL function was quantified on sixteen cadaveric hips, by measuring ROM by internally (IR) and externally rotating (ER) the hip in six functional positions, ranging from full extension with abduction to full flexion with adduction (squatting). Native ROM was compared to ROM after posterior capsulotomy (right hips) or anterior capsulotomy (left hips), and HRA, and C-THA and DM-THA, before and after CL repair. Independent of approach, ROM increased most following C-THA (max 62°), then DM-THA (max 40°), then HRA (max 19°), indicating later CL engagement and reduced biomechanical function with smaller head-size. Dislocations also occurred in squatting after C-THA and DM-THA. CL-repair following HRA restored ROM to the native hip (max 8°). CL-repair following DM-THA reduced ROM hypermobility in flexed positions only and prevented dislocation (max 36°). CL-repair following C-THA did not reduce ROM or prevent dislocation. For HRA and repair, native anatomy was preserved and ligament function was restored. For DM-THA with repair, ligament function depended on the movement of the mobile-bearing, with increased ROM in positions when ligaments could not wrap around head/neck. For C-THA, the reduced head-size resulted in inferior capsular mechanics in all positions as the ligaments remained slack, irrespective of repair. Choosing devices with anatomic head-sizes (HRA/DM-THA) with capsular repair may have greater effect than surgical approach to protect against instability in the early postoperative period

Anterior cruciate ligament reconstruction (ACLR) using a semitendinosus (ST) autograft, with or without gracilis (GR), results in donor muscle atrophy and varied tendon regeneration. The effects of harvesting these muscles on muscle moment arm and torque generating capacity have not been well described. This study aimed to determine between-limb differences (ACLR vs uninjured contralateral) in muscle moment arm and torque generating capacity across a full range of hip and knee motions.

A secondary analysis of magnetic resonance imaging was undertaken from 8 individuals with unilateral history of ST-GR ACLR with complete ST tendon regeneration. All hamstring muscles and ST tendons were manually segmented. Muscle length (cm), peak cross-sectional area (CSA) (cm²), and volume (cm³) were measured in ACLR and uninjured contralateral limbs. OpenSim was used to simulate and evaluate the mechanical consequences of changes in normalised moment arm (m) and torque generating capacity (N.m) between ACLR and uninjured contralateral limbs.

Compared to uninjured contralateral limbs, regenerated ST tendon re-insertion varied proximal (+) (mean = 0.66cm, maximum = 3.44cm, minimum = −2.17cm, range = 5.61cm) and posterior (+) (mean = 0.38cm maximum = 0.71cm, minimum = 0.02cm, range = 0.69cm) locations relative to native anatomical positions. Compared to uninjured contralateral limbs, change in ST tendon insertion point in ACLR limbs resulted in 2.5% loss in peak moment arm and a 3.4% loss in peak torque generating capacity. Accounting for changes to both max isometric force and ST moment arm, the ST had a 14.8% loss in peak torque generating capacity.

There are significant deficits in ST muscle morphology and insertion points following ST-GR ACLR. The ST atrophy and insertion point migration following ACLR may affect force transmission and distribution within the hamstrings and contribute to persistent deficits in knee flexor and internal rotator strength.

Ligament integrity is directly associated with ankle stability. Nearly 40% of ankle sprains result in chronic ankle instability, affecting biomechanics and potentially causing osteoarthritis. Ligament replacement could restore stability and avoid this degenerative pathway, but a greater understanding of ankle ligament behaviour is required. Additionally, autograft or allograft use is limited by donor-site morbidity and inflammatory responses respectively. Decellularised porcine grafts could address this, by removing cellular material to prevent acute immune responses, while preserving mechanical properties.

This project will characterise commonly injured ankle ligaments and damage mechanisms, identify ligament reconstruction requirements, and investigate the potential of decellularised porcine grafts as a replacement material.

Several porcine tendons were evaluated to identify suitable candidates for decellularisation. The viscoelastic properties of native tissues were assessed using dynamic mechanical analysis (DMA), followed by ramp to ‘sub-rupture’ at 1% strain/s, and further DMA. Multiple samples (n=5) were taken along the graft to assess variation along the tendon.

