Alleviation of osteoarthritis by Trichostatin A, a histone deacetylase inhibitor, in experimental osteoarthritis

This study investigated the effects of the histone deacetylase (HDAC) inhibitor trichostatin A (TSA) on cartilage degradation in an experimental model of osteoarthritis (OA). Thirty-two male New Zealand rabbits underwent unilateral anterior cruciate ligament transection (ACLT) on left knee joints to induce OA and were randomly divided into two groups (n = 16), the TSA group was injected intra-articularly with 0.3 ml TSA [250 ng/ml in the dimethylsulphoxide (DMSO)], the OA group received DSMO since 4 weeks after operation once a week for 5 weeks. Rabbits were killed seven days after the last injection. Left knee cartilage was harvested for morphological, histological and genetic analysis. Another ten rabbits were used for normal control and received no injection. The TSA group showed less cartilage degradation as compared to the OA group assessed by morphological and histological evaluation. Gene expression of matrix metalloproteinase-1 (MMP-1), MMP-3, MMP-13, and interleukin-1 (IL-1) was increased significantly in the OA group compared to the normal group. The elevated expression was reduced by TSA. Our results suggest that TSA could be considered as a potential agent for treatment for OA.     

Unstable Stifles without Clinical or Radiographic Osteoarthritis in Young Goats: An Experimental Study

Thirteen young, castrated male goats had instability of one stifle (knee joint) created by surgical transection of the cranial cruciate ligament, but did not develop any signs of osteoarthritis (OA) in treated joints when confined in limited space for 8 months. At the end of the experiment, the instability in the stifles had not improved, the joints were normal at radiographic examination, there were no signs of inflammation in the synovial membrane or joint capsule, and fibrosis in these tissues was not evident. The articular cartilage was normal both visually and histologically. This may indicate that the young age of the goats and the restricted physical activity on soft floor had prevented the expected development of OA in the experimantally operated joints. Synovial fluid volumes and proteoglycan concentration were measured in the treated and control joints in 6 of the goats. There seemed to be increased quantity of the proteoglycan aggrecan in the synovial fluid from the treated joints compared to the contralateral joints throughout the course of this study. It was concluded that the turnover of aggrecan in the articular cartilage of the treated joints may have been increased.     

Degeneration of normal articular cartilage induced by late phase osteoarthritic synovial fluid in beagle dogs

Objective

To investigate the pathogenesis of late phase osteoarthritic (OA) synovial fluid (SF) on normal articular cartilage in vivo and provide an understanding of degenerative cartilage extending in OA joint.

Methods

A random knee, each of 8 beagle dogs, received anterior cruciate ligament transection (ACLT) and was confirmed to have late phase OA degenerative changes at 24 weeks after operation. Thereafter, one random elbow of each canine was injected with autologous late phase OA knee SF. The contralateral elbow was injected with normal saline (NS) of the same volume as SF aspirated from ACLT knee. These two groups of elbows were labeled “SF” and “NS”. 8 other beagle dogs were left intact and placed in Group Control. After aseptic arthrocentesis was performed weekly on both elbows for 24 weeks, morphological changes were observed in the cartilage of the elbows, and expressions of 7 biological etiological factors of chondrocytes of the elbows were determined in Group SF, Group NS and Group Control, respectively.

Results

Morphological changes were observed in articular cartilage of the elbows in Group SF. Levels of unit area of collagen type I in the noncalcified, calcified and full zones of articular cartilage of the elbows in Group SF increased significantly. Level of unit area of collagen type III in the calcified zone of articular cartilage of the elbows in Group SF remained unchanged. Meanwhile, expressions of MMP-1 and MMP-3 of chondrocytes of the elbows in Group SF increased significantly. There was almost no difference between articular cartilage in Group NS and Group Control.

Conclusion

Based on these results, we conclude that OA degeneration of normal articular cartilage can be independently induced by late phase OA SF. Endogenous OA biological etiological factor may be one of the reasons causing degenerative cartilage extending in OA joint.     

