Bilateral Anterior Cruciate Ligament Transection Research Articles

Prophylactic administration of miR-451 inhibitor decreases osteoarthritis severity in rats

Transfection of chondrocytes with microRNA-451(miR-451), present in growth zone cartilage of the growth plate, upregulates production of enzymes association with extracellular matrix degradation. miR-451 is also present in articular cartilage and exacerbates IL-1β effects in articular chondrocytes. Moreover, when osteoarthritis (OA) was induced in Sprague Dawley rats via bilateral anterior cruciate ligament transection (ACLT), miR-451 expression was increased in OA cartilage compared to control, suggesting its inhibition might be used to prevent or treat OA. To examine the prophylactic and therapeutic potential of inhibiting miR-451, we evaluated treatment with miR-451 power inhibitor (451-PI) at the onset of joint trauma and treatment after OA had developed. The prophylactic animal cohort received twice-weekly intra-articular injections of either 451-PI or a negative control (NC-PI) beginning on post-surgical day 3. OA was allowed to develop for 24 days in the therapeutic cohort before beginning injections. All rats were killed on day 45. Micro-CT, histomorphometrics, OARSI scoring, and muscle force testing were performed on samples. 451-PI mitigated OA progression compared to NC-PI limbs in the prophylactic cohort based on histomorphometric analysis and OARSI scoring, but no differences were detected by micro-CT. 451-PI treatment beginning 24 days post-surgery was not able to reduce OA severity. Prophylactic administration of 451-PI mitigates OA progression in a post-trauma ACLT rat model supporting its potential to prevent OA development following an ACLT injury clinically.

Pulsed electromagnetic field alleviates synovitis and inhibits the NLRP3/Caspase-1/GSDMD signaling pathway in osteoarthritis rats

ABSTRACT Low-grade inflammation is a key mediator of the pathogenesis of Osteoarthritis (OA). Pulsed electromagnetic field (PEMF) can improve the symptoms of OA and potentially acts as an anti-inflammatory. The aim of this study was to investigate the effect of the PEMF on OA and its relationship with the NLRP3/Caspase-1/GSDMD signaling pathway.18 Three-month-old Sprague-Dawley (SD) rats were randomly divided into three groups (n = 6 per group): 1) OA group, 2) OA+PEMF group (OA with PEMF exposure), 3) Control group (sham operation with placebo PEMF). Rats in the OA and OA+PEMF groups were subjected to bilateral anterior cruciate ligament transection and ovariectomy. PEMF scheme: Pulse waveform, 3.82 mT, 8 Hz, 40 min/day, 5 days a week, for 12 weeks. The expression levels of NLRP3, Caspase-1, GSDMD, IL-1β, and MMP-13 were detected by qRT-PCR and Western blot. The pathological structures of OA were monitored with Safranin O/fast green staining and hematoxylin eosin staining. Our results showed that PEMF alleviated the degree of inflammation and degeneration of cartilage in rats with OA, based on the histopathological changes and decline of the expression of IL-1β and MMP-13. Moreover, the over-expression of NLRP3, Caspase-1, and GSDMD in the cartilage of the OA rats decreased after PEMF treatment. These results suggested that PEMF could be a highly promising noninvasive strategy to slow down the progression of OA and inhibition of the NLRP3/Caspase-1/GSDMD signaling pathway might be involved in the beneficial effect of PEMF.

