Response of Articular Cartilage to Hyperosmolar Stress: Report of an Ex Vivo Injury Model.

BACKGROUND: Physiological 0.9% saline is commonly used as an irrigation fluid in modern arthroscopy. There is a growing body of evidence that a hyperosmolar saline solution has chondroprotective effects, especially if iatrogenic injury occurs. PURPOSE: To (1) corroborate the superiority of a hyperosmolar saline solution regarding chondrocyte survival after mechanical injury and (2) observe the modulatory response of articular cartilage to osmotic stress and injury. STUDY DESIGN: Controlled laboratory study. METHODS: Osteochondral explants were isolated from bovine stifle joints and exposed to either 0.9% saline (308 mOsm) or hyperosmolar saline (600 mOsm) and then damaged with a sharp dermatome blade to attain a confined full-thickness cartilage injury site, incubated in the same fluids for another 3 hours, and transferred to chondropermissive medium for further culture for 1 week. Chondrocyte survival was assessed by confocal imaging, while the cellular response was evaluated over 1 week by relative gene expression for apoptotic and inflammatory markers and mediator release into the medium. RESULTS: The full-thickness cartilage cut resulted in a confined zone of cell death that mainly affected superficial zone chondrocytes. Injured samples that were exposed to hyperosmolar saline showed less expansion of cell death in both the axial (P < .007) and the coronal (P < .004) plane. There was no progression of cell death during the following week of culture. Histological assessment revealed an intact cartilage matrix and normal chondrocyte morphology. Inflammatory and proapoptotic genes were upregulated on the first days postexposure with a notable downregulation toward day 7. Mediator release into the medium was concentrated on day 3. CONCLUSION: This in vitro cartilage injury model provides further evidence for the chondroprotective effect of a hyperosmolar saline irrigation fluid, as well as novel data on the capability of articular cartilage to quickly regain joint homeostasis after osmotic stress and injury. CLINICAL RELEVANCE: Raising the osmolarity of an irrigating solution may be a simple and safe strategy to protect articular cartilage during arthroscopic surgery.

Effect of expansion media and fibronectin coating on growth and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells.

In the field of regenerative medicine, considerable advances have been made from the technological and biological point of view. However, there are still large gaps to be filled regarding translation and application of mesenchymal stromal cell (MSC)-based therapies into clinical practice. Indeed, variables such as cell type, unpredictable donor variation, and expansion/differentiation methods lead to inconsistencies. Most protocols use bovine serum (FBS) derivatives during MSC expansion. However, the xenogeneic risks associated with FBS limits the use of MSC-based products in clinical practice. Herein we compare a chemically defined, xenogeneic-free commercial growth medium with a conventional medium containing 10% FBS and 5 ng/ml FGF2. Furthermore, the effect of a fibronectin-coated growth surface was investigated. The effect of the different culture conditions on chondrogenic commitment was assessed by analyzing matrix deposition and gene expression of common chondrogenic markers. Chondrogenic differentiation potential was similar between the FBS-containing αMEM and the chemically defined medium with fibronectin coating. On the contrary, the use of fibronectin coating with FBS-containing medium appeared to reduce the differentiation potential of MSCs. Moreover, cells that were poorly responsive to in vitro chondrogenic stimuli were shown to improve their differentiation potential after expansion in a TGF-ß1 containing medium. In conclusion, the use of a xenogeneic-free medium provides a suitable alternative for human bone marrow MSC expansion, due the capability to maintain cell characteristic and potency. To further improve chondrogenic potential of BMSCs, priming the cells with TGF-ß1 during expansion is a promising strategy.

Preclinical ex-vivo Testing of Anti-inflammatory Drugs in a Bovine Intervertebral Degenerative Disc Model.