Early and stable upregulation of collagen type II,collagen type I and YKL40 expression levels in cartilage during early experimental osteoarthritis occurs independent of joint location and histological grading

While morphologic and biochemical aspects of degenerative joint disease (osteoarthritis [OA]) have been elucidated by numerous studies, the molecular mechanisms underlying the progressive loss of articular cartilage during OA development remain largely unknown. The main focus of the present study was to gain more insight into molecular changes during the very early stages of mechanically induced cartilage degeneration and to relate molecular alterations to histological changes at distinct localizations of the joint. Studies on human articular cartilage are hampered by the difficulty of obtaining normal tissue and early-stage OA tissue, and they allow no progressive follow-up. An experimental OA model in dogs with a slow natural history of OA (Pond–Nuki model) was therefore chosen. Anterior cruciate ligament transection (ACLT) was performed on 24 skeletally mature dogs to induce joint instability resulting in OA. Samples were taken from different joint areas after 6, 12, 24 and 48 weeks, and gene expression levels of common cartilage molecules were quantified in relation to the histological grading (modified Mankin score) of adjacent tissue. Histological changes reflected early progressive degenerative OA. Soon after ACLT, chondrocytes responded to the altered mechanical conditions by significant and stable elevation of collagen type II, collagen type I and YKL40 expression, which persisted throughout the study. In contrast to the mild to moderate histological alterations, these molecular changes were not progressive and were independent of the joint localization (tibia, femur, lateral, medial) and the extent of matrix degeneration. MMP13 remained unaltered until 24 weeks, and aggrecan and tenascinC remained unaltered until 48 weeks after ACLT. These findings indicate that elevated collagen type II, collagen type I and YKL40 mRNA expression levels are early and sensitive measures of ACLT-induced joint instability independent of a certain grade of morphological cartilage degeneration. A second phase of molecular changes in OA may begin around 48 weeks after ACLT with altered expression of further genes, such as MMP13, aggrecan and tenascin. Molecular changes observed in the present study suggest that dog cartilage responds to degenerative conditions by regulating the same genes in a similar direction as that observed for chondrocytes in late human OA.     

Early and stable upregulation of collagen type II, collagen type I and YKL40 expression levels in cartilage during early experimental osteoarthritis occurs independent of joint location and histological grading

While morphologic and biochemical aspects of degenerative joint disease (osteoarthritis [OA]) have been elucidated by numerous studies, the molecular mechanisms underlying the progressive loss of articular cartilage during OA development remain largely unknown. The main focus of the present study was to gain more insight into molecular changes during the very early stages of mechanically induced cartilage degeneration and to relate molecular alterations to histological changes at distinct localizations of the joint. Studies on human articular cartilage are hampered by the difficulty of obtaining normal tissue and early-stage OA tissue, and they allow no progressive follow-up. An experimental OA model in dogs with a slow natural history of OA (Pond-Nuki model) was therefore chosen. Anterior cruciate ligament transection (ACLT) was performed on 24 skeletally mature dogs to induce joint instability resulting in OA. Samples were taken from different joint areas after 6, 12, 24 and 48 weeks, and gene expression levels of common cartilage molecules were quantified in relation to the histological grading (modified Mankin score) of adjacent tissue. Histological changes reflected early progressive degenerative OA. Soon after ACLT, chondrocytes responded to the altered mechanical conditions by significant and stable elevation of collagen type II, collagen type I and YKL40 expression, which persisted throughout the study. In contrast to the mild to moderate histological alterations, these molecular changes were not progressive and were independent of the joint localization (tibia, femur, lateral, medial) and the extent of matrix degeneration. MMP13 remained unaltered until 24 weeks, and aggrecan and tenascinC remained unaltered until 48 weeks after ACLT. These findings indicate that elevated collagen type II, collagen type I and YKL40 mRNA expression levels are early and sensitive measures of ACLT-induced joint instability independent of a certain grade of morphological cartilage degeneration. A second phase of molecular changes in OA may begin around 48 weeks after ACLT with altered expression of further genes, such as MMP13, aggrecan and tenascin. Molecular changes observed in the present study suggest that dog cartilage responds to degenerative conditions by regulating the same genes in a similar direction as that observed for chondrocytes in late human OA.     

Evaluation of cartilage damage by measuring collagen degradation products in joint extracts in a traumatic model of osteoarthritis

The objective of this work was to investigate whether collagen degradation products in protein extract from joints could provide quantitative information on cartilage damage. Osteoarthritis (OA) was surgically induced in rat knee joints. Joints were isolated 7, 14 and 28 days after surgery for protein extraction and histology. C-terminal telopeptide of type II collagen (CTX-II), CTX-I and hydroxyproline were measured in protein extracts. Matrix metalloproteinase (MMP)-2 and -9 activity was evaluated by gelatinase zymography and joint pathology was visualized by histology and immunohistochemistry. The results showed that levels of CTX-II were significantly increased in anterior cruciate ligament transection (ACLT)-operated compared with sham-operated knee joints on days 7 and 28, whereas the levels of hydroxyproline and CTX-I epitopes showed no difference. MMP activity was slightly increased in ACLT-operated joints. The CTX-II epitope was highly expressed and co-localized to damaged articular cartilage in ACLT-operated joints. We have therefore demonstrated an increased type II collagen degradation in knees after surgical induction of OA, and propose assessment of collagen degradation epitopes as a quantitative measure of cartilage damage.     