Regulation of inflammatory and catabolic responses to IL-1β in rat articular chondrocytes by microRNAs miR-122 and miR-451

miR-122 stimulates proliferation of growth plate chondrocytes whereas miR-451 stimulates terminal differentiation and matrix turnover. Here, we examined the potential of these microRNA as regulators of articular chondrocytes using an in vitro model of osteoarthritis. miR-122 and miR-451 presence in rat articular cartilage was assessed using the anterior cruciate ligament transection model of OA. In vitro testing used first passage rat articular chondrocytes (rArCs) that were transfected with lipofectamine (Lipo) and miR-122 or miR-451 for 24-h, then treated with 10ng/mL IL-1β in order to mimic an osteoarthritic environment. Conditioned media were collected and MMP13, PGE2 and OA-related cytokines were measured. Matrix vesicles were collected from cell layer lysates using ultra-centrifugation. Cells were treated with miR-122 or miR-451 inhibitors to verify miR-specific effects. Both miR-122 and miR-451 were increased in the OA articular cartilage compared to healthy tissue; rArCs expressed both microRNAs in MVs. miR-122 prevented IL-1β-dependent increases in MMP-13 and PGE2, whereas miR-451 significantly increased the IL-1β effect. Multiplex data indicated that miR-122 reduced the stimulatory effect of IL-1β on IL-1α, IL-2, Il-4, IL-6, GM-CSF, MIP-1A, RANTES and VEGF. In contrast, IL-2, IL-4, IL-6, GM-CSF, and MIP-1A were increased by miR-451 while VEGF was decreased. Inhibiting miR-122 exacerbated the response to IL-1β indicating endogenous levels of miR-122 were present. There were no differences in MMP-13 or PGE2 with miR-451 Locked Nucleic Acid (LNA) inhibitor treatment. Both miRs were elevated in OA in a rat bilateral anterior cruciate ligament transection (ACLT) model. miR-122 prevented, while miR-451 exacerbated the effects of IL-1β on rArCs.

Retracted] Differential miRNAomics of the synovial membrane in knee osteoarthritis induced by bilateral anterior cruciate ligament transection in rats.

Following the publication of the above article, a number of errors were identified in the paper, and after having consulted with the editor of Molecular Medicine Reports, a corrigendum was published last year ("[Corrigendum] Differential miRNAomics of the synovial membrane in knee osteoarthritis induced by bilateral anterior cruciate ligament transection in rats." Zhou J, Zhao Y, Wu G, Lin B, Li Z and Liu X. Mol Med Rep 20: 5363, 2019). However, following publication of the above corrigendum, the paper was re‑examined by the authors, and additional errors were identified; therefore, the authors are going to retract this paper from the publication. All the authors agree to this retraction, and apologize to the Editor of Molecular Medicine Reports and to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 18: 4051‑4057, 2018; DOI: 10.3892/mmr.2018.9385].

Corrigendum] Differential miRNAomics of the synovial membrane in knee osteoarthritis induced by bilateral anterior cruciate ligament transection in rats.

Following the publication of the above article, an interested reader drew to our attention that, in Fig.4A, in which the authors had presented a western blot image depicting protein expression of IL‑18, IL‑1β, NLRP3 and β‑actin from synovial tissue lysates from osteoarthritic rats, upon close examination of the figure a striking similarity was noted between the bands shown for the IL‑18‑ and NLRP3‑stained Sham group experiments, although the bands appeared in an inverted position relative to each other. Following an enquiry with the authors, they realized that they had included incorrect data for this figure; an amended version of Fig.4, showing the correct data for NLRP3, is shown opposite. Secondly, the authors have realized that, at various points throughout the paper, two miRNAs were written incorrectly: Specifically, references to an 'miR‑352' should have appeared as miR‑532, and 'miR‑233' should have been written as miR‑223. This error affected the presentation of Fig.2; therefore, a revised version of this figure, with the miRNAs correctly labelled as miR‑532 and miR‑223 respectively, is also shown opposite. Furthermore, mi‑233 should have been written as miR‑223 at the following places in the text: p.4051, right‑hand column (RHC), line10 ("Furthermore, the miR‑223‑regulated...); p.4054, Results section, left‑hand column (LHC), third subheading ("miR‑223 negatively regulates the expression of NLRP3."); and in the concluding paragraph of the Discussion on p.4056, LHC, miR‑233 should have been written as miR‑223 in all five instances where this occurred (lines4, 7, 8, 9, and 10 of this paragraph). Finally, the second sentence featured in the subsection of the Results section entitled "Expression validation of miRs by RT‑qPCR" on p.4054 contained additional errors. This sentence should have appeared as follows (changed text is highlighted in bold): "The expression of miR‑223, ‑100, ‑345, ‑130, ‑382, ‑9a and 183 were upregulated, whereas miR‑377, ‑532, ‑200b were downregulated with a fold change of ≥1.5, similar to the microarray data (Fig.2). All the authors agree to the contents of this Corrigendum, and apologize to the Editor of Molecular Medicine Reports and to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 18: 4051‑4057, 2018; DOI: 10.3892/mmr.2018.9385].

