Maximilian Rudert

A Prospective Multicenter Study on the Outcome of Type I Collagen Hydrogel–Based Autologous Chondrocyte Implantation (CaReS) for the Repair of Articular Cartilage Defects in the Knee

Background: The Cartilage Regeneration System (CaReS) is a novel matrix-associated autologous chondrocyte implantation (ACI) technique for the treatment of chondral and osteochondral lesions (Outerbridge grades III and IV). For this technology, no expansion of the chondrocytes in a monolayer culture is needed, and a homogeneous cell distribution within the gel is guaranteed. Purpose: To report a prospective multicenter study of matrix-associated ACI of the knee using a new type I collagen hydrogel (CaReS). Study Design: Case series; Level of evidence, 4. Methods: From 2003 to 2008, 116 patients (49 women and 67 men; mean age, 32.5 ± 8.9 years) had CaReS implantation of the knee in 9 different centers. On the basis of the International Cartilage Repair Society (ICRS) Cartilage Injury Evaluation Package 2000, the International Knee Documentation Committee (IKDC) score, pain score (visual analog scale [VAS]), SF-36 score, overall treatment satisfaction and the IKDC functional status were evaluated. Patient follow-up was performed at 3, 6, and 12 months after surgery and annually thereafter. Mean follow-up was 30.2 ± 17.4 months (range, 12-60 months). There were 67 defects of the medial condyle, 14 of the lateral, 22 of the patella/trochlea, and 3 of the tibial plateau, and 10 patients had 2 lesions. The mean defect size was 5.4 ± 2.4 cm2. Thirty percent of the defects were <4 cm2 and 70% were >4 cm2. Results: The IKDC score improved significantly from 42.4 ± 13.8 preoperatively to 70.5 ± 18.7 (P < .001) at latest follow-up. Global pain level significantly decreased (P < .001) from 6.7 ± 2.2 preoperatively to 3.2 ± 3.1 at latest follow-up. There also was a significant increase of both components of the SF-36 score. The overall treatment satisfaction was judged as very good or good in 88% by the surgeon and 80% by the patient. The IKDC functional knee status was grade I in 23.4%, II in 56.3%, III in 17.2%, and IV in 3.1% of the patients. Conclusion: Matrix-associated ACI employing the CaReS technology for the treatment of chondral or osteochondral defects of the knee is a safe and clinically effective treatment that yields significant functional improvement and improvement in pain level. However, further investigation is necessary to determine the long-term viability and clinical outcome of this procedure.

Techniques of cartilage growth enhancement: A review of the literature

This review attempts to present an overview of the literature pertaining to the techniques of cartilage growth enhancement. Only cartilage has an incomplete capacity for self-repair, especially of superficial defects. Full-thickness defects involving the subchondral bone can be repaired with the use of pluripotent progenitor cells from bone marrow or from transplanted perichondreum or periosteum. Bone-cartilage autografts and allografts transplanted into the cartilage defect heal primarily, but they loose their long-term biomechanical qualities because of transformation into fibrous cartilage. Cultivated human chondrocyte autografts may make cartilage healing possible in the future.

Stem cell- and growth factor-based regenerative therapies for avascular necrosis of the femoral head

Avascular necrosis (AVN) of the femoral head is a debilitating disease of multifactorial genesis, predominately affects young patients, and often leads to the development of secondary osteoarthritis. The evolving field of regenerative medicine offers promising treatment strategies using cells, biomaterial scaffolds, and bioactive factors, which might improve clinical outcome. Early stages of AVN with preserved structural integrity of the subchondral plate are accessible to retrograde surgical procedures, such as core decompression to reduce the intraosseous pressure and to induce bone remodeling. The additive application of concentrated bone marrow aspirates, ex vivo expanded mesenchymal stem cells, and osteogenic or angiogenic growth factors (or both) holds great potential to improve bone regeneration. In contrast, advanced stages of AVN with collapsed subchondral bone require an osteochondral reconstruction to preserve the physiological joint function. Analogously to strategies for osteochondral reconstruction in the knee, anterograde surgical techniques, such as osteochondral transplantation (mosaicplasty), matrix-based autologous chondrocyte implantation, or the use of acellular scaffolds alone, might preserve joint function and reduce the need for hip replacement. This review summarizes recent experimental accomplishments and initial clinical findings in the field of regenerative medicine which apply cells, growth factors, and matrices to address the clinical problem of AVN.