Evaluation of cartilage damage by measuring collagen degradation products in joint extracts in a traumatic model of osteoarthritis

The objective of this work was to investigate whether collagen degradation products in protein extract from joints could provide quantitative information on cartilage damage. Osteoarthritis (OA) was surgically induced in rat knee joints. Joints were isolated 7, 14 and 28 days after surgery for protein extraction and histology. C-terminal telopeptide of type II collagen (CTX-II), CTX-I and hydroxyproline were measured in protein extracts. Matrix metalloproteinase (MMP)-2 and -9 activity was evaluated by gelatinase zymography and joint pathology was visualized by histology and immunohistochemistry. The results showed that levels of CTX-II were significantly increased in anterior cruciate ligament transection (ACLT)-operated compared with sham-operated knee joints on days 7 and 28, whereas the levels of hydroxyproline and CTX-I epitopes showed no difference. MMP activity was slightly increased in ACLT-operated joints. The CTX-II epitope was highly expressed and co-localized to damaged articular cartilage in ACLT-operated joints. We have therefore demonstrated an increased type II collagen degradation in knees after surgical induction of OA, and propose assessment of collagen degradation epitopes as a quantitative measure of cartilage damage.     

Periarticular cancellous bone changes following anterior cruciate ligament injury.

To understand more fully the early bone changes in an experimental model of osteoarthrosis, we quantified periarticular bone mineral density and bone mechanical properties in anterior cruciate ligament transected (ACLX) knee joints (4, 10, 32, and 39 wk post-ACLX) compared with contralateral joints and unoperated normal joints of skeletally mature animals. Maximal stress and energy were significantly reduced in ACLX cancellous bone from the medial femoral condyles at 4 wk postinjury. All mechanical properties (e.g., yield stress and elastic modulus) declined after 4 wk and were significantly reduced at 10 wk. ACLX bone mineral density was significantly reduced at all measured time points. Ash content was significantly reduced at 10 and 32 wk. Changes in the lateral condyles were similar but less pronounced than in the medial condyles. These bony changes accompanied the earliest articular cartilage molecular changes and preceded changes in the articular cartilage gross morphology. We suggest that these early changes in bone mechanical behavior contribute to the progression of osteoarthrosis and pathogenic changes in the joint.     

Subchondral bone and ligament changes precede cartilage degradation in guinea pig osteoarthritis

There is increasing recognition that osteoarthritis (OA) is a complex disease involving the whole synovial joint, rather than the articular cartilage alone, however its aetiology and pathogenesis is not understood. Our initial studies revealed elevated turnover of bone and ligament collagen in human and mouse OA, respectively. To investigate the relative appearance of pathology in cartilage, bone and ligament, we studied the progression of spontaneous OA in the Dunkin-Hartley (DH) guinea pig knee, and compared with age-matched control Bristol Strain 2 (BS2) knees. The classical radiographic OA score of the DH knees compared to BS2 knees was 2-fold higher at 24 weeks of age. The patella perimeter and subchondral bone density was significantly greater in the DHs at 24 and 36 weeks compared to BS2. The femoral intercondylar notch width was found to be significantly lower in the DHs at 24 and 36 weeks, compared to BS2, indicating bone remodelling at the cruciate ligament (CL) insertion site. We found significantly greater laxity of the DH anterior CL at 12, 16 and 20 weeks compared to BS2. This elevated laxity was associated with increased remodelling of the CLs, based on markers of collagen turnover, and occurred prior to bone and cartilage pathology. We propose that the laxity of the CL leads to remodelling of the subchondral bone, and intercondylar notch, due to a change in load through the joint. Remodelling of the CLs and bone occurs prior to and concomitant with histopathological changes in the articular cartilage respectively, demonstrating the fundamental role of the ligament and subchondral bone in the aetiology of knee OA.     

Osteoarthritis in the Knee Joints of G?ttingen Minipigs after Resection of the Anterior Cruciate Ligament? Missing Correlation of MRI,Gene and Protein Expression with Histological Scoring

IntroductionThe Göttingen Minipig (GM) is used as large animal model in articular cartilage research. The aim of the study was to introduce osteoarthritis (OA) in the GM by resecting the anterior cruciate ligament (ACLR) according to Pond and Nuki, verified by histological and magnetic resonance imaging (MRI) scoring as well as analysis of gene and protein expression.

