Joint Cartilage Regeneration Research Articles

Application of Injectable Hydrogels as Delivery Systems in Osteoarthritis and Rheumatoid Arthritis.

Osteoarthritis and rheumatoid arthritis, though etiologically distinct, are both inflammatory joint diseases that cause progressive joint injury, chronic pain, and loss of function. Therefore, long-term treatment with a focus on relieving symptoms is needed. At present, the primary treatment for arthritis is drug therapy, both oral and intravenous. Although significant progress has been achieved for these treatment methods in alleviating symptoms, certain prominent drawbacks such as the substantial side effects and limited absorption of medications call for an urgent need for improved drug delivery methods. Injected hydrogels can be used as a delivery system to deliver drugs to the joint cavity in a controlled manner and continuously release them, thereby enhancing drug retention in the joint cavity to improve therapeutic effectiveness, which is attributed to the desirable attributes of the delivery system such as low immunogenicity, good biodegradability and biocompatibility. This review summarizes the types of injectable hydrogels and analyzes their applications as delivery systems in arthritis treatment. We also explored how hydrogels counteract inflammation, bone and cartilage degradation, and oxidative stress, while promoting joint cartilage regeneration in the treatment of osteoarthritis (OA) and rheumatoid arthritis (RA). This review also highlights new approaches to developing injectable hydrogels as delivery systems for OA and RA.

Comparative Efficacy of Endogenous Stem Cells Recruiting Hydrogels and Stem Cell-loaded Hydrogels in Knee Cartilage Regeneration: A Meta- analysis.

Cartilage defects remain a challenge in diseases such as osteoarthritis (OA) and fractures. Scientists have explored the use of hydrogels in conjunction with stem cell technology as a tissue engineering method to treat cartilage defects in joints. In recent years, research into hydrogels containing stem cell technology for cartilage repair has mainly focused on two categories: stem cell-loaded hydrogels and endogenous stem cell recruiting hydrogels. The latter, utilizing cell-free products, represents a novel concept with several advantages, including easier dose standardization, wider sources, and simpler storage. This meta-analysis aims to assess and compare the therapeutic effects of endogenous stem cell recruiting hydrogels and stem cell-loaded hydrogels in promoting articular cartilage regeneration in animal models, with the goal of exploring endogenous stem cell recruiting hydrogels as a promising replacement therapy for knee cartilage regeneration in preclinical animal studies. We systematically searched PubMed, Web of Science, Cochrane Library, and Embase until January 2023 using key words related to stem cells, cartilage regeneration and hydrogel. A random-effects meta-analysis was performed to evaluate the therapeutic effect on newborn cartilage formation. Stratified analyses were also carried out by independently classifying trials according to similar characteristics. The level of evidence was determined using the GRADE method. Twenty-eight studies satisfied the inclusion criteria. Comprehensive analyses revealed that the use of endogenous stem cell recruiting hydrogels significantly promoted the formation of new cartilage in the knee joint, as evidenced by the histological score (3.77, 95% CI 2.40, 5.15; p < 0.0001) and the International Cartilage Repair Society (ICRS) macroscopic score (3.00, 95% CI 1.83, 4.18; p = 0.04), compared with the control group. The stem cell-loaded hydrogels also increased cartilage regeneration in the knee with the histological score (3.13, 95% CI 2.22, 4.04; p = 0.02) and the ICRS macroscopic score (2.49, 95% CI 1.16, 3.82; p = 0.03) in comparison to the control. Significant heterogeneity between studies was observed, and further stratified and sensitivity analyses identified the transplant site and modelling method as the sources of heterogeneity. The current study indicates that both endogenous stem cell recruiting hydrogels and stem cell loaded hydrogels can effectively promote knee joint cartilage regeneration in animal trials.

FGF-stimulated tendon cells embrace a chondrogenic fate with BMP7 in newt tissue culture.

