Locally Directed Recombinant Adeno- Associated Virus-Mediated IGF-1 Gene Therapy Enhances Osteochondral Repair and Counteracts Early Osteoarthritis In Vivo.

BACKGROUND: Restoration of osteochondral defects is critical, because osteoarthritis (OA) can arise. HYPOTHESIS: Overexpression of insulin-like growth factor 1 (IGF-1) via recombinant adeno-associated viral (rAAV) vectors (rAAV-IGF-1) would improve osteochondral repair and reduce parameters of early perifocal OA in sheep after 6 months in vivo. STUDY DESIGN: Controlled laboratory study. METHODS: Osteochondral defects were created in the femoral trochlea of adult sheep and treated with rAAV-IGF-1 or rAAV-lacZ (control) (24 defects in 6 knees per group). After 6 months in vivo, osteochondral repair and perifocal OA were assessed by well-established macroscopic, histological, and immunohistochemical scoring systems as well as biochemical and micro-computed tomography evaluations. RESULTS: Application of rAAV-IGF-1 led to prolonged (6 months) IGF-1 overexpression without adverse effects, maintaining a significantly superior overall cartilage repair, together with significantly improved defect filling, extracellular matrix staining, cellular morphology, and surface architecture compared with rAAV-lacZ. Expression of type II collagen significantly increased and that of type I collagen significantly decreased. Subchondral bone repair and tidemark formation were significantly improved, and subchondral bone plate thickness and subarticular spongiosa mineral density returned to normal. The OA parameters of perifocal structure, cell cloning, and matrix staining were significantly better preserved upon rAAV-IGF-1 compared with rAAV-lacZ. Novel mechanistic associations between parameters of osteochondral repair and OA were identified. CONCLUSION: Local rAAV-mediated IGF-1 overexpression enhanced osteochondral repair and ameliorated parameters of perifocal early OA. CLINICAL RELEVANCE: IGF-1 gene therapy may be beneficial in repair of focal osteochondral defects and prevention of perifocal OA.

Modulation of early osteoarthritis by tibiofemoral re-alignment in sheep.

OBJECTIVE: To investigate whether tibiofemoral alignment influences early knee osteoarthritis (OA). We hypothesized that varus overload exacerbates early degenerative osteochondral changes, and that valgus underload diminishes early OA. METHOD: Normal, over- and underload were induced by altering alignment via high tibial osteotomy in adult sheep (n = 8 each). Simultaneously, OA was induced by partial medial anterior meniscectomy. At 6 weeks postoperatively, OA was examined in five individual subregions of the medial tibial plateau using Kellgren-Lawrence grading, quantification of macroscopic OA, semiquantitative histopathological OA and immunohistochemical type-II collagen, ADAMTS-5, and MMP-13 scoring, biochemical determination of DNA and proteoglycan contents, and micro-computed tomographic evaluation of the subchondral bone. RESULTS: Multivariate analyses revealed that OA cartilaginous changes had a temporal priority over subchondral bone changes. Underload inhibited early cartilage degeneration in a characteristic topographic pattern (P ≥ 0.0983 vs. normal), in particular below the meniscal damage, avoided alterations of the subarticular spongiosa (P ≥ 0.162 vs. normal), and prevented the disturbance of otherwise normal osteochondral correlations. Overload induced early alterations of the subchondral bone plate microstructure towards osteopenia, including significantly decreased percent bone volume and increased bone surface-to-volume ratio (all P ≤ 0.0359 vs. normal). CONCLUSION: The data provide high-resolution evidence that tibiofemoral alignment modulates early OA induced by a medial meniscus injury in adult sheep. Since underload inhibits early OA, these data also support the clinical value of strategies to reduce the load in an affected knee compartment to possibly decelerate structural OA progression.

Accuracy of Personalized Computed Tomographic 3D Templating for Acetabular Cup Placement in Revision Arthroplasty.