Materials and Methods

The eight included skeletally mature female GM were assessed after ACLR in the left and a sham operation in the right knee, which served as control. 26 weeks after surgery the knee joints were scanned using a 3-Tesla high-field MR tomography unit with a 3 T CP Large Flex Coil. Standard proton-density weighted fat saturated sequences in coronal and sagittal direction with a slice thickness of 3 mm were used. The MRI scans were assessed by two radiologists according to a modified WORMS-score, the X-rays of the knee joints by two evaluators. Osteochondral plugs with a diameter of 4mm were taken for histological examination from either the main loading zone or the macroscopic most degenerated parts of the tibia plateau or condyle respectively. The histological sections were blinded and scored by three experts according to Little et al. Gene expression analysis was performed from surrounding cartilage. Expression of adamts4, adamts5, acan, col1A1, col2, il-1ß, mmp1, mmp3, mmp13, vegf was determined by qRT-PCR. Immunohistochemical staining (IH) of Col I and II was performed. IH was scored using a 4 point grading (0—no staining; 3-intense staining).

Results and Discussion

Similar signs of OA were evident both in ACLR and sham operated knee joints with the histological scoring result of the ACLR joints with 6.48 ± 5.67 points and the sham joints with 6.86 ± 5.84 points (p = 0.7953) The MRI scoring yielded 0.34 ± 0.89 points for the ACLR and 0.03 ± 0.17 for the sham knee joints. There was no correlation between the histological and MRI scores (r = 0.10021). The gene expression profiles as well as the immunohistochemical findings showed no significant differences between ACLR and sham knee joints. In conclusion, both knee joints showed histological signs of OA after 26 weeks irrespective of whether the ACL was resected or not. As MRI results did not match the histological findings, MRI was obviously unsuitable to diagnose the OA in GM. The analysis of the expression patterns of the 10 genes could not shed light on the question, whether sham operation also induced cartilage erosion or if the degeneration was spontaneous. The modified Pond-Nuki model may be used with reservation in the adult minipig to induce an isolated osteoarthritis.     

Efficacy test of Polycan, a beta-glucan originated from Aureobasidium pullulans SM-2001, on anterior cruciate ligament transection and partial medial meniscectomy-induced-osteoarthritis rats

The object of this study was to assess the efficacy of Polycan from Aureobasidium pullulans SM-2001, which is composed mostly of beta-1,3-1,6-glucan, on osteoarthritis (OA)-induced by anterior cruciate ligament transection and partial medial meniscectomy (ACLT&PMM). Three different dosages of Polycan (85, 42.5, and 21.25 mg/kg) were orally administered once a day for 84 days to male rats a week after ACLT&PMM surgery. Changes in the circumference and maximum extension angle of each knee, and in cartilage histopathology were assessed using Mankin scores 12 weeks after Polycan administration. In addition, cartilage proliferation was evaluated using bromodeoxyuridine (BrdU). As the result of ACLT&PMM, classic OA was induced with increases in maximum extension angles, edematous knees changes, and capsule thickness, as well as decreases in chondrocyte proliferation, cartilages degenerative changes, and loss of articular cartilage. However, these changes (except for capsule thickness) were markedly inhibited in all Polycan- and diclofenac sodium-treated groups compared with OA control. Although diclofenac sodium did not influence BrdU uptake, BrdU-immunoreactive cells were increased with all dosages of Polycan, which means that Polycan treatment induced proliferation of chondrocytes in the surface articular cartilage of the tibia and femur. The results obtained in this study suggest that 84 days of continuous oral treatment of three different dosages of Polycan led to lesser degrees of articular stiffness and histological cartilage damage compared with OA controls 91 days after OA inducement, suggesting that the optimal Polycan dosage to treat OA is 42.5 mg/kg based on the present study.     

Bevacizumab,an anti-vascular endothelial growth factor antibody,inhibits osteoarthritis

Introduction

Angiogenesis is an important factor in the development of osteoarthritis (OA). We investigated the efficacy of bevacizumab, an antibody against vascular endothelial growth factor and an inhibitor of angiogenesis, in the treatment of OA using a rabbit model of anterior cruciate ligament transection.