Newts can regenerate functional elbow joints after amputation at the joint level. Previous studies have suggested the potential contribution of cells from residual tendon tissues to joint cartilage regeneration. A serum-free tissue culture system for tendons was established to explore cell dynamics during joint regeneration. Culturing isolated tendons in this system, stimulated by regeneration-related factors, such as fibroblast growth factor (FGF) and platelet-derived growth factor, led to robust cell migration and proliferation. Moreover, cells proliferating in an FGF-rich environment differentiated into Sox9-positive chondrocytes upon BMP7 introduction. These findings suggest that FGF-stimulated cells from tendons may aid in joint cartilage regeneration during functional elbow joint regeneration in newts.

New composition of hyaluronic acid and new effectiveness

Hyaluronic acid is currently one of the most universal and comprehensive tools of modern medicine. Intensive research is still being carried out to demonstrate further new possibilities of its use in the prevention and treatment of diseases and injuries of the musculoskeletal system, because HA is one of the few medicinal substances characterized not only by high therapeutic effectiveness, but also by high safety of use for the human body. The introduction of hybrid complexes of high and low molecular weight hyaluronic acid (H/L-HA) has increased the clinical utility of hyaluronic preparations by increasing the viscoelastic properties, increasing the anti-inflammatory and chondroprotective effects and thermodynamic stabilization of the preparation, which determines the extension of its half-life. Thus, it became more effective and useful in non-surgical treatment of osteoarthritis. Looking at the effectiveness of hybrid complexes, it is worth trying and using in your practice, for example, Sinodrial, which, in addition to the characteristics of the hybrid, has amino acids in its composition, which can also directly affect the production of collagen and the regeneration of joint cartilage.

Autologous Adipose-Derived Tissue Stromal Vascular Fraction (AD-tSVF) for Knee Osteoarthritis.

Adipose tissue contains adult mesenchymal stem cells that may modulate the metabolism when applied to other tissues. Stromal vascular fraction (SVF) can be isolated from adipose tissue mechanically and/or enzymatically. SVF was recently used to decrease the pain and improve the function of knee osteoarthritis (OA) patients. Primary and/or secondary OA causes inflammation and degeneration in joints, and regenerative approaches that may modify the natural course of the disease are limited. SVF may modulate inflammation and initiate regeneration in joint tissues by initiating a paracrine effect. Chemokines released from SVF may slow down degeneration and stimulate regeneration in joints. In this review, we overviewed articular joint cartilage structures and functions, OA, and macro-, micro-, and nano-fat isolation techniques. Mechanic and enzymatic SVF processing techniques were summarized. Clinical outcomes of adipose tissue derived tissue SVF (AD-tSVF) were evaluated. Medical devices that can mechanically isolate AD-tSVF were listed, and publications referring to such devices were summarized. Recent review manuscripts were also systematically evaluated and included. Transferring adipose tissues and cells has its roots in plastic, reconstructive, and aesthetic surgery. Micro- and nano-fat is also transferred to other organs and tissues to stimulate regeneration as it contains regenerative cells. Minimal manipulation of the adipose tissue is recently preferred to isolate the regenerative cells without disrupting them from their natural environment. The number of patients in the follow-up studies are recently increasing. The duration of follow up is also increasing with favorable outcomes from the short- to mid-term. There are however variations for mean age and the severity of knee OA patients between studies. Positive outcomes are related to the higher number of cells in the AD-tSVF. Repetition of injections and concomitant treatments such as combining the AD-tSVF with platelet rich plasma or hyaluronan are not solidified. Good results were obtained when combined with arthroscopic debridement and micro- or nano-fracture techniques for small-sized cartilage defects. The optimum pressure applied to the tissues and cells during filtration and purification of the AD-tSVF is not specified yet. Quantitative monitoring of articular joint cartilage regeneration by ultrasound, MR, and synovial fluid analysis as well as with second-look arthroscopy could improve our current knowledge on AD-tSVF treatment in knee OA. AD-tSVF isolation techniques and technologies have the potential to improve knee OA treatment. The duration of centrifugation, filtration, washing, and purification should however be standardized. Using gravity-only for isolation and filtration could be a reasonable approach to avoid possible complications of other methodologies.

Regeneration of Jaw Joint Cartilage in Adult Zebrafish.