Background: Revision hip arthroplasty presents a surgical challenge, necessitating meticulous preoperative planning to avert complications like periprosthetic fractures and aseptic loosening. Historically, assessment of the accuracy of three-dimensional (3D) versus two-dimensional (2D) templating has focused exclusively on primary hip arthroplasty. Materials and Methods: In this retrospective study, we examined the accuracy of 3D templating for acetabular revision cups in 30 patients who underwent revision hip arthroplasty. Utilizing computed tomography scans of the patients' pelvis and 3D templates of the implants (Aesculap Plasmafit, B. Braun; Aesculap Plasmafit Revision, B. Braun; Avantage Acetabular System, Zimmerbiomet, EcoFit 2M, Implantcast; Tritanium Revision, Stryker), we performed 3D templating and positioned the acetabular cup implants accordingly. To evaluate accuracy, we compared the planned sizes of the acetabular cups in 2D and 3D with the sizes implanted during surgery. Results: An analysis was performed to examine potential influences on templating accuracy, specifically considering factors such as gender and body mass index (BMI). Significant statistical differences (p < 0.001) in the accuracy of size prediction were observed between 3D and 2D templating. Personalized 3D templating exhibited an accuracy rate of 66.7% for the correct prediction of the size of the acetabular cup, while 2D templating achieved an exact size prediction in only 26.7% of cases. There were no statistically significant differences between the 2D and 3D templating methods regarding gender or BMI. Conclusion: This study demonstrates that 3D templating improves the accuracy of predicting acetabular cup sizes in revision arthroplasty when compared to 2D templating. However, it should be noted that the predicted implant size generated through 3D templating tended to overestimate the implanted implant size by an average of 1.3 sizes.

A Photopolymerizable Biocompatible Hyaluronic Acid Hydrogel Promotes Early Articular Cartilage Repair in a Minipig Model In Vivo.

Articular cartilage defects represent an unsolved clinical challenge. Photopolymerizable hydrogels are attractive candidates supporting repair. This study investigates the short-term safety and efficacy of two novel hyaluronic acid (HA)-triethylene glycol (TEG)-coumarin hydrogels photocrosslinked in situ in a clinically relevant large animal model. It is hypothesized that HA-hydrogel-augmented microfracture (MFX) is superior to MFX in enhancing early cartilage repair, and that the molar degree of substitution and concentration of HA affects repair. Chondral full-thickness defects in the knees of adult minipigs are treated with either 1) debridement (No MFX), 2) debridement and MFX, 3) debridement, MFX, and HA hydrogel (30% molar derivatization, 30 mg mL-1 HA; F3) (MFX+F3), and 4) debridement, MFX, and HA hydrogel (40% molar derivatization, 20 mg mL-1 HA; F4) (MFX+F4). After 8 weeks postoperatively, MFX+F3 significantly improves total macroscopic and histological scores compared with all other groups without negative effects, besides significantly enhancing the individual repair parameters "defect architecture," "repair tissue surface" (compared with No MFX, MFX), and "subchondral bone" (compared with MFX). These data indicate that photopolymerizable HA hydrogels enable a favorable metastable microenvironment promoting early chondrogenesis in vivo. This work also uncovers a mechanism for effective HA-augmented cartilage repair by combining lower molar derivatization with higher concentrations.

Subgroup analysis of scientific performance in the field of arthroplasty.

Introduction: Arthroplasty is the final treatment option for maintaining mobility and quality of life in many primary degenerative and (post-) traumatic joint diseases. Identification of research output and potential deficits for specific subspecialties may be an important measure to achieve long-term improvement of patient care in this field. Methods: Using specific search terms and Boolean operators, all studies published since 1945 to the subgroups of arthroplasty listed in the Web of Science Core Collection were included. All identified publications were analysed according to bibliometric standards, and comparative conclusions were drawn regarding the scientific merit of each subgroup. Results: Most publications investigated the subgroups of septic surgery and materials followed by approach, navigation, aseptic loosening, robotic and enhanced recovery after surgery (ERAS). In the last 5 years, research in the fields of robotic and ERAS achieved the highest relative increase in publications In contrast, research on aseptic loosening has continued to lose interest over the last 5 years. Publications on robotics and materials received the most funding on average while those on aseptic loosening received the least. Most publications originated from USA, Germany, and England, except for research on ERAS in which Denmark stood out. Relatively, publications on aseptic loosening received the most citations, whereas the absolute scientific interest was highest for the topic infection. Discussion: In this bibliometric subgroup analysis, the primary scientific outputs focused on septic complications and materials research in the field of arthroplasty. With decreasing publication output and the least financial support, intensification of research on aseptic loosening is urgently recommended.