Discogenic low back pain (LBP) is a main cause of disability and inflammation is presumed to be a major driver of symptomatic intervertebral disc degeneration (IDD). Anti-inflammatory agents are currently under investigation as they demonstrated to alleviate symptoms in patients having IDD. However, their underlying anti-inflammatory and regenerative activity is poorly explored. The present study sought to investigate the potential of Etanercept and Tofacitinib for maintaining disc homeostasis in a preclinical intervertebral disc (IVD) organ culture model within IVD bioreactors allowing for dynamic loading and nutrient exchange. Bovine caudal IVDs were cultured in a bioreactor system for 4 days to simulate physiological or degenerative conditions: (1) Phy-physiological loading (0.02-0.2 MPa; 0.2 Hz; 2 h/day) and high glucose DMEM medium (4.5 g/L); (2) Deg+Tumor necrosis factor α (TNF-α)-degenerative loading (0.32-0.5 MPa; 5 Hz; 2 h/day) and low glucose DMEM medium (2 g/L), with TNF-α injection. Etanercept was injected intradiscally while Tofacitinib was supplemented into the culture medium. Gene expression in the IVD tissue was measured by RT-qPCR. Release of nitric oxide (NO), interleukin 8 (IL-8) and glycosaminoglycan (GAG) into the IVD conditioned medium were analyzed. Cell viability in the IVD was assessed using lactate dehydrogenase and ethidium homodimer-1 staining. Immunohistochemistry was performed to assess protein expression of IL-1ß, IL-6, IL-8, and collagen type II in the IVD tissue. Etanercept and Tofacitinib downregulated the expression of IL-1ß, IL-6, IL-8, Matrix metalloproteinase 1 (MMP1), and MMP3 in the nucleus pulposus (NP) tissue and IL-1ß, MMP3, Cyclooxygenase-2 (COX2), and Nerve growth factor (NGF) in the annulus fibrosus (AF) tissue. Furthermore, Etanercept significantly reduced the IL-1ß positively stained cells in the outer AF and NP regions. Tofacitinib significantly reduced IL-1ß and IL-8 positively stained cells in the inner AF region. Both, Etanercept and Tofacitinib reduced the GAG loss to the level under physiological culture condition. Etanercept and Tofacitinib are able to neutralize the proinflammatory and catabolic environment in the IDD organ culture model. However, combined anti-inflammatory and anabolic treatment may be required to constrain accelerated IDD and relieving inflammation-induced back pain.

Comparing case-control study for treatment of proximal tibia fractures with a complete metaphyseal component in two centers with different distinct strategies: fixation with Ilizarov frame or locking plates.

BACKGROUND: The purpose of this study was to compare two methods of stabilization for proximal tibia fractures (AO 41) with a complete metaphyseal component, external fixation with the Ilizarov wire frame, and internal fixation with locking plates. METHODS: Patients from two level 1 trauma centers treated between 2009 and 2015 were included in a retrospective comparing cohort study. The first center stabilized the non-pathological, proximal tibia fractures exclusively with external fixation and the second with internal plating. Combined clinically and radiologically evaluated, bone healing was the primary outcome. The secondary outcomes included complications, range of motion (ROM) and axial alignment of the knee, the reoperation rate within 6 months, heterotopic ossifications (HTO), and signs of posttraumatic osteoarthritis (PTOA). A logistic regression analysis corrected for uneven distributed parameters. RESULTS: The 62 patients treated with Ilizarov frame and the 68 patients treated with plate fixation were comparable regarding epidemiological parameters, injury characteristics, and comorbidity except for injury severity score (ISS) and smoking behavior. The time of healing was shorter in the group undergoing plate fixation (p = 0.041); however, the incidence of non-unions was equal. Furthermore, there was no difference regarding the rate of deep infections, thrombosis, alignment, reoperations, PTOA, and ROM. Heterotopic ossifications were more prevalent following plate fixation (13.2 vs 1.6%, p = .013). External fixation was associated with a higher rate of superficial infections (40.4 vs 2.9%, p = .000). The initial displacement, the incidence of deep infections, and the classification significantly influenced the incidence of non-unions in both groups (p < 0.02). CONCLUSIONS: Fixation of proximal tibia fractures with plates resulted in a slightly shorter healing time compared to Ilizarov frame stabilization. Furthermore, the complication profiles differ with more heterotopic ossifications and less superficial infections following internal plating. TRIAL REGISTRATION: DRKS, DRKS00013275 , Registered 11/2/2017, Retrospectively registered.

Work-family conflict and neck and back pain in surgical nurses.