When identifying suitable porcine tendons, a lack of literature on human ankle ligaments was identified. Inconsistencies in measurement methods and properties reported makes comparison between studies difficult.

Preliminary testing on porcine tendons suggested there is little variation in viscoelastic properties along the length of tendon. Testing also suggested strain rates of 1%/s sub-rupture was not large enough to affect viscoelastic properties (no changes in storage or loss moduli or tanẟ). Further testing is underway to improve upon low initial sample numbers and confirm these results, with varying strain rates to identify suitable sub-rupture sprain conditions.

This work highlights need for new data on human ankle ligaments to address knowledge gaps and identify suitable replacement materials. Future work will generate this data and decellularise porcine tendons of similar dimensions. Collagen damage will be investigated using histology and lightsheet microscopy, and viscoelastic changes through DMA.

Introduction and Objective

Osteonecrosis of the femoral head (ONFH) is an evolving and disabling condition that often leads to subchondral collapse in late stages. It is the underlying diagnosis for approximately 3%–12% of total hip arthroplasties (THAs) and the most frequent aetiology for young patients undergoing THA. To date, the pathophysiological mechanisms underlying ONFH remain poorly understood. In this study, we investigated whether ONFH without an obvious etiological factor is related to impaired osteoblast activities, as compared to age-matched patients with primary OA.

No therapeutic strategy, administered in the early stage of osteoarthritis (OA), is fully able to block the degenerative and inflammatory progress of the pathology, whose only solution remains surgery. Aiming to identify minimally invasive therapies able to act on both degenerative and inflammatory processes, infiltrative treatments based on mesenchymal stem cells represent a promising solution due to their proliferative, immunomodulatory, anti-inflammatory, and paracrine ability. Accordingly, the aim of the present study was to investigate the performance of different cell therapies (stem cells from adipose tissue, ADSCs, stromal vascular fraction, SVF, and culture expanded, AECs vs negative control NaCl) in the treatment of OA. An in vivo model of early OA was developed in sheep knee (research protocol N.62/2018-PR date 29/01/2018 approved by the local Ethical Committee). Three and six months after the treatments injections, gross evaluation of articular surfaces (damage score, DS), histological (cartilage thickness, Th; fibrillation index, FI; collagen II content, C2) and mechanical assessment (elastic modulus, E; stress-relaxation time, τ) of cartilage were carried out. Due to the importance of the relationship between structure/composition (histology) and function (mechanics), this study investigated which of the revealed parameters were involved in such relation and how they were influenced by the level of degeneration and by the specific cell treatment, thus to better understand cell-tissue interaction. A statistically significant multi-variable linear regression model was found between τ and Th, FI, C2 (R2 0.7, p-value 8.39E-5). The relation was particularly strong between τ and C2 (p-value 7E-4), with a positive coefficient of 0.92. This is in agreement with literature, where a higher cartilage viscosity was related to a major content of collagen. By dividing the samples in two groups depending on cartilage damage, the more degenerated group (DS > 5) showed statistically significant lower C2 (p-value 0.0124) and τ (p-value 0.05), confirming that collagen content and viscosity decrease with OA grade increasing. Averaging the entire group of samples, the OA degeneration progressed between 3 and 6 months after, and despite, the treatment. But focusing on specific treatments, SVF and AECs differed from the general trend, inducing a higher amount of collagen at 6 months respect to 3 months. Moreover, articular cartilage treated by AECs and, overall, SVF showed a higher content of collagen and a major viscosity respect to the other treatments. We conclude that an injection of mesenchymal stem cells from stromal vascular fraction in early OA articulations could hinder the degenerative process, preserving or even restoring collagen content and viscosity of the articular cartilage