Axial alignment is a critical regulator of knee osteoarthritis.

Although osteoarthritis (OA), a leading cause of disability, has been associated with joint malalignment, scientific translational evidence for this link is lacking. In a clinical case study, we provide evidence of osteochondral recovery upon unloading symptomatic isolated medial tibiofemoral knee OA associated with varus malalignment. By mapping response correlations at high resolution, we identify spatially complex degenerative changes in cartilage after overloading in a clinically relevant ovine model. We further report that unloading diminishes OA cartilage degeneration and alterations of critical parameters of the subchondral bone plate in a similar topographic fashion. Last, therapeutic unloading shifted the articular cartilage and subchondral bone phenotype to normal and restored several physiological correlations disturbed in neutral and varus OA, suggesting a protective effect on the integrity of the entire osteochondral unit. Collectively, these findings identify modifiable trajectories with considerable translational potential to reduce the burden of human OA.

Hydrogel-Guided, rAAV-Mediated IGF-I Overexpression Enables Long-Term Cartilage Repair and Protection against Perifocal Osteoarthritis in a Large-Animal Full-Thickness Chondral Defect Model at One Year In Vivo.

The regeneration of focal articular cartilage defects is complicated by the reduced quality of the repair tissue and the potential development of perifocal osteoarthritis (OA). Biomaterial-guided gene therapy may enhance cartilage repair by controlling the release of therapeutic sequences in a spatiotemporal manner. Here, the benefits of delivering a recombinant adeno-associated virus (rAAV) vector coding for the human insulin-like growth factor I (IGF-I) via an alginate hydrogel (IGF-I/AlgPH155) to enhance repair of full-thickness chondral defects following microfracture surgery after one year in minipigs versus control (lacZ/AlgPH155) treatment are reported. Sustained IGF-I overexpression is significantly achieved in the repair tissue of defects treated with IGF-I/AlgPH155 versus those receiving lacZ/AlgPH155 for one year and in the cartilage surrounding the defects. Administration of IGF-I/AlgPH155 significantly improves parameters of cartilage repair at one year relative to lacZ/AlgPH155 (semiquantitative total histological score, cell densities, matrix deposition) without deleterious or immune reactions. Remarkably, delivery of IGF-I/AlgPH155 also significantly reduces perifocal OA and inflammation after one year versus lacZ/AlgPH155 treatment. Biomaterial-guided rAAV gene transfer represents a valuable clinical approach to promote cartilage repair and to protect against OA.

Thermosensitive Hydrogel Based on PEO-PPO-PEO Poloxamers for a Controlled In Situ Release of Recombinant Adeno-Associated Viral Vectors for Effective Gene Therapy of Cartilage Defects.

Advanced biomaterial-guided delivery of gene vectors is an emerging and highly attractive therapeutic solution for targeted articular cartilage repair, allowing for a controlled and minimally invasive delivery of gene vectors in a spatiotemporally precise manner, reducing intra-articular vector spread and possible loss of the therapeutic gene product. As far as it is known, the very first successful in vivo application of such a biomaterial-guided delivery of a potent gene vector in an orthotopic large animal model of cartilage damage is reported here. In detail, an injectable and thermosensitive hydrogel based on poly(ethylene oxide) (PEO)-poly(propylene oxide) (PPO)-PEO poloxamers, capable of controlled release of a therapeutic recombinant adeno-associated virus (rAAV) vector overexpressing the chondrogenic sox9 transcription factor in full-thickness chondral defects, is applied in a clinically relevant minipig model in vivo. These comprehensive analyses of the entire osteochondral unit with multiple standardized evaluation methods indicate that rAAV-FLAG-hsox9/PEO-PPO-PEO hydrogel-augmented microfracture significantly improves cartilage repair with a collagen fiber orientation more similar to the normal cartilage and protects the subchondral bone plate from early bone loss.