OBJECTIVE: Surgical nurses' work is physically and mentally demanding, possibly leading to work-family conflict (WFC). The current study tests WFC to be a risk factor for neck and lower back pain (LBP). Job influence and social support are tested as resources that could buffer the detrimental impact of WFC. METHODS: Forty-eight surgical nurses from two university hospitals in Germany and Switzerland were recruited. WFC was assessed with the Work-Family Conflict Scale. Job influence and social support were assessed with the Copenhagen Psychosocial Questionnaire, and back pain was assessed with the North American Spine Society Instrument. RESULTS: Multiple linear regression analyses confirmed WFC as a significant predictor of cervical pain (ß = 0.45, p < 0.001) and LBP (ß = 0.33, p = 0.012). Job influence and social support did not turn out to be significant predictors and were not found to buffer the impact of WFC in moderator analyses. CONCLUSION: WFC is likely to affect neck and back pain in surgery nurses. Work-life interventions may have the potential to reduce WFC in surgery nurses.

The Potential for Synovium-derived Stem Cells in Cartilage Repair.

BACKGROUND: Articular cartilage defects often result in pain, loss of function and finally osteoarthritis. Developing cell-based therapies for cartilage repair is a major goal of orthopaedic research. Autologous chondrocyte implantation is currently the gold standard cell-based surgical procedure for the treatment of large, isolated, full thickness cartilage defects. Several disadvantages such as the need for two surgical procedures or hypertrophic regenerative cartilage, underline the need for alternative cell sources. OBJECTIVE: Mesenchymal stem cells, particularly synovium-derived mesenchymal stem cells, represent a promising cell source. Synovium-derived mesenchymal stem cells have attracted considerable attention since they display great chondrogenic potential and less hypertrophic differentiation than mesenchymal stem cells derived from bone marrow. The aim of this review was to summarize the current knowledge on the chondrogenic potential for synovial stem cells in regard to cartilage repair purposes. RESULTS: A literature search was carried out identifying 260 articles in the databases up to January 2017. Several in vitro and initial animal in vivo studies of cartilage repair using synovia stem cell application showed encouraging results. Since synvoium-derived stem cells are located in the direct vicinity of cartilage and cartilage lesions these cells might even contribute to natural cartilage regeneration. The only one published human in vivo study with 10 patients revealed good results concerning postoperative outcome, MRI, and histologic features after a two-stage implantation of synovial stem cells into an isolated cartilage defect of the femoral condyle. CONCLUSION: Synovium-derived stem cells possess great chondrogenic potential and showed encouraging results for cartilage repair purposes. Furthermore, synovial stem cells play an important role in joint homeostasis and possibly in natural cartilage repair. Further studies are needed to elucidate the interplay of synovial stem cells and chondrocytes, and the promising role of synovium-derived stem cells in cartilage tissue engineering.

Examination of concomitant glenohumeral pathologies in patients treated arthroscopically for calcific tendinitis of the shoulder and implications for routine diagnostic joint exploration.

BACKGROUND: Glenohumeral exploration is routinely performed during arthroscopic removal of rotator cuff calcifications in patients with calcific tendinitis of the shoulder (CTS). However, evidence on the prevalence of intraarticular co-pathologies is lacking and the benefit of glenohumeral exploration remains elusive. The aim of the present study was to assess and quantify intraoperative pathologies during arthroscopic removal of rotator cuff calcifications in order to determine whether standardized diagnostic glenohumeral exploration appears justified in CTS patients. METHODS: One hundred forty five patients undergoing arthroscopic removal of calcific depots (CD) that failed conservative treatment were included in a retrospective cohort study. Radiographic parameters including number/localization of calcifications and acromial types, intraoperative arthroscopic findings such as configuration of glenohumeral ligaments, articular cartilage injuries, and characteristics of calcifications and sonographic parameters (characteristics/localization of calcification) were recorded. RESULTS: One hundred forty five patients were analyzed. All CDs were removed by elimination with a blunt hook probe via "squeeze-and-stir-technique" assessed postoperatively via conventional X-rays. Neither subacromial decompression nor refixation of the rotator cuff were performed in any patient. Prevalence of glenohumeral co-pathologies, such as partial tears of the proximal biceps tendon (2.1%), superior labral tears from anterior to posterior (SLAP) lesions (1.4%), and/or partial rotator cuff tears (0.7%) was low. Most frequently, glenohumeral articular cartilage was either entirely intact (ICRS grade 0 (humeral head/glenoid): 46%/48%) or showed very mild degenerative changes (ICRS grade 1: 30%/26%). Two patients (1.3%) required intraarticular surgical treatment due to a SLAP lesion type III (n = 1) and an intraarticular rupture of CD (n = 1). CONCLUSIONS: Routine diagnostic glenohumeral exploration does not appear beneficial in arthroscopic treatment of CTS due to the low prevalence of intraarticular pathologies which most frequently do not require surgical treatment. Exploration of the glenohumeral joint in arthroscopic removal of CD should only be performed in case of founded suspicion of relevant concomitant intraarticular pathologies.