Indian hedgehog gene transfer is a chondrogenic inducer of human mesenchymal stem cells

IntroductionTo date, no single most-appropriate factor or delivery method has been identified for the purpose of mesenchymal stem cell (MSC)-based treatment of cartilage injury. Therefore, in this study we tested whether gene delivery of the growth factor Indian hedgehog (IHH) was able to induce chondrogenesis in human primary MSCs, and whether it was possible by such an approach to modulate the appearance of chondrogenic hypertrophy in pellet cultures in vitro.MethodsFirst-generation adenoviral vectors encoding the cDNA of the human IHH gene were created by cre-lox recombination and used alone or in combination with adenoviral vectors, bone morphogenetic protein-2 (Ad.BMP-2), or transforming growth factor beta-1 (Ad.TGF-β1) to transduce human bone-marrow derived MSCs at 5 × 102 infectious particles/cell. Thereafter, 3 × 105 cells were seeded into aggregates and cultured for 3 weeks in serum-free medium, with untransduced or marker gene transduced cultures as controls. Transgene expressions were determined by ELISA, and aggregates were analysed histologically, immunohistochemically, biochemically and by RT-PCR for chondrogenesis and hypertrophy.ResultsIHH, TGF-β1 and BMP-2 genes were equipotent inducers of chondrogenesis in primary MSCs, as evidenced by strong staining for proteoglycans, collagen type II, increased levels of glycosaminoglycan synthesis, and expression of mRNAs associated with chondrogenesis. IHH-modified aggregates, alone or in combination, also showed a tendency to progress towards hypertrophy, as judged by the expression of alkaline phosphatase and stainings for collagen type X and Annexin 5.ConclusionAs this study provides evidence for chondrogenic induction of MSC aggregates in vitro via IHH gene delivery, this technology may be efficiently employed for generating cartilaginous repair tissues in vivo.

How Frequent Is Rotational Mismatch Within 0°±10° in Kinematically Aligned Total Knee Arthroplasty?

Rotational mismatch of the tibial component on the femoral component within 0°±10° is associated with better function after mechanically aligned total knee arthroplasty (TKA). Kinematically aligned TKA has gained interest; however, the percentage of kinematically aligned TKA within 0°±10° is unknown. The authors prospectively followed all patients who underwent TKA for primary osteoarthritis between December 2011 and April 2012 (194 patients, 195 knees). Each underwent kinematically aligned TKA with manual instruments. Aligning the anteroposterior axis of the tibial component parallel to the line that bisects the oval boundary of the lateral tibial condyle set internal/external rotation. Removing bone from the posterior femoral condyles equal in thickness to the femoral component after correction for cartilage wear set internal/external rotation and anteroposterior translation of the femoral component. Rotational mismatch of the tibial component on the femoral component was determined from a computed tomography scan of the knee. Ninety-seven percent of kinematically aligned TKA with fixed-bearing components had a rotational mismatch within 0°±10° (overall range, -11° to 11°). This percentage was higher and the range narrower than the 85% of TKA within 0°±10° and the -14° to 16° range reported for mechanically aligned TKA. The use of manual instruments to kinematically aligned TKA reliably limited rotational mismatch to within 0°±10°, which has been associated with better function.