Ligaments and tendons are vital musculoskeletal soft tissues, which are commonly injured due to overuse and trauma. Their distinct functions are well known however their unique structure and biochemical composition and how they change with disease is poorly described. The most commonly injured ligament in the dog and man is the cranial cruciate (CCL) and anterior cruciate ligament (ACL) respectively. Therefore, the structure, function and pathophysiology of disease of this ligament has been most commonly studied in both species. Canine cranial cruciate ligament rupture (CCLR) most commonly occurs following gradual ligament degeneration or disease (CCLD) followed by a non-contact injury or a minor trauma. Several studies have described marked degenerative histological changes in ligament structure prior to and following rupture which consist of loss of the collagen fascicular structure, areas of poor collagen fibril staining, a marked increase in “chondroid” type cells and mineralisation. The ECM protein profile is also altered with increased sulphated glycosaminoglycans content, increased immature collagen cross-links as well as enzymes involved in collagen remodelling. In man, similar findings have been described in the ACL with age and in osteoarthritis (OA). Previously it had been thought that ligament degeneration occurred following OA but these more recent studies suggest that ligament degeneration can lead to joint destabilisation and OA. Being able to determine early degenerative ligament changes in spontaneous clinical cohorts and the mechanisms which cause them are ideal starting points to determine targets for future therapies in the prevention of ligament degradation and rupture. Further identification of ligament cell types in terms of degenerative, responsive and regenerative (stem) types is essential to try and alter ligament cellular and extracellular matrices harnessing their therapeutic potential

Background. Assessment of functional outcome after total hip arthroplasty (THA) often involves subjective patient-reported outcome measures (PROMs) whereas analysis of gait allows more objective assessment. The aims of the study were to compare longitudinal changes of WOMAC function score and ambulatory gait analysis after THA, between patients with low and high self-reported levels of physical function. Methods. Patients undergoing primary THA (n=36; m/f=18/18; mean age=63.9; SD=9.8yrs; BMI=26.3 SD=3.5) were divided in a high and low function group, on their preoperative WOMAC function score. Patients were prospectively measured preoperatively and 3 and 12 months postoperatively. WOMAC function scores 0–100) were compared to inertial sensor based ambulatory gait analysis. Results. WOMAC function scores significantly improved in both low and high groups at 3 months postoperatively whereas gait parameters only improved in with a low pre-operative function. Between 3 and 12 months postoperatively, function scores had not significantly further improved whereas several gait parameters significantly improved in the low function group. WOMAC function scores parameters were only moderately correlated (Spearman's r = 0.33–0.51). Discussion. In routine longitudinal assessment of physical function following THA, ambulatory gait analysis can be supplementary to WOMAC. As gait significantly improved during the first 3 months and following 9 months after THA in patients with a low preoperative level of physical function only, assessment of more demanding tasks than gait may be more sensitive to capture functional improvement in patients with high preoperative function. Acknowledgements. This article presents independent research funded by the National Institute for Health Research (NIHR) under its Programme Grants for Applied Research programme (RP-PG-0407-10070). The views expressed in this article are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health. The research team acknowledges the support of the NIHR, through the Comprehensive Clinical Research Network