Regenerative Features of Adipose Tissue for Osteoarthritis Treatment in a Rabbit Model: Enzymatic Digestion Versus Mechanical Disruption.

Evaluating cell migration after cell-based treatment is important for several disorders, including osteoarthritis (OA), as it might influence the clinical outcome. This research explores migrating expanded-adipose stromal cells (ASCs) and adipose niches after enzymatic and mechanical processes. Bilateral anterior cruciate ligament transection induced a mild grade of OA at eight weeks in adult male New Zealand rabbits. ASCs, enzymatic stromal vascular fraction (SVF), and micro fragmented adipose tissue (MFAT) were intra-articularly injected in the knee joint. Assessments of cell viability and expression of specific markers, including CD-163 wound-healing macrophages, were done. Cell migration was explored through labelling with PKH26 dye at 7 and 30 days alongside co-localization analyses for CD-146. All cells showed good viability and high percentages of CD-90 and CD-146. CD-163 was significantly higher in MFAT compared to SVF. Distinct migratory potential and time-dependent effects were observed among cell-based treatments. At day 7, both ASCs and SVF migrated towards synovium, whereas for MFAT versus cartilage, a different migration pattern was noticed at day 30. The long-term distinct cell migration of ASCs, SVF, and MFAT open interesting clinical insights on their potential use for OA treatment. Moreover, the highest expression of CD-163 in MFAT, rather than SVF, might have an important role in directly mediating cartilage tissue repair responses.

Differential miRNAomics of the synovial membrane in knee osteoarthritis induced by bilateral anterior cruciate ligament transection in rats

The differential microRNA (miRNA) omics of the synovial membrane were investigated using a rat model of knee osteoarthritis (KOA) induced by bilateral anterior cruciate ligament transection, which produced pathological biomarkers in KOA. Sprague‑Dawley rats were randomly divided into two groups; Sham‑operated and KOA‑operated group. The KOA rats were subjected to bilateral anterior cruciate ligament transection. After 6weeks, total RNA was extracted from the knee joint synovial membrane of the rats and a microRNA (miR) microarray was performed to identify differentially expressed miRs. Subsequently, the obtained differentially expressed miRs were validated by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis. A total of 24 miRs were identified with alterations ≥1.5‑fold in the synovial membrane in the KOA‑operated group compared with the sham‑operated group, of which 4 miRs (miR‑532‑5p, ‑200b‑5p, ‑377‑3p and ‑759‑5p) were decreased and 20 miRs (miR‑382‑3p, ‑223‑3p, ‑100‑5p, ‑30d‑5p, ‑183‑5p, ‑130, ‑92b‑3p, ‑125b‑3p, ‑151‑3p, ‑155‑3p, 27a‑3p, ‑146b‑3p, ‑885‑5p, ‑352, ‑184, ‑345‑5p, ‑30a‑5p and ‑9a‑5p) were increased. Subsequently, RT‑qPCR was used to validate the expressions of miR‑223, ‑100, ‑345, ‑130, ‑382, ‑377, ‑352, ‑200b, ‑9a and ‑183, which were upregulated by a fold change of ≥1.5 in synovial membranes of KOA rats compared with shams. Furthermore, invitro miR‑223 mimic could suppress the luciferase activity of NACHT, LRR and PYD domains‑containing protein 3 (NLRP3) 3' untranslated region by detecting of dual luciferase reporter vector. Additionally, the expression of NLRP3, interleukin (IL)‑1β and IL‑18 significantly increased in the synovial membrane of KOA rats. A total of 24 different miRs were determined by comparing the miRNAomics in the synovial membrane of the KOA model rats. Furthermore, the miR‑233‑regulated NLRP3 inflammasome was implicated in synovial membrane injury, which may be an important mechanism of KOA pathogenesis.