The poor intrinsic repair capacity of mammalian joint cartilage likely contributes to the high incidence of arthritis worldwide. Adult zebrafish can regenerate many structures that show limited or no healing capacity in mammals, including the jawbone. To test whether zebrafish can also regenerate damaged joints, we developed a surgical injury model in which the zebrafish jaw joint is destabilized via transection of the major jaw joint ligament, the interopercular–mandibular (IOM). Unilateral transection of the IOM ligament in 1-year-old fish resulted in an initial reduction of jaw joint cartilage by 14 days, with full regeneration of joint cartilage by 28 days. Joint cartilage regeneration involves the re-entry of articular chondrocytes into the cell cycle and the upregulated expression of sox10, a marker of developing chondrocytes in the embryo that becomes restricted to a subset of joint chondrocytes in adults. Genetic ablation of these sox10-expressing chondrocytes shows that they are essential for joint cartilage regeneration. To uncover the potential source of new chondrocytes during joint regeneration, we performed single-cell RNA sequencing of the uninjured adult jaw joint and identified multiple skeletal, connective tissue, and fibroblast subtypes. In particular, we uncovered a joint-specific periosteal population expressing coch and grem1a, with the jaw joint chondrocytes marked by grem1a expression during regeneration. Our findings demonstrate the capacity of zebrafish to regenerate adult joint cartilage and identify candidate cell types that can be tested for their roles in regenerative response.

Dual-crosslinked 3D printed gelatin scaffolds with potential for temporomandibular joint cartilage regeneration

A promising alternative to current treatment options for degenerative conditions of the temporomandibular joint (TMJ) is cartilage tissue engineering, using 3D printed scaffolds and mesenchymal stem cells. Gelatin, with its inherent biocompatibility and printability has been proposed as a scaffold biomaterial, but because of its thermoreversible properties, rapid degradation and inadequate strength it must be crosslinked to be stable in physiological conditions. The aim of this study was to identify non-toxic and effective crosslinking methods intended to improve the physical properties of 3D printed gelatin scaffolds for cartilage regeneration. Dehydrothermal (DHT), ribose glycation and dual crosslinking with both DHT and ribose treatments were tested. The crosslinked scaffolds were characterized by chemical, mechanical, and physical analysis. The dual-crosslinked scaffolds had the highest degree of crosslinking and the greatest resistance to hydrolytic and enzymatic degradation. Compared to the dual-crosslinked group, the ribose-crosslinked scaffolds had thinner printed strands, larger pore surface area and higher fluid uptake. The compressive modulus values were 2 kPa for ribose, 37.6 kPa for DHT and 30.9 kPa for dual-crosslinked scaffolds. None of the crosslinking methods had cytotoxic effects on the seeded rat bone marrow-derived mesenchymal stem cells (rBMSC). After 4 and 7 d, the dual-crosslinked scaffolds exhibited better cell proliferation than the other groups. Although all scaffolds supported chondrogenic differentiation of rBMSC, dual-crosslinked scaffolds demonstrated the lowest expression of the hypertrophy-related collagen 10 gene after 21 d. The results show that 3D printed gelatin scaffolds, when dually crosslinked with ribose and DHT methods, are not toxic, promote chondrogenic differentiation of rBMSC and have potential application in tissue engineering of TMJ condylar cartilage.

Regeneration of joint cartilage: perspectives and future

Los tratamientos de las lesiones del cartílago cambiaron significativamente desde la incorporación de las técnicas de biotecnología, inicialmente con el cultivo de condrocitos autólogos en un medio líquido, posteriormente con la incorporación de matrices, embebidas o no con células cultivadas. El reto continúa y esas técnicas que han mostrado unos buenos resultados clínicos deben mejorarse para establecer un protocolo que sea capaz de regenerar el tejido cartilaginoso. Desde nuestro punto de vista, los ensayos clínicos y los trabajos de investigación son primordiales, para conocer la biología articular y ser capaces de dirigir la condrogénesis.

Evolution of hydrogels for cartilage tissue engineering of the knee: A systematic review and meta-analysis of clinical studies.