Attachment to laminin-111 facilitates transforming growth factor β-induced expression of matrix metalloproteinase-3 in synovial fibroblasts

Background: In the synovial membrane of patients with rheumatoid arthritis (RA), a strong expression of laminins and matrix degrading proteases was reported. Aim: To investigate the regulation of matrix metalloproteinases (MMPs) in synovial fibroblasts (SFs) of patients with osteoarthritis (OA) and RA by attachment to laminin-1 (LM-111) and in the presence or absence of costimulatory signals provided by transforming growth factor β (TGFβ). Methods: SFs were seeded in laminin-coated flasks and activated by addition of TGFβ. The expression of genes was investigated by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR), immunocytochemistry and ELISA, and intracellular signalling pathways by immunoblotting, and by poisoning p38MAPK by SB203580, MEK-ERK by PD98059 and SMAD2 by A-83-01. Results: Attachment of SF to LM-111 did not activate the expression of MMPs, but addition of TGFβ induced a fivefold higher expression of MMP-3. Incubation of SF on LM-111 in the presence of TGFβ induced a significant 12-fold higher expression of MMP-3 mRNA, and secretion of MMP-3 was elevated 20-fold above controls. Functional blocking of LM-111–integrin interaction reduced the laminin-activated MMP-3 expression significantly. Stimulation of SF by LM-111 and TGFβ activated the p38MAPK, ERK and SMAD2 pathways, and inhibition of these pathways by using SB203580, PD98059 or A-83-01 confirmed the involvement of these pathways in the regulation of MMP-3. Conclusion: Attachment of SF to LM-111 by itself has only minor effects on the expression of MMP-1 or MMP-3, but it facilitates the TGFβ-induced expression of MMP-3 significantly. This mode of MMP-3 induction may therefore contribute to inflammatory joint destruction in RA independent of the proinflammatory cytokines interleukin (IL)1β or tumour necrosis factor (TNF)α.

Direct bone morphogenetic protein 2 and Indian hedgehog gene transfer for articular cartilage repair using bone marrow coagulates

OBJECTIVE Bone morphogenetic protein 2 (BMP-2, encoded by BMP2) and Indian hedgehog protein (IHH, encoded by IHH) are well known regulators of chondrogenesis and chondrogenic hypertrophy. Despite being a potent chondrogenic factor BMP-2 was observed to induce chondrocyte hypertrophy in osteoarthritis (OA), growth plate cartilage and adult mesenchymal stem cells (MSCs). IHH might induce chondrogenic differentiation through different intracellular signalling pathways without inducing subsequent chondrocyte hypertrophy. The primary objective of this study is to test the efficacy of direct BMP2 and IHH gene delivery via bone marrow coagulates to influence histological repair cartilage quality in vivo. METHOD Vector-laden autologous bone marrow coagulates with 10(11) adenoviral vector particles encoding BMP2, IHH or the Green fluorescent protein (GFP) were delivered to 3.2 mm osteochondral defects in the trochlea of rabbit knees. After 13 weeks the histological repair cartilage quality was assessed using the ICRS II scoring system and the type II collagen positive area. RESULTS IHH treatment resulted in superior histological repair cartilage quality than GFP controls in all of the assessed parameters (with P < 0.05 in five of 14 assessed parameters). Results of BMP2 treatment varied substantially, including severe intralesional bone formation in two of six joints after 13 weeks. CONCLUSION IHH gene transfer is effective to improve repair cartilage quality in vivo, whereas BMP2 treatment, carried the risk intralesional bone formation. Therefore IHH protein can be considered as an attractive alternative candidate growth factor for further preclinical research and development towards improved treatments for articular cartilage defects.

Can Bone Tissue Engineering Contribute to Therapy Concepts after Resection of Musculoskeletal Sarcoma

Resection of musculoskeletal sarcoma can result in large bone defects where regeneration is needed in a quantity far beyond the normal potential of self-healing. In many cases, these defects exhibit a limited intrinsic regenerative potential due to an adjuvant therapeutic regimen, seroma, or infection. Therefore, reconstruction of these defects is still one of the most demanding procedures in orthopaedic surgery. The constraints of common treatment strategies have triggered a need for new therapeutic concepts to design and engineer unparalleled structural and functioning bone grafts. To satisfy the need for long-term repair and good clinical outcome, a paradigm shift is needed from methods to replace tissues with inert medical devices to more biological approaches that focus on the repair and reconstruction of tissue structure and function. It is within this context that the field of bone tissue engineering can offer solutions to be implemented into surgical therapy concepts after resection of bone and soft tissue sarcoma. In this paper we will discuss the implementation of tissue engineering concepts into the clinical field of orthopaedic oncology.