Intra-Articular Injection of Alginate-Microencapsulated Adipose Tissue-Derived Mesenchymal Stem Cells for the Treatment of Osteoarthritis in Rabbits.

We investigated the effects of intra-articular injections of alginate-microencapsulated adipose tissue-derived mesenchymal stem cells (ASCs) during osteoarthritis (OA) development in a rabbit model of anterior cruciate ligament transection (ACLT). We induced OA in mature New Zealand white rabbits by bilateral ACLT. Stifle joints were categorised into four groups according to intra-articular injection materials. Alginate microbeads and microencapsulated ASCs were prepared using the vibrational nozzle technology. Two weeks after ACLT, the rabbits received three consecutive weekly intra-articular injections of 0.9% NaCl, alginate microbeads, ASCs, or microencapsulated ASCs, into each joint. Nine weeks after ACLT, we euthanised the rabbits and collected bilateral femoral condyles for macroscopic, histological, and immunohistochemical analyses. Macroscopic evaluation using the modified OA Research Society International (OARSI) score and total cartilage damage score showed that cartilage degradation on the femoral condyle was relatively low in the microencapsulated-ASC group. Histological analysis of the lateral femoral condyles indicated that microencapsulated ASCs had significant chondroprotective effects. Immunohistochemically, the expression of MMP-13 after the articular cartilage damage was relatively low in the microencapsulated-ASC-treated stifle joints. During the development of experimental OA, as compared to ASCs alone, intra-articular injection of microencapsulated ASCs significantly decreased the progression and extent of OA.

Local intra‑articular injection of vascular endothelial growth factor accelerates articular cartilage degeneration in rat osteoarthritis model.

In the pathophysiology of osteoarthritis (OA), articular cartilage degeneration exhibits a significant role. Vascular endothelial growth factor (VEGF) is considered to be an effective angiogenic factor and a crucial regulator of articular cartilage degeneration in the development of OA. Therefore, the present study aimed to investigate the underlying influences of exogenous VEGF on articular cartilage degeneration in OA model rat. A total of 24 male Sprague-Dawley rats were randomly allocated into 3 groups. In the normal saline (NS) and VEGF groups, animals received bilateral anterior cruciate ligament (ACL) transection to establish the OA model; at 4 weeks post-surgery, the rats received local intra-articular injections of 100 µl NS or VEGF solution, respectively, every week for 4 weeks. The Control group received neither surgery nor injections. All animals were sacrificed at 12 weeks following surgery. Prominent cartilage degeneration was observed in rats in the NS- and VEGF-injected groups. The extent and the grade of cartilage damage in the VEGF-injected group were notably more severe compared with those in the NS-treated group. Western blotting results demonstrated that the expression levels of aggrecan and type II collagen were significantly reduced in OA model rats that were treated with VEGF. In addition, the expression levels of matrix metalloproteinase (MMP)-3, MMP-9, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (a disintegrin and metalloproteinase; ADAMTS)-4, −5 and −12, type III collagen and transforming growth factor-β1 were significantly increased following VEGF administration. Results from the present study indicated that VEGF may exhibit a promoting role in the development of OA by destroying articular cartilage matrix.

Differential proteomics of the synovial membrane between bilateral and unilateral knee osteoarthritis in surgery‑induced rabbit models.