Topographic modeling of early human osteoarthritis in sheep.

Articular cartilage damage occurring during early osteoarthritis (OA) is a key event marking the development of the disease. Here, we modeled early human OA by gathering detailed spatiotemporal data from surgically induced knee OA development in sheep. We identified a specific topographical pattern of osteochondral changes instructed by a defined meniscal injury, showing that both cartilage and subchondral bone degeneration are initiated from the region adjacent to the damage. Alterations of the subarticular spongiosa arising locally and progressing globally disturbed the correlations of cartilage with subchondral bone seen at homeostasis and were indicative of disease progression. We validated our quantitative findings against human OA, showing a similar pattern of early OA correlating with regions of meniscal loss and an analogous late critical disturbance within the entire osteochondral unit. This translational model system can be used to elucidate mechanisms of OA development and provides a roadmap for investigating regenerative therapies.

Effects of TGF-ß Overexpression via rAAV Gene Transfer on the Early Repair Processes in an Osteochondral Defect Model in Minipigs.

BACKGROUND: Application of the chondrogenic transforming growth factor beta (TGF-ß) is an attractive approach to enhance the intrinsic biological activities in damaged articular cartilage, especially when using direct gene transfer strategies based on the clinically relevant recombinant adeno-associated viral (rAAV) vectors. PURPOSE: To evaluate the ability of an rAAV-TGF-ß construct to modulate the early repair processes in sites of focal cartilage injury in minipigs in vivo relative to control (reporter lacZ gene) vector treatment. STUDY DESIGN: Controlled laboratory study. METHODS: Direct administration of the candidate rAAV-human TGF-ß (hTGF-ß) vector was performed in osteochondral defects created in the knee joint of adult minipigs for macroscopic, histological, immunohistochemical, histomorphometric, and micro-computed tomography analyses after 4 weeks relative to control (rAAV- lacZ) gene transfer. RESULTS: Successful overexpression of TGF-ß via rAAV at this time point and in the conditions applied here triggered the cellular and metabolic activities within the lesions relative to lacZ gene transfer but, at the same time, led to a noticeable production of type I and X collagen without further buildup on the subchondral bone. CONCLUSION: Gene therapy via direct, local rAAV-hTGF-ß injection stimulates the early reparative activities in focal cartilage lesions in vivo. CLINICAL RELEVANCE: Local delivery of therapeutic (TGF-ß) rAAV vectors in focal defects may provide new, off-the-shelf treatments for cartilage repair in patients in the near future.

Subchondral drilling for articular cartilage repair: a systematic review of translational research.

Articular cartilage defects may initiate osteoarthritis. Subchondral drilling, a widely applied clinical technique to treat small cartilage defects, does not yield cartilage regeneration. Various translational studies aiming to improve the outcome of drilling have been performed; however, a robust systematic analysis of its translational evidence was still lacking. Here, we performed a systematic review of the outcome of subchondral drilling for knee cartilage repair in translational animal models. A total of 12 relevant publications studying 198 animals was identified, detailed study characteristics were extracted, and methodological quality and risk of bias were analyzed. Subchondral drilling led to improved repair outcome compared with defects that were untreated or treated with abrasion arthroplasty for cartilage repair in multiple translational models. Within the 12 studies, considerable subchondral bone changes were observed, including subchondral bone cysts and intralesional osteophytes. Furthermore, extensive alterations of the subchondral bone microarchitecture appeared in a temporal pattern in small and large animal models, together with specific topographic aspects of repair. Moreover, variable technical aspects directly affected the outcomes of osteochondral repair. The data from this systematic review indicate that subchondral drilling yields improved short-term structural articular cartilage repair compared with spontaneous repair in multiple small and large animal models. These results have important implications for future investigations aimed at an enhanced translation into clinical settings for the treatment of cartilage defects, highlighting the importance of considering specific aspects of modifiable variables such as improvements in the design and reporting of preclinical studies, together with the need to better understand the underlying mechanisms of cartilage repair following subchondral drilling.