In recent years, studies have boosted our knowledge about the biology and disorders of articular cartilage. In this regard, the design of hydrogel-based scaffolds has advanced to improve cartilage repair. However, the efficacy of knee cartilage repair using hydrogels remains unclear. The aim of systematic review and meta-analysis was to scrutinize the efficiency of hydrogel-based therapy in correcting cartilage defects of knee (femoral condyle, patella, tibia plateau and trochlea). The search was conducted in PubMed to gather articles published from 2004/1/1 to 2019/10/01, addressing the effects of implant of hydrogel on knee joint cartilage regeneration. The Cochrane Collaboration's tool for estimating the risk of bias was applied to check the quality of articles. The clinical data for meta-analysis was recorded using the visual analog scale (VAS), Lysholm score, WOMAC, and IKDC. The guidelines of Cochrane Handbook for Systematic Reviews of Interventions were utilized to conduct the review and meta-analysis in the RevMan 5.3 software. The search resulted in 50 clinical trials that included 2846 patients, 986 of whom received cell-based hydrogel implants while 1860 patients used hydrogel without cell. There were significant differences comparing the pain scores based on the VAS (MD: -2.97; 95% CI: -3.15 to -2.79, P<0.00001) and WOMAC (MD: -25.22; 95% CI: -31.22 to -19.22, P<0.00001) between pre- and post-treatment with hydrogels. Furthermore, there were significant improvements in the functional scores based on the IKDC total score (MD: 30.67; P<0.00001) and the Lysholm knee scale (MD: 29.26; 95% CI: 26.74 to 31.78, P<0.00001). According to the Lysholm and IKDC score and after cumulative functional analysis, there was a significant improvement in this parameter (MD: 29.25; 95% CI: 27.26 to 31.25, P<0.00001). This meta-analysis indicated clinically and statistically significant improvements in the pain score (VAS and WOMAC) and the functional score (IKDC and Lysholm) after the administration of hydrogel compared to pretreatment status. So, the current evidence shows the efficiency of hydrogel-based therapy in correcting and repairing knee cartilage defects.

Herbal Remedies as Potential in Cartilage Tissue Engineering: An Overview of New Therapeutic Approaches and Strategies.

It is estimated that by 2023, approximately 20% of the population of Western Europe and North America will suffer from a degenerative joint disease commonly known as osteoarthritis (OA). During the development of OA, pro-inflammatory cytokines are one of the major causes that drive the production of inflammatory mediators and thus of matrix-degrading enzymes. OA is a challenging disease for doctors due to the limitation of the joint cartilage’s capacity to repair itself. Though new treatment approaches, in particular with mesenchymal stem cells (MSCs) that integrate the tissue engineering (TE) of cartilage tissue, are promising, they are not only expensive but more often do not lead to the regeneration of joint cartilage. Therefore, there is an increasing need for novel, safe, and more effective alternatives to promote cartilage joint regeneration and TE. Indeed, naturally occurring phytochemical compounds (herbal remedies) have a great anti-inflammatory, anti-oxidant, and anabolic potential, and they have received much attention for the development of new therapeutic strategies for the treatment of inflammatory diseases, including the prevention of age-related OA and cartilage TE. This paper summarizes recent research on herbal remedies and their chondroinductive and chondroprotective effects on cartilage and progenitor cells, and it also emphasizes the possibilities that exist in this research area, especially with regard to the nutritional support of cartilage regeneration and TE, which may not benefit from non-steroidal anti-inflammatory drugs (NSAIDs).

Human pluripotent stem cell-derived chondroprogenitors for cartilage tissue engineering.