The present study investigated the differential proteomics of synovial membranes between bilateral and unilateral anterior cruciate ligament transection (ACLT) in rabbits with knee osteoarthritis (KOA), in order to elucidate the pathological biomarkers of different degrees of KOA. A total of 6New Zealand rabbits were randomly divided into groups AandB (three rabbits per group). The two groups were subjected to bilateral and unilateral ACLT, respectively. A total of 6weeks following surgery, proteins were extracted from the knee joint synovial membranes of KOA rabbits and were separated by two‑dimensional polyacrylamide gel electrophoresis. The differentially expressed proteins in the OA synovial membranes were selected for further analysis by linear ion trap‑Fourier transform ion cyclotron resonance mass spectrometry. Ten protein spots were identified to be different between the synovial membranes of the bilateral and unilateral KOA rabbits. Protein disulfide‑isomerase and creatine kinaseM‑type were identified in the unilateral KOA rabbit synovial membranes. Serum albumin (three spots), lumican, α‑2‑HS‑glycoprotein and three uncharacterized proteins were identified in the synovial membranes of the bilateral KOA rabbits. The differential proteomic expression demonstrated the different biomarkers associated with bilateral and unilateral KOA, and indicated that spontaneous and secondary KOA require diverse methods of treatment; thus the underlying mechanism of KOA requires further investigation.

Protective effect of LR-90 on articular cartilage in rabbit model of osteoarthritis

To investigate the protective effect of LR-90 on articular cartilage in rabbit model of osteoarthritis. The cultured rabbits chondrocytes were assigned to be treated with IL-1β (10ng/ml) or IL-1β (10ng/ml)+LR-90 (50 mg/L). The mRNA expression of MMP-13, ADAMTS-5, aggrecan and collagen II in chondrocytes were assessed by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). Twenty male New Zealand white rabbits underwent bilateral anterior cruciate ligament transection (ACLT) to establish a animal model of osteoarthritis. Four weeks after model established, on the basis of randomization one knee of each rabbit was treated with 50 mg/L LR-90 in normal saline (NS) (experimental group) and the other knee was treated with same volume of NS (control group), 1/week × 5. Nine weeks after ACLT all rabbits were sacrificed and the knee joints were evaluated by gross morphology and histology. The mRNA expression of IL-1β, MMP-13, ADAMTS-5, aggrecan and collagen Ⅱ in articular cartilage was analyzed by RT-PCR. Gross morphology and Mankin histological evaluation showed that the extent and grade of cartilage damage in the experimental group were less severe than those in the control group.Compared to IL-1β group, LR-90 treatment suppressed the mRNA expression of MMP-13 and ADAMTS-5, and enhanced aggrecan and collagen Ⅱ mRNA expression. Consistent with the in vitro results, the intraarticular LR-90 administration suppressed the mRNA expression of IL-1β,MMP-13 and ADAMTS-5 (all P<0.01), while enhanced mRNA expression of aggrecan and collagen Ⅱ in cartilage (all P<0.01). LR-90 protects against cartilage degradation and inhibits the progression of osteoarthritis in rabbit mode1 of osteoarthritis, which is associated with the suppressing IL-1β, MMP-13, ADAMTS-5 and promoting aggrecan and collagen Ⅱ mRNA expression in cartilage.

Combined Effect of Bilateral Ovariectomy and Anterior Cruciate Ligament Transection With Medial Meniscectomy on the Development of Osteoarthritis Model.