Sustained spatiotemporal release of TGF-ß1 confers enhanced very early chondrogenic differentiation during osteochondral repair in specific topographic patterns.

The continuous presence of TGF-ß is critically important to induce effective chondrogenesis. To investigate chondrogenesis in a cartilage defect, we tested the hypothesis that the implantation of TGF-ß1-releasing scaffolds improves very early cartilage repair in vivo. Spatiotemporal controlled release of TGF-ß1 was achieved from multiblock scaffolds that were implanted in osteochondral defects in the medial femoral condyles of adult minipigs. We observed a sustained presence of TGF-ß1 at 4 wk in vivo, which significantly promoted structural aspects of early overall cartilage repair, especially cellularity, cellular morphology, and safranin O staining intensity. Furthermore, early aggrecan and type II collagen production were both increased in specific topographic patterns in cartilaginous repair tissue. Sustained release of TGF-ß1 also increased cell numbers and proliferation, staining intensities for the stem cell surface marker, CD105, and number of stromal cell-derived factor-1 (SDF-1) -positive cells within cartilaginous repair tissue. These data identify a mechanism by which TGF-ß1 modulates early chondrogenesis by primarily increasing the number of progenitor cells arising from the subchondral bone marrow compartment via the SDF-1/chemokine (CXC motif) receptor 4 pathway, their proliferation, differentiation, and extracellular matrix deposition in specific topographic patterns, highlighting the pivotal role played by TGF-ß1 during this crucial phase.-Asen, A.-K., Goebel, L., Rey-Rico, A., Sohier, J., Zurakowski, D., Cucchiarini, M., Madry, H. Sustained spatiotemporal release of TGF-ß1 confers enhanced very early chondrogenic differentiation during osteochondral repair in specific topographic patterns.

Reliable landmarks for precise topographical analyses of pathological structural changes of the ovine tibial plateau in 2-D and 3-D subspaces.

Selecting identical topographical locations to analyse pathological structural changes of the osteochondral unit in translational models remains difficult. The specific aim of the study was to provide objectively defined reference points on the ovine tibial plateau based on 2-D sections of micro-CT images useful for reproducible sample harvesting and as standardized landmarks for landmark-based 3-D image registration. We propose 5 reference points, 11 reference lines and 12 subregions that are detectable macroscopically and on 2-D micro-CT sections. Their value was confirmed applying landmark-based rigid and affine 3-D registration methods. Intra- and interobserver comparison showed high reliabilities, and constant positions (standard errors < 1%). Spatial patterns of the thicknesses of the articular cartilage and subchondral bone plate were revealed by measurements in 96 individual points of the tibial plateau. As a case study, pathological phenomena 6 months following OA induction in vivo such as osteophytes and areas of OA development were mapped to the individual subregions. These new reference points and subregions are directly identifiable on tibial plateau specimens or macroscopic images, enabling a precise topographical location of pathological structural changes of the osteochondral unit in both 2-D and 3-D subspaces in a region-appropriate fashion relevant for translational investigations.

Early loss of subchondral bone following microfracture is counteracted by bone marrow aspirate in a translational model of osteochondral repair.

Microfracture of cartilage defects may induce alterations of the subchondral bone in the mid- and long-term, yet very little is known about their onset. Possibly, these changes may be avoided by an enhanced microfracture technique with additional application of bone marrow aspirate. In this study, full-thickness chondral defects in the knee joints of minipigs were either treated with (1) debridement down to the subchondral bone plate alone, (2) debridement with microfracture, or (3) microfracture with additional application of bone marrow aspirate. At 4 weeks after microfracture, the loss of subchondral bone below the defects largely exceeded the original microfracture holes. Of note, a significant increase of osteoclast density was identified in defects treated with microfracture alone compared with debridement only. Both changes were significantly counteracted by the adjunct treatment with bone marrow. Debridement and microfracture without or with bone marrow were equivalent regarding the early cartilage repair. These data suggest that microfracture induced a substantial early resorption of the subchondral bone and also highlight the potential value of bone marrow aspirate as an adjunct to counteract these alterations. Clinical studies are warranted to further elucidate early events of osteochondral repair and the effect of enhanced microfracture techniques.