The cartilage of joints, such as meniscus and articular cartilage, is normally long lasting (i.e., permanent). However, once damaged, especially in large animals and humans, joint cartilage is not spontaneously repaired. Compensating the lack of repair activity by supplying cartilage-(re)forming cells, such as chondrocytes or mesenchymal stromal cells, or by transplanting a piece of normal cartilage, has been the basis of therapy for biological restoration of damaged joint cartilage. Unfortunately, current biological therapies face problems on a number of fronts. The joint cartilage is generated de novo from a specialized cell type, termed a 'joint progenitor' or 'interzone cell' during embryogenesis. Therefore, embryonic chondroprogenitors that mimic the property of joint progenitors might be the best type of cell for regenerating joint cartilage in the adult. Pluripotent stem cells (PSCs) are expected to differentiate in culture into any somatic cell type through processes that mimic embryogenesis, making human (h)PSCs a promising source of embryonic chondroprogenitors. The major research goals toward the clinical application of PSCs in joint cartilage regeneration are to (1) efficiently generate lineage-specific chondroprogenitors from hPSCs, (2) expand the chondroprogenitors to the number needed for therapy without loss of their chondrogenic activity, and (3) direct the in vivo or in vitro differentiation of the chondroprogenitors to articular or meniscal (i.e., permanent) chondrocytes rather than growth plate (i.e., transient) chondrocytes. This review is aimed at providing the current state of research toward meeting these goals. We also include our recent achievement of successful generation of "permanent-like" cartilage from long-term expandable, hPSC-derived ectomesenchymal chondroprogenitors.

Composite Silk-Extracellular Matrix Scaffolds for Enhanced Chondrogenesis of Mesenchymal Stem Cells.

This study presented a new method to fabricate SF-ECM scaffolds that potentially promote chondrogenesis of BMSCs, and open up new possibilities for using SF-ECM scaffolds as an off-the-shelf strategy for joint cartilage regeneration. It is worthy of further investigation in knee joints of animals, and beyond knee cartilage, this scaffold may also serve as an ideal biomaterial for the regeneration of other joint cartilages.

Surgical techniques of mesenchymal stimulation of hyaline cartilage reparative chondrogenesis

The main direction of surgical treatment of cartilage injuries are the surgeries that create conditions for chondrogenesis stimulation by the method of perforation of subchondral bone plate – osteoperforative methods. The modern osteoperforative method is a microfracturization of the fundus of cartilage defect. The aim. of experimental research was the study of features of clinical course of reparative chondrogenesis on the background of chondromalacia using two osteoperforative methods. Methods. Experimental researches were carried out on 45 sexually mature male rats that underwent modeling of knee joint chondromalacia. There were carried out histological examinations of cartilaginous tissue of the joint. For study of the source of oligopotent cells of mesenchymal nature immunohistochemical research was carried out. Results. The most significant advantages of joint cartilage regeneration were observed at tunnelization to the medullary cavity. The most cells-precursors of chondroblasts were localized on the border of endosteum and medullary cavity. An access of stem cells of mesenchymal nature to an injured cartilage can be ensured by tunnelization to the medullary cavity. Conclusions. Subchondral tunnalization of the defect area in conditions of chondromalacia is less effective that tunnelization of medullary cavity. Arthroscopic surgical technology of the treatment of patients with chondromalacia of articular cartilage was grounded.

Effect of low-intensity pulsed ultrasound on regeneration of joint cartilage in patients with second and third degree osteoarthritis of the knee

ObjectiveTo determine if the application of low intensity pulsed ultrasound (LIPUS) therapy has a positive effect over the cartilage repair, functional status, and reduction of pain in patients with grade 2 or 3 osteoarthrosis of the knee. DesignThis trial was an observational, before and after study without a control group, in which 10 patients (11 knees) were studied. We applied LIPUS therapy with an intensity of 0.3 W/cm2, duty cycle of 50%, giving a total of 36 J/cm2 per session during 36 sessions (3 months). The clinical measures were obtained before the first session and at the end of the 36th session, and were: cartilage thickness by the analysis of magnetic resonance images (MRI) measured by 2 rheumatologists and a radiology specialist, pain by a visual analog scale (1–10cm) and function/severity by the Lequesne index. We used the non parametric tests of Wilcoxon for comparing medians and the Spearmans rho for the correlation of the inter observer cartilage thickness measurements defining a P value of <.05 as significant. ResultsWe observed an effect on pain (VAS mean before 7.09 [2.54]; mean after 4.18 [2.22]; P=.005) and on the function/severity index (Lequesne mean before 10.55 [5.42]; mean after 5 [4.45]; P=.008). There was poor consistency regarding the cartilage thickness measures by resonance imaging between the 3 observers (2 rheumatologists and 1 radiologist) so we were not able to define the presence or absence of effect on cartilage thickness augmentation. ConclusionsLIPUS has a benefic effect over pain and functionality/severity in patients with Kellgren and Lawrence grade 2 and 3 osteoarthritis of the knee. Unfortunately in this study we did not count with a reliable measure method to conclude on its effect over cartilage thickness measured by MRI.