ObjectiveTo investigate the combined effect of bilateral ovariectomy (OVX) and anterior cruciate ligament transection (ACLT) with medial meniscectomy (MM) on the development of osteoarthritis (OA).MethodsTwenty female 15-week-old Sprague-Dawley rats were used. Five rats in each group underwent bilateral OVX (OVX group), bilateral ACLT with MM (ACLT with MM group), bilateral OVX plus ACLT with MM (OVX plus ACLT with MM group), and sham surgery (SHAM group). All the rats were subjected to treadmill running for 4 weeks. The behavioral evaluation for induction of OA used the number of rears method, and this was conducted at 1, 2, and 4 weeks post-surgery. Bone mineral density (BMD) was calculated with micro-computerized tomography images and the modified Mankin's scoring was used for the histological changes.ResultsThe number of rears in the OVX plus ACLT with MM group decreased gradually and more rapidly in the ACLT with MM group. Histologically, the OVX plus ACLT with MM group had a significantly higher modified Mankin's score than the OVX group (p=0.008) and the SHAM group (p=0.008). BMDs of the OVX plus ACLT with MM group were significantly lower than the SHAM group (p=0.002), and the ACLT with MM group (p=0.003).ConclusionWe found that bilateral OVX plus ACLT with MM induced definite OA change in terms of histology and BMD compared to bilateral OVX and ACLT with MM alone. Therefore, OVX and ACLT with MM was an appropriate degenerative OA rat model.

Pathology of the calcified zone of articular cartilage in post-traumatic osteoarthritis in rat knees.

ObjectivesThis study aimed to investigate the pathology occurring at the calcified zone of articular cartilage (CZC) in the joints afflicted with post-traumatic osteoarthritis (PTOA).MethodsRats underwent bilateral anterior cruciate ligament (ACL) transection and medial meniscectomy to induce PTOA. Sham surgery was performed on another five rats to serve as controls. The rats were euthanized after four weeks of surgery and tibial plateaus were dissected for histology. The pathology of PTOA, CZC area and the tidemark roughness at six pre-defined locations on the tibial plateaus were quantified by histomorphometry.ResultsPTOA developed in the knees, generally more severe at the medial plateau than the lateral plateau, of rats in the experimental group. The CZC area was unchanged in the PTOA joints, but the topographic variations of CZC areas that presented in the control knees were reduced in the PTOA joints. The tidemark roughness decreased in areas of the medial plateau of PTOA joints and that was inversely correlated with the Mankin’s score of PTOA pathology.ConclusionReduced tidemark roughness and unchanged CZC area differentiate PTOA from primary osteoarthritis, which is generally believed to have the opposite pathology at CZC, and may contribute to the distinct disease progression of the two entities of arthropathy.

Chondroprotective effects of high-molecular-weight cross-linked hyaluronic acid in a rabbit knee osteoarthritis model

We hypothesized that high-molecular-weight (MW) cross-linked (CL) hyaluronic acid (HA) improves joint lubrication and has an enhanced chondroprotective effect. We examined the histopathological changes and friction coefficients in osteoarthritic knee joints after injecting high-MW CL HA. A bilateral anterior cruciate ligament transection (ACLT) model in 20 Japanese white rabbits was used. From week 5 after transection, low-MW HA (0.8×10(6)Da; HA80) or high-MW CL HA (6×10(6)Da; HA600) was injected weekly into 10 right knee for 3 weeks; normal saline (NS) was injected into the 10 left knee. A sham operation was undertaken to exclude spontaneous osteoarthritis (OA) in five knees. Results were evaluated with macroscopy, histopathology (Kikuchi's score), biomechanical testing, and rheological assessment of the joint fluid viscoelasticity. Statistical analysis was performed using one-way analysis of variance with a 95% confidence interval (CI) (P<0.05). The macroscopic findings showed severely damaged cartilage in 30% of the NS group and 20% of the HA80 and HA600 groups and intact cartilage in 100% of the sham group. The histological scores and friction coefficients of the HA600 group were significantly lower than those of the NS group (P=0.007 and P=0.002, respectively). Viscoelasticity measurements of the joint fluid showed no significant differences between the three treatment groups. High-MW CL HA exerts potential chondroprotective effects and produces superior friction coefficients. Our results suggest that HA600 delays the progression of OA effectively and improves joint lubrication significantly.