Botulinum toxin A (BoNT-A) is a substance that requires repeated application due to its effectiveness being lost 12–16 weeks post application. Performing these intra-muscular injections under anesthesia reduces pain and distress during applications, ensuring effective and successful functional results. This study evaluates motor development of patients undergoing 3 or more repeated BoNT-A application in a tertiary pediatric hospital and the safety as well as effect of 3 different types of anesthesia. 75 children with cerebral palsy who underwent BoNT-A application at least three times consecutively with 6-month intervals and a total of 320 procedures admitted between January 2008 and January 2018 were retrospectively examined. Gross Motor Function Classification System (GMFCS) was employed in motor development evaluation. To observe the improvement in motor development, those with 2-1-0 level decreases in GMFCS classes were grouped and compared in terms of birth time, birth weight, cerebral palsy type and first BoNT-A application age. The 3 types of anesthesia methods (sedation analgesia, larengeal mask anesthesia (LMA) and inhalation mask anesthesia) applied during the procedures were compared in terms of sedation, procedure, recovery and total operation room time. The mean age of the children for all procedures was 45.51 ± 22.40 months. As a result of procedures, significant motor development was observed in 60 (80%) patients (p <0,000∗). No significant difference was observed when the children with cerebral palsy whose GMFCS declined in the form of level 2, 1 and unchanged were compared in terms of first application age, birth weight and gestational age. It was found that 106 (33.1%) were applied sevoflurane with anesthesia mask, 103 (32.1%) were administered sevoflurane with laryngeal mask, and 111 (34.6%) were sedation-analgesia. Only 10 out of 320 procedures were seen to develop side effects (8 vomiting, 2 bronchospasm). In the patients who underwent sedation analgesia during the first 3 BoNT-A procedures, the duration of recovery and total operating room time was seen to be significantly shorter than the others, while there was no difference between the anesthesia methods in the 4th and subsequent procedures. Regardless of the type of anesthesia, the recovery and total operating room times of those having undergone 6 or more procedures were longer than those with less than 6 procedures (p <0.009, p <0.016, respectively). As conclusion, repeated BoNT-A applications in children with CP provides progress in motor steps, it can be applied safely and effectively under anesthesia. Sedation analgesia application provides easier recovery compared to general anesthesia with LMA and mask only in the first three applications. However, recovery time increases with 4 and more repeated applications as the number of applications increases

Cell-based therapies require removal of cells from their optimal in vivotissue context and propagation in vitroto attain suitable number. However, bereft of their optimal tissue niche, cells lose their phenotype and with it their function and therapeutic potential. Biophysical signals, such as surface topography and substrate stiffness, and biochemical signals, such as collagen I, have been shown to maintain permanently differentiated cell phenotype and to precisely regulate stem cell lineage commitment (1, 2). Herein, we developed and characterised substrates of variable rigidity and constant nanotopographical features to offer control over cellular functions during ex vivoexpansion. PDMS substrates with varying ratios of monomer to curing agent (0:1, 1:1, 5:1) were fabricated based on established protocols. Grooved substrates were created using a silinated wafer with groove dimensions of 2µm × 2µm × 2µm; planar control groups were created using flat glass. The aforementioned PDMS solutions were poured onto the wafer/glass, cured at 200 ºC and treated with oxygen plasma. Substrates were then investigated with/without collagen I coating. (0.1, 0.5, and 1 mg/ml). Atomic force microscopy (AFM) and optical profilometry were used to assess the topographical features of the substrates. Dynamic mechanical analysis (DMA) was used to determine the mechanical properties of the substrates. The simultaneous effect of surface topography / substrate rigidity on cell phenotype and function was assessed using human permanently differentiated cells (dermal fibroblasts, tenocytes) and stem cells (human bone marrow stem cells) and various morphometric and gene / protein assays. PDMS substrates of varying stiffness (1000 kPa, 130 kPa, 50 kPa) can be made by varying the Sylgard ratio, while maintaining topographical features. Human adult dermal fibroblasts, tenocytes, and tenocytes attach, align, elongate and deposit aligned extracellular matrix on the grooved PDMS substrate surface of all 3 stiffnesses. Preliminary in vitrodata indicate that surface topography and substrate stiffness play crucial role in maintaining cell phenotype and the prevention of phenotypic drift in vitro