In situ guided tissue regeneration in musculoskeletal diseases and aging

In situ guided tissue regeneration, also addressed as in situ tissue engineering or endogenous regeneration, has a great potential for population-wide “minimal invasive” applications. During the last two decades, tissue engineering has been developed with remarkable in vitro and preclinical success but still the number of applications in clinical routine is extremely small. Moreover, the vision of population-wide applications of ex vivo tissue engineered constructs based on cells, growth and differentiation factors and scaffolds, must probably be deemed unrealistic for economic and regulation-related issues. Hence, the progress made in this respect will be mostly applicable to a fraction of post-traumatic or post-surgery situations such as big tissue defects due to tumor manifestation. Minimally invasive procedures would probably qualify for a broader application and ideally would only require off the shelf standardized products without cells. Such products should mimic the microenvironment of regenerating tissues and make use of the endogenous tissue regeneration capacities. Functionally, the chemotaxis of regenerative cells, their amplification as a transient amplifying pool and their concerted differentiation and remodeling should be addressed. This is especially important because the main target populations for such applications are the elderly and diseased. The quality of regenerative cells is impaired in such organisms and high levels of inhibitors also interfere with regeneration and healing. In metabolic bone diseases like osteoporosis, it is already known that antagonists for inhibitors such as activin and sclerostin enhance bone formation. Implementing such strategies into applications for in situ guided tissue regeneration should greatly enhance the efficacy of tailored procedures in the future.

Krüppel-like factors KLF2 and 6 and Ki-67 are direct targets of zoledronic acid in MCF-7 cells.

Bisphosphonates (BP) are used for the treatment of osteoporosis and bone metastases due to breast and prostate cancer. Recent clinical studies indicated a benefit in survival and tumor relapse with the supportive treatment of breast cancer using zoledronic acid (ZA), thus stimulating the debate about its putative anti-tumor activity in vivo. MCF-7 breast cancer cells were treated for 3 h (pulse treatment) and 72 h (permanent treatment) with ZA, and apoptosis rates and cell viability, defined as ATP content, were determined after 72 h. Permanent and pulse stimulation with ZA inhibited the viability of MCF-7 cells, which could partly be rescued by atorvastatin (Ator) pre-treatment but not by geranylgeranyl pyrophosphate (GGPP) co-treatment. Microarray analysis of ZA treated MCF-7 cells identified genes of the mevalonate pathway as significantly upregulated, which was verified by qPCR. Additionally the putative tumor suppressors krüppel-like factor 2 and 6 (KLF2 and KLF6) were markedly upregulated, while the classical proliferation marker Ki-67 was clearly downregulated. The expression of all three genes was confirmed by qPCR on mRNA level and by immunocytochemistry or Western blot staining. Expression of target genes were also analyzed in other breast (MDA-MB-231, BT-20, ZR75-1, T47D) and prostate (LNCaP, PC3) cancer cell lines by qPCR. ZA responsiveness of KLF2, KLF6 and Ki-67 could be verified in PC3 and T47D cells, KLF6 responsiveness in LNCaP and KLF2 responsiveness in MDA-MB-231 and BT-20 cells. Here we demonstrate in the apoptosis insensitive MCF-7 cell line a remarkable impact of ZA exposure on cell viability and on the regulation of putative tumor suppressors of the KLF family. The molecular mechanism involved might be the accumulation of isopentenyl pyrophosphate (IPP) and ApppI, since we could partly rescue the ZA effect by Ator pre-treatment and GGPP co-treatment. These data should stimulate further research into both the role of the mevalonate pathway and the accumulation of pyrophosphate compounds like ApppI in tumorigenesis and differentiation and their potential apart from the inhibition of mitochondrial ADP/ATP translocase and apoptosis, since such effects might well be responsible for the adjuvant ZA treatment benefit of patients suffering from breast cancer.