Surgical anatomy of medial open-wedge high tibial osteotomy: crucial steps and pitfalls.

PURPOSE: To give an overview of the basic knowledge of the functional surgical anatomy of the proximal lower leg and the popliteal region relevant to medial high tibial osteotomy (HTO) as key anatomical structures in spatial relation to the popliteal region and the proximal tibiofibular joint are usually not directly visible and thus escape a direct inspection. METHODS: The surgical anatomy of the human proximal lower leg and its relevance for HTO are illustrated with a special emphasis on the individual steps of the operation involving creation of the osteotomy planes and plate fixation. RESULTS: The posteriorly located popliteal neurovascular bundle, but also lateral structures such as the peroneal nerve, the head of the fibula and the lateral collateral ligament must be protected from the instruments used for osteotomy. Neither positioning the knee joint in flexion, nor the posterior thin muscle layer of the popliteal muscle offers adequate protection of the popliteal neurovascular bundle when performing the osteotomy. Tactile feedback through a loss-of-resistance when the opposite cortex is perforated is only possible when sawing and drilling is performed in a pounding fashion. Kirschner wires with a proximal thread, therefore, always need to be introduced under fluoroscopic control. Due to anatomy of the tibial head, the tibial slope may increase inadvertently. CONCLUSIONS: Enhanced surgical knowledge of anatomical structures that are at a potential risk during the different steps of osteotomy or plate fixation will help to avoid possible injuries. LEVEL OF EVIDENCE: Expert opinion, Level V.

A novel algorithm for a precise analysis of subchondral bone alterations.

Subchondral bone alterations are emerging as considerable clinical problems associated with articular cartilage repair. Their analysis exposes a pattern of variable changes, including intra-lesional osteophytes, residual microfracture holes, peri-hole bone resorption, and subchondral bone cysts. A precise distinction between them is becoming increasingly important. Here, we present a tailored algorithm based on continuous data to analyse subchondral bone changes using micro-CT images, allowing for a clear definition of each entity. We evaluated this algorithm using data sets originating from two large animal models of osteochondral repair. Intra-lesional osteophytes were detected in 3 of 10 defects in the minipig and in 4 of 5 defects in the sheep model. Peri-hole bone resorption was found in 22 of 30 microfracture holes in the minipig and in 17 of 30 microfracture holes in the sheep model. Subchondral bone cysts appeared in 1 microfracture hole in the minipig and in 5 microfracture holes in the sheep model (n = 30 holes each). Calculation of inter-rater agreement (90% agreement) and Cohen's kappa (kappa = 0.874) revealed that the novel algorithm is highly reliable, reproducible, and valid. Comparison analysis with the best existing semi-quantitative evaluation method was also performed, supporting the enhanced precision of this algorithm.

Comprehensive analysis of translational osteochondral repair: Focus on the histological assessment.

Articular cartilage guarantees for an optimal functioning of diarthrodial joints by providing a gliding surface for smooth articulation, weight distribution, and shock absorbing while the subchondral bone plays a crucial role in its biomechanical and nutritive support. Both tissues together form the osteochondral unit. The structural assessment of the osteochondral unit is now considered the key standard procedure for evaluating articular cartilage repair in translational animal models. The aim of this review is to give a detailed overview of the different methods for a comprehensive evaluation of osteochondral repair. The main focus is on the histological assessment as the gold standard, together with immunohistochemistry, and polarized light microscopy. Additionally, standards of macroscopic, non-destructive imaging such as high resolution MRI and micro-CT, biochemical, and molecular biological evaluations are addressed. Potential pitfalls of analysis are outlined. A second focus is to suggest recommendations for osteochondral evaluation.