Intra-articular delivery of adipose derived stromal cells attenuates osteoarthritis progression in an experimental rabbit model

IntroductionCell therapy is a rapidly growing area of research for the treatment of osteoarthritis (OA). This work is aimed to investigate the efficacy of intra-articular adipose-derived stromal cell (ASC) injection in the healing process on cartilage, synovial membrane and menisci in an experimental rabbit model.MethodsThe induction of OA was performed surgically through bilateral anterior cruciate ligament transection (ACLT) to achieve eight weeks from ACLT a mild grade of OA. A total of 2 × 106 and 6 × 106 autologous ASCs isolated from inguinal fat, expanded in vitro and suspended in 4% rabbit serum albumin (RSA) were delivered in the hind limbs; 4% RSA was used as the control. Local bio-distribution of the cells was verified by injecting chloro-methyl-benzamido-1,1'-dioctadecyl-3,3,3'3'-tetra-methyl-indo-carbocyanine per-chlorate (CM-Dil) labeled ASCs in the hind limbs. Cartilage and synovial histological sections were scored by Laverty's scoring system to assess the severity of the pathology. Protein expression of some extracellular matrix molecules (collagen I and II), catabolic (metalloproteinase-1 and -3) and inflammatory (tumor necrosis factor- α) markers were detected by immunohistochemistry. Assessments were carried out at 16 and 24 weeks.ResultsLabeled-ASCs were detected unexpectedly in the synovial membrane and medial meniscus but not in cartilage tissue at 3 and 20 days from ASC-treatment. Intra-articular ASC administration decreases OA progression and exerts a healing contribution in the treated animals in comparison to OA and 4% RSA groups.ConclusionsOur data reveal a healing capacity of ASCs in promoting cartilage and menisci repair and attenuating inflammatory events in synovial membrane inhibiting OA progression. On the basis of the local bio-distribution findings, the benefits obtained by ASC treatment could be due to a trophic mechanism of action by the release of growth factors and cytokines.

Stigmasterol blocks cartilage degradation in rabbit model of osteoarthritis.

Stigmasterol has been shown exhibit anti-osteoarthritic properties in vitro studies. However, the in vivo effects of stigmasterol on cartilage are still unclear. This study investigated the anti-osteoarthritic properties of stigmasterol on cartilage degradation in a rabbit model of osteoarthritis (OA). Twenty rabbits underwent bilateral anterior cruciate ligament transection (ACLT) to induce OA. Five rabbits were used as normal control. Two weeks after operation, the rabbits were randomly divided into two groups. Each group of 10 rabbits received intra-articular injection with 0.3 ml of stigmasterol in left knees and vehicle in right knees, once weekly. Group 1 was killed 6 weeks after ACLT and 2 were sacrificed 9 weeks after ACLT. The knee joints were assessed by gross morphology, histology and gene expression analysis. We found that expression of genes encoding matrix metalloproteinases (MMPs) was significantly higher while tissue inhibitors of metalloproteinase (TIMP)-1 was significantly lower in the both joints of the two OA groups compared to normal controls. Stigmasterol reduced the cartilage degradation as assessed by histological analysis and markedly suppressed MMPs expression both in group 1 and group 2. Our results suggest that stigmasterol may be considered as a possible therapeutical agent in the treatment of OA.

Effect of dehydroepiandrosterone on aggrecanase expression in articular cartilage in a rabbit model of osteoarthritis

To investigate the in vivo effect of dehydroepiandrosterone (DHEA) on the expression of aggrecanases and their endogenous inhibitor in a rabbit model of OA. Ten New Zealand white rabbits underwent bilateral anterior cruciate ligament transection (ACLT). One knee of each rabbit was randomly assigned to receive 100 μM DHEA dissolved in dimethylsulphoxide (DMSO) and the other was treated with DMSO only. The treatment was given once a week for 5 weeks, starting 4 weeks after transection. All rabbits were euthanized 9 weeks after ACLT treatment, and the knee joints were evaluated by gene expression analysis. Intra-articular administration of DHEA significantly reduced the gene expression of aggrecanases, while markly increasing that of tissue inhibitor of metalloproteinase-3 (TIMP-3), an endogenous inhibitor of aggrecanases. DHEA may have beneficial effects on OA by influencing the balance between aggrecanases and TIMP-3 through which DHEA may protect against OA.