In-vitro chondrogenic potential of synovial stem cells and chondrocytes allocated for autologous chondrocyte implantation - a comparison : Synovial stem cells as an alternative cell source for autologous chondrocyte implantation.

PURPOSE: The use of passaged chondrocytes is the current standard for autologous chondrocyte implantation (ACI). De-differentiation due to amplification and donor site morbidity are known drawbacks highlighting the need for alternative cell sources. METHODS: Via clinically validated flow cytometry analysis, we compared the expression of human stem cell and cartilage markers (collagen type 2 (Col2), aggrecan (ACAN), CD44) of chondrocytes (CHDR), passaged chondrocytes for ACI (CellGenix™), bone marrow derived mesenchymal stem cells (BMSC), and synovial derived stem cells (SDSC). RESULTS: Primary, human BMSC and SDSC revealed similar adipogenic, osteogenic, and chondrogenic differentiation potential and stem cell marker expression. However, the expression of the chondrogenic markers Col2 and ACAN was statistically significant higher in SDSC. CHDR and SDSC expressed ACAN and CD44 equally, but Col2 was expressed more strongly on the SDSC surface. The marker expression of SDSC from osteoarthritic joints (Kellgren-Lawrence score ≥3) versus normal knees (Kellgren-Lawrence score ≤2) did not differ. Similarly, there was no difference between temporarily frozen and fresh SDSC. Col2 and ACAN surface expression declined with further passaging, whereas CD44 remained unchanged. We observed the same effect after reducing the serum content. When comparing CHDR for ACI with SDSC of the same passage (P2/3), both Col2 and ACAN, correlating with clinical outcome, were expressed higher in SDSC. CONCLUSIONS: In summary, SDSC demonstrated high differentiation potential and a stable chondrogenic phenotype. They might therefore be better suitable for ACI than BMSC or passaged CHDR.

Quantitative work demands, emotional demands, and cognitive stress symptoms in surgery nurses.

In surgery, cognitive stress symptoms, including problems in concentrating, deciding, memorising, and reflecting are risks to patient safety. Recent evidence points to social stressors as antecedents of cognitive stress symptoms in surgery personnel. The current study tests whether cognitive stress symptoms are positively associated with emotional abuse, emotional- and task-related demands and resources in surgery work. Forty-eight surgery nurses from two hospitals filled out the Copenhagen Psychosocial Questionnaire in its German version. Task-related and emotional demands were positively related to cognitive stress symptoms. In a stepwise, multiple, linear regression of cognitive stress symptoms on task-related and emotional demands, emotional abuse and emotional demands were unique predictors (p < .05). Efforts to increase patient safety should address emotional abuse, emotional demands, and, therefore, communication and cooperation team climate in surgery personnel.

The trans-well coculture of human synovial mesenchymal stem cells with chondrocytes leads to self-organization, chondrogenic differentiation, and secretion of TGFß.

BACKGROUND: Synovial mesenchymal stem cells (SMSC) possess a high chondrogenic differentiation potential, which possibly supports natural and surgically induced healing of cartilage lesions. We hypothesized enhanced chondrogenesis of SMSC caused by the vicinity of chondrocytes (CHDR). METHODS: Human SMSC and CHDR interactions were investigated in an in-vitro trans-well monolayer coculture over a time period of up to 21 days. Protein expression was analyzed using histology, immunostaining, or enzyme-linked immunosorbent assay. Additionally, mRNA expression was assessed by quantitative PCR. RESULTS: After 7 days, phase-contrast microscopy revealed cell aggregation of SMSC in coculture with CHDR. Afterwards, cells formed spheres and lost adherence. However, this phenomenon was not observed when culturing SMSC alone. Fluorescence labeling showed concurrent collagen type II expression. Addition of transforming growth factor beta (TGFß) to the cocultures induced SMSC aggregation in less time and with higher intensity. Additionally, alcian blue staining demonstrated enhanced glycosaminoglycan expression around SMSC aggregates after 1 and 2 weeks. Although TGFß mRNA was expressed in all SMSC, the protein was measured with constantly increasing levels over 21 days only in supernatants of the cocultures. Considering the enhanced mRNA levels following supplementation with TGFß, a positive feedback mechanism can be supposed. In line with the development of a chondrogenic phenotype, aggrecan mRNA expression increased after 7 and 14 days in the cocultures with and without TGFß. Coculture conditions also amplified collagen type II mRNA expression after 2 weeks without and already after 1 week with TGFß. There was no difference in collagen type I and type X expression between SMSC alone and the coculture with CHDR. Expression of both collagens increased following addition of TGFß. mRNA data correlated with the intensity of immunofluorescence staining. CONCLUSIONS: Paracrine effects of CHDR induce a chondrogenic phenotype in SMSC possibly mimicking joint homeostasis. Coculture approaches may lead to a better understanding of cellular interactions with potential implications for cartilage repair procedures.