Biological aspects of bone, cartilage and tendon regeneration

Current orthopaedic procedures in supporting regeneration of bone, cartilage and tendon are dependant on our understanding of the molecular processes responsible for tissue repair. At present we know how to regenerate bone when physiological mechanisms of fracture repair fail [13]. Since the original description of the potential of demineralised bone matrix to induce bone at an ectopic site, it has taken more than 3 decades to bring bone morphogenetic proteins (BMPs) to clinical use. By the end of 2007 nearly 1 million patients worldwide will have been treated with BMPs for spinal fusions, non-unions, acute fractures and maxillofacial reconstruction. Use of animal models, genomics and proteomics has deciphered new mechanisms and candidate molecules for the regeneration of joint cartilage and tendons, opening new avenues in regenerative orthopaedics. This special issue reviews novel strategies in the regeneration of bone, cartilage and tendon. Bishop and Einhorn [2] describe the clinical use of recombinant BMP-2 and BMP-7. They predict that current techniques in engineering bone for restoring defined skeletal defects represent a unique opportunity for BMPs in the future. McKay and colleagues [9] review the clinical applications of BMP-2 (INFUSE Bone Graft), while Vaccaro, McKee and colleagues [14] describe the clinical applications of BMP-7 (OP-1 implant or Osigraft). The authors suggest that recombinant human BMP-based devices, when properly applied, can eliminate the need to harvest autologous bone for grafting procedures, benefiting both the surgeon and patients. Grgurevic and collaborators [6] describe molecules discovered by proteomic analysis in the plasma of patients with an acute bone fracture. The characterisation and use of potential new biomarkers, like TGF-β-induced protein IG-H3, cartilage acidic protein 1, procollagen C proteinase enhancer protein and TGF-β receptor III, for bone and cartilage regeneration are discussed. Sendak and colleagues [11] review the newly discovered role of thyroid-stimulating hormone (TSH) and follicle-stimulating hormone (FSH) in bone remodelling. It has been recently found that low doses of TSH increase bone volume and improve bone microarchitecture and strength in aged osteoporotic rats, suggesting that TSH directly affects bone remodelling in vivo [10]. Grasser and collaborators [5], by using a genomic approach, decipher the potential mechanism of bone augmentation following systemic administration of BMP-6, suggesting that IGF-1 and EGF mediate, at least in part, the effects of BMP-6 on bone, which has been recently reported [12]. The role of prostaglandin E2 receptors and their analogs in bone formation is reviewed by Li and colleagues [8], demonstrating that the PGE2 receptor is essential for restoring bone defects in animal models. Cartilage regeneration and chronic back pain due to degenerative disc disease are among major obstacles in orthopaedics. Chubinskaya and colleagues [4] summarise the role of osteogenic protein-1 (BMP-7) in animal models of osteochondral and chondral defects, osteoarthritis and degeneration in intervertebral disc cartilage. They show that OP-1 exhibits unique pro-anabolic and prominent anti-catabolic properties having a potential for treating cartilage and disc lesions. Overuse tendon disorders are common and associated with a significant morbidity. Tendon and ligament function engineering and repair are discussed in articles by Aspenberg [1] and by Hoffman and Gross [7]. Aspenberg found that the repair of subcutaneous tendon ruptures can be stimulated by a single application of one of several growth factors, including PDGF, TGF-β, IGF-1, VEGF and GDF-5,-6,-7, or by a thrombocite concentrate. The response is dependent on the mechanical microenvironment, which is crucial for the repair process. Hoffmann and Gross specifically discuss gene-therapeutic approaches using mesenchymal stem cells for a potential future application in tendon/ligament regeneration. Borovecki and colleagues [3] discuss the potential of a genomic approach in analysing bone homeostasis and diseases. Gene expression profiling studies yielded novel insights into the complex interplay of osteoblast and osteoclast regulation, paracrine and endocrine control of bone and cartilage remodelling, as well as the pathophysiology of osteoporosis and bone tumours. We believe that this issue will provide basic and clinical scientists with the state of the art knowledge in regenerative orthopaedics.