Physical functioning in patients undergoing hip surgery is commonly assessed in three ways: patient-reported outcome measure (PROM), performance test, or clinician-administered measure. It is recommended that several types of measures are used concurrently to capture an extended picture of function. Patient fatigue and burden, time, resources and logistical constraints of clinic and research appointments mean that collecting multiple measures is seldom feasible, leading to focus on a limited number of measures, if not a single one. While there is evidence that performance-tests and PROMs do not fully correlate, correlations between PROMs, performance tests and clinician-administrated measures are yet to be evaluated. It is also not known if the associations between function and patient characteristics depend on how function is measured. The aim of our study was to use different measures to assess function in the same group of patients before their hip surgery to determine 1. how well PROMs, performance tests and clinician-administrated measures correlate with one another and 2. Whether these measures are associated with the same patient characteristics. We conducted a cross-sectional analysis of the pre-operative information of 125 participants listed for hip replacement. The WOMAC function subscale, Harris Hip Score (HHS) and walk-, step- and balance-tests were assessed by questionnaire or during a clinic visit. Participant socio-demographics and medical characteristics were also collected. Correlations between functional measures were investigated with correlation coefficients (r). Regression models were used to test the association between the patient's characteristics and each of the three types of functional measures. None of the correlations between the PROM, clinician-administrated measure and performance tests were very high (r<0.90). The highest correlations were found between the WOMAC-function and the HHS (r=0.7) or the Walk-test (r=0.6), and between the HHS and the walk-test(r=0.7). All the other performance-tests had low correlations with the other measures(r ranging between 0.3 and 0.5). The associations between patient characteristics and functional scores varied by type of measure. Psychological status was associated with the WOMAC function (p-value<0.0001) but not with the other measures. Age was associated with the performance test measures (p-value ranging from ≤0.01 to <0.0001) but not with the WOMAC function. The clinician-administered (HHS) measure was not associated with age or psychological status. When evaluating function prior to hip replacement clinicians and researchers should be aware that each assessment tool captures different aspects of function and that patient characteristics should be taken into account. Psychological status influences the perception of function; patients may be able to do more than they think they can do, and may need encouragement to overcome anxiety. A performance test like a walk-test would provide a more comprehensive assessment of function limitations than a step or balance test, although performance tests are influenced by age. For the most precise description of functional status a combination of measures should be used. Clinicians should supplement their pre-surgery assessment of function with patient-reported measure to include the patient's perspective

Introduction. NF-κB transcription factors regulate a number of genes that are activated under stress conditions. Blockage of the the canonical NF-κB pathway has been emerged as a possible strategy to cure osteoarthritis and rheumatoid arthritis. However, the roles of κNF-B in normal skeletal physiology are largely unknown owing to the lack of suitable animal models. Here, we investigated the function of canonical κNF-B pathway in the cartilaginous skeleton by ablating Nemo (NF-κB essential modulator) in chondrocytes using the Col2a1 transgene. Methods. Mice were analyzed by skeletal staining, histology, proliferation and apoptosis assays at various stages. Histochemistry, GAG assay and immunohistochemistry were utilized to assess the impact of NEMO-deficiency in cytokine-induced cartilage degradation of hip explants. To identify genes regulated through the canonical NF-κB pathway in response to injury, an ex vivo hip avulsion model was applied. 24 genes known to be induced early following cartilage injury were assessed in wildtype and mutant hips by RT-PCR. Time lapse photography was used to investigate chondrocyte migration in vitro. Atomic force microscopy (AFM) was applied to assess biomechanical properties of the cartilage. Pathological changes of articular cartilage were scored in aged joints. Results. Mutant mice exhibited moderate dwarfism postnatally characterized by disorganized growth plate, abnormal chondrocyte proliferation, apoptosis and migration. AFM indentation experiments showed no changes in biomechanical properties of the mutant growth plate compared with control. Exposure of aggrecan degradation neoepitopes and release of GAGs were less pronounced in mutant hip explants stimulated by cytokines. Of the 24 genes regulated 4h following injury in wildtype hips, only Arginase-1 was suppressed in the mutant hips, while the expression levels of most other inflammatory response genes e.g. TSG-6, NOS2, COX2, IL6 and IL1b were unaffected. A small number of genes, IL-18, MMP-3 and Has-2 were further upregulated upon injury in Nemo-deficient compared with wildtype hips. Aging mutant mice showed signs of osteoarthritis comparable to wildtype. Conclusion. Nemo-deficient mice have demonstrated an important role for canonical NF-κB signaling in skeletal growth by modulating chondrocyte behavior. Even though the catabolic effects of pro-inflammatory cytokines in cartilage could be partially eased by blocking the canonical NF-κB pathway, canonical NF-κB signaling seems to play only a minor role in injury-induced inflammatory gene expression and the development of spontaneous OA