Clinical trial and in-vitro study comparing the efficacy of treating bony lesions with allografts versus synthetic or highly-processed xenogeneic bone grafts.

BACKGROUND: Our study aim was to compare allogeneic cancellous bone (ACB) and synthetic or highly-processed xenogeneic bone substitutes (SBS) in the treatment of skeletal defects in orthopedic surgery. METHODS: 232 patients treated for bony lesions with ACB (n = 116) or SBS (n = 116) within a 10-year time period were included in this case-control study. Furthermore, both materials were seeded with human osteoblasts (hOB, n = 10) and analyzed by histology, for viability (AlamarBlue®) and protein expression activity (Luminex®). RESULTS: The complication rate was 14.2 %, proportion of defects without bony healing 3.6 %; neither outcome parameter differed comparing the intervention groups. Failed consolidation correlated with an increase in complications (p < 0.03). The rate of complications was further highly significant in association with the location of use (p < 0.001), but did not depend on age, ASA risk classification, BMI, smoking behavior or type of insurance. However, those factors did significantly influence the bony healing rate (p < 0.02). Complication and consolidation rates were independent of gender and the filling substances employed within the different locations. Histological examination revealed similar bone structures, whereas cell remnants were apparent only in the allografts. Both materials were biocompatible in-vitro, and seeded with human osteoblasts. The cells remained vital over the 3-week culture period and produced microscopically typical bone matrix. We observed initially increased expression of osteocalcin, osteopontin, and osteoprotegerin as well as leptin and adiponectin secretion declining after 1 week, especially in the ACB group. CONCLUSION: Although both investigated materials appeared to be similarly suitable for the treatment of skeletal lesions in-vivo and in-vitro, outcome was decisively influenced by other factors such as the site of use or epidemiological parameters.

Clinical outcome and T2 assessment following autologous matrix-induced chondrogenesis in osteochondral lesions of the talus.

PURPOSE: Scientific evidence for the treatment of osteochondral lesions (OCLs) of the talus is limited. The aim of this study was an evaluation of the clinical outcome after a one-step autologous subchondral cancellous bone graft and autologous matrix-induced chondrogenesis (AMIC) in medial OCLs of the talus and the assessment of the repair tissue (RT). METHODS: Seventeen patients (eight women, nine men; mean age, 38.8 ± 15.7 years) with an OCL of the medial talus underwent surgery. Clinical and radiological assessment was performed after a mean follow-up of 39.5 ± 18.4 months, including established scoring systems (American Orthopaedic Foot and Ankle Society [AOFAS] Score, Foot Function Index [FFI], visual analogue scale [VAS]), evaluation of Magnetic Resonance Observation of Cartilage Repair Tissue scoring system (MOCART Score) and T2 mapping. RESULTS: Preoperative pain (7.8 ± 2.1) significantly improved to an average of 3.2 ± 2.4 postoperatively. AOFAS Score averaged 82.6 ± 13.4, MOCART Score 52.7 ± 15.9. Mean T2 relaxation time of the RT was 41.6 ± 6.3 ms and showed no significant differences to the surrounding cartilage (mean, 38.8 ± 8.5; p = 0.58). MOCART Score significantly correlated with the AOFAS Score (rho = 0.574, p = 0.040). T2 relaxation time of the RT significantly correlated with the MOCART Score (rho = 0.593, p = 0.033). CONCLUSIONS: The one-step autologous subchondral cancellous bone grafting and AMIC leads to a significant reduction in postoperative pain and satisfying postoperative functional outcome in mid-term follow-up. Magnetic resonance imaging (MRI) assessment demonstrated a good quality of regenerative tissue similar to the MRI ultrastructure of the surrounding cartilage.