Hyaluronan Synthases in Normal and Regenerating Joint Cartilage

Repair of full thickness joint cartilage defects is within reach of routine clinical practice. The quality of regenerating hyaline cartilage, however, is difficult to assess. Synthesis of an extracellular matrix with high hyaluronan content is crucial for its metabolic and functional properties. We studied hyaluronan synthase (HAS) expression in knee joints of adult sheep as a novel cellular marker for chondrocyte function. Six house-bred Merino sheep (age 4–6 years) underwent two-stage surgery of their femoral condyles for autologous chondrocyte transplantation (ACT). First, cells were isolated from biopsies and expanded in vitro for 2 weeks using standard culture techniques. In a second session, three defects were made and either left untreated, covered with periosteal flap alone, or in combination with chondrocyte suspensions injected under the flaps. After 16 weeks, biopsies were taken from the operated knees at the defect sites and from the untreated condyle. Specimens were processed for safranin O and electron microscopy, and for immunofluorescence using three different polyclonal anti-HAS antibodies recognizing one or all of the three known mammalian HAS. Control and regenerating tissues were compared regarding their morphology and the expression of HAS, in relation to collagens type I and II, and adult cartilage proteoglycan core protein. In comparison with untreated defects or with periosteal flap alone, ACT provided a neocartilage with better-differentiated morphology. In healthy joint cartilage, about 50% of the chondrocytes expressed HAS, independent of antibody. Following ACT, a higher density of chondrocytes was observed, of which more than 75% expressed HAS, whereas the regenerates without ACT showed a lower density of HAS-expressing cells. We propose to use HAS immunofluorescence as an additional marker of matrix synthesis by chondrocytes and joint cartilage regeneration.

Joint cartilage regeneration by tissue engineering.

The research field of tissue engineering combines cells biology, biomaterial science, and surgery. Major long-term goals are tissue and organ replacement therapies using the patients' own cells. Our work is focused on the treatment of severe joint defects and on plastic surgery using in vitro engineered cartilage tissues. The practical approaches in cartilage engineering face problems with three-dimensional cell distribution or cell immobilization raising biocompatibility problems. The tissue engineering of cartilage is based on combining biocompatible cell embedding substances such as fibrin, agarose, alginate, hyaluronic acid and fiber fleece scaffolds of poly alpha-hydroxy acids (PLLA/PGLA). Different technical approaches were established: a) three-dimensional in vitro cultures of chondrocytes for the development of vital tissue transplants and b) interacting three-dimensional cultures consisting of different cell populations, such as BMP-transfected mesenchymal cells. The preshaped artificial tissue constructs were cultured in perfusion chambers to maintain a stable diffusion of nutrients during the in vitro pre-formation step. Subsequently, pre-formed tissues were implanted into nude mice and into 4 mm articular joint defects of rabbits. Transplants were found to produce cartilage typic morphological patterns and matrix. 80% of the transplants remained stable in vivo. However, 20% of the tissues are resorbed or replaced by a fibrous tissue. These results demonstrate that current artificial cartilage transplants are already feasible for plastic reconstruction. The treatment of severe joint defects, however, faces additional problems which are addressed in ongoing studies: (a) the fixation of engineered cartilage in joints, (b) the protection against chronic inflammatory degradation, and (c) the required enormous mechanical stability.

Stimulation of regeneration of joint cartilage by cod liver oil and ASD (3rd fraction)

Intracapsular injuries were made in the bone cartilage of the lower epiphysis of the femur. The experiments were performed on 20 dogs aged less than 9 months. It was found that cod liver oil and particularly ASD(3rd fraction) dissolved in cod liver oil had a marked stimulating effect on the regeneration of joint cartilage, but only when the defect extended as far as the spongy substance of the bone. Growth of blood vessels into the clot filling the wound was then accelerated; the resorption of the injured edge of bone cartilage was also intensified, thus favoring the escape of chondrocytes (from the isogenous groups), which later changed into chondroblasts.