The Effect of Skeletal Maturity on Functional Healing of the Anterior Cruciate Ligament

The effects of skeletal maturity on functional ligament healing are unknown. Prior studies have suggested that ligament injuries in skeletally mature animals heal with improved mechanical properties. In this study, we hypothesized that skeletally immature animals have improved functional healing compared with skeletally mature animals. Twenty-one Yucatan minipigs (eight juvenile, eight adolescent, and five adult animals) underwent bilateral anterior cruciate ligament transection. On one side, the ligament injury was left untreated to determine the intrinsic healing response as a function of age. On the contralateral side, an enhanced suture repair incorporating a collagen-platelet composite was performed. Biomechanical properties of the repairs were measured after fifteen weeks of healing, and histologic analysis was performed. Anterior cruciate ligaments from skeletally immature animals had significantly improved structural properties over those of adult animals at three months after transection in both the untreated and repair groups. Use of the enhanced suture technique provided the most improvement in the adolescent group, in which an increase of 85% in maximum load was noted with repair. The repair tissue in the adult tissue had the highest degree of hypercellularity at the fifteen-week time point. Functional ligament healing depends on the level of skeletal maturity of the animal, with immature animals having a more productive healing response than mature animals.

Trichostatin A inhibits expression of cathepsins in experimental osteoarthritis

The aim of this study was to investigate the effects of trichostatin A (TSA) on expression of cathepsins in cartilage in experimental osteoarthritis (OA). OA was induced in 18 rabbits by bilateral anterior cruciate ligament transection (ACLT). Four weeks after surgery, rabbits received intra-articular injection with TSA dissolved in the dimethylsulphoxide (DMSO) in the right knees and DMSO in the left knees once a week for 5 weeks. Rabbits were killed 7 days after the last injection. The knee joints were assessed by morphological and histological examination. Messenger RNA expression of cathepsins K, B, L, S and cystatin C was studied by real-time PCR. TSA inhibited the expression of cathepsins K, B, L, S and cystatin C accompanied with the less degradation in cartilage. The results suggest that TSA exhibits protective effects against cartilage degradation in rabbits with OA and the effects may be associated with the inhibition of cathepsins.

Effects of dehydroepiandrosterone on articular cartilage during the development of osteoarthritis.

To investigate the in vivo effects of dehydroepiandrosterone (DHEA) on knee joints during the development of experimentally induced osteoarthritis (OA). Twenty-two mature NZW rabbits underwent bilateral anterior cruciate ligament transection (ACLT) and received 0.3-ml intraarticular injections of DHEA (at a concentration of 100 microM in phosphate buffered saline) and control solution in the right and left knees, respectively, beginning 4 weeks after ACLT and continuing once weekly for 5 weeks. All animals were killed 9 weeks after surgery, and the knee joints were assessed by gross morphologic, histologic, histomorphometric, and biochemical methods. Gross morphologic inspection following India ink application showed that the right femoral condyles, which received DHEA, demonstrated less severe cartilage damage than did the contralateral condyles. The thickness, area, and roughness of the DHEA-treated femoral condyles provided evidence of a cartilage-protecting effect of DHEA following ACLT. These results were supported by gene expression analysis. Messenger RNA expression of a proinflammatory cytokine, interleukin-1beta, and catabolic enzymes, matrix metalloproteinases 1 and 3, was reduced in the cartilage of the DHEA-treated knee joints, and expression of tissue inhibitor of metalloproteinase 1 was increased. Results of the present study demonstrate a cartilage-protecting effect of DHEA during the development of OA following ACLT in a rabbit model.