Methods

First, we evaluated the response of gene expression and histology of the normal joint to bevacizumab treatment. Next, in a rabbit model of OA induced by anterior cruciate ligament transection, we used macroscopic and histological evaluations and real-time polymerase chain reaction (PCR) to examine the responses to intravenous (systemic) administration of bevacizumab (OAB IV group). We also investigated the efficacy of intra-articular (local) administration of bevacizumab in OA-induced rabbits (OAB IA group).

Results

Histologically, bevacizumab had no negative effect in normal joints. Bevacizumab did not increase the expression of genes for catabolic factors in the synovium, subchondral bone, or articular cartilage, but it increased the expression of collagen type 2 in the articular cartilage. Macroscopically and histologically, the OAB IV group exhibited a reduction in articular cartilage degeneration and less osteophyte formation and synovitis compared with the control group (no bevacizumab; OA group). Real-time PCR showed significantly lower expression of catabolic factors in the synovium in the OAB IV group compared with the OA group. In articular cartilage, expression levels of aggrecan, collagen type 2, and chondromodulin-1 were higher in the OAB IV group than in the OA group. Histological evaluation and assessment of pain behaviour showed a superior effect in the OAB IA group compared with the OAB IV group 12 weeks after administration of bevacizumab, even though the total dosage given to the OAB IA group was half that received by the OAB IV group.

Conclusions

Considering the dosage and potential adverse effects of bevacizumab, the local administration of bevacizumab is a more advantageous approach than systemic administration. Our results suggest that intra-articular bevacizumab may offer a new therapeutic approach for patients with post-traumatic OA.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-014-0427-y) contains supplementary material, which is available to authorized users.     

Structural and functional changes of the articular surface in a post-traumatic model of early osteoarthritis measured by atomic force microscopy

The functional integrity of the articulating cartilage surface is a critical determinant of joint health. Although a variety of techniques exist to characterize the structural changes in the tissue with osteoarthritis (OA), some with extremely high resolution, most lack the ability to detect and monitor the functional changes that accompany the structural deterioration of this essential bearing surface. Atomic force microscopy (AFM) enables the acquisition of both structural and mechanical properties of the articular cartilage surface, with up to nanoscale resolution, making it particularly useful for evaluating the functional behavior of the macromolecular network forming the cartilage surface, which disintegrates in OA.In the present study, AFM was applied to the articular cartilage surfaces from six pairs of canine knee joints with post-traumatic OA. Microstructure (RMS roughness) and micromechanics (dynamic indentation modulus, E^?) of medial femoral condyle cartilages were compared between contralateral controls and cruciate-transected knee joints, which develop early signs of OA by three months after surgery.Results reveal a significant increase in RMS roughness and a significant four-fold decrease in E^? in cartilages from cruciate-transected joints versus contralateral controls. Compared to previous reports of changes in bulk mechanics, AFM was considerably more sensitive at detecting early cartilage changes due to cruciate-deficiency. The use of AFM in this study provides important new information on early changes in the natural history of OA because of its ability to sensitively detect and measure local structural and functional changes of the articular cartilage surface, the presumptive site of osteoarthritic initiation.     

Combined use of adipose derived stem cells and TGF-β3 microspheres promotes articular cartilage regeneration in vivo

We investigated enhancement of articular cartilage regeneration using a combination of human adipose derived stem cells (hADSCs) and TGF-β3 microspheres (MS) in vivo. Poly-lactic-co-glycolic acid (PLGA)MS were prepared using a solid/oil/water emulsion solvent evaporation-extraction method. The morphology of the MS was evaluated by scanning electron microscopy (SEM). The release characteristic of the TGF-β3 MS was evaluated. A New Zealand rabbit model for experimental osteoarthritis (OA) was established using the anterior medial meniscus excision method. Thirty OA rabbits were divided randomly into three groups according to different treatments of the right knee joints on day 7 after surgery: hADSCs/MS group received injection of both hADSCs and TGF-β3 MS; hADSCs group was injected with hADSCs; control group was injected with normal saline. Gross observation, histological staining and RT-PCR for collagen II and aggrecan) were used to assess the severity of OA and for evaluating the effect of combined use of hADSCs and TGF-β3 MS on articular cartilage regeneration in vivo. The MS were spherical with a smooth surface and the average diameter was 28 ± 2.3 µm. The encapsulation efficiency test showed that 73.8 ± 2.9% of TGF-β3 were encapsulated in the MS. The release of TGF- β3 lasted for at least 30 days. At both 6 and 12 weeks after injection, three groups exhibited different degrees of OA. Histological analysis showed that the hADSCs/MS group exhibited less OA than the hADSCs group, and the control group exhibited the most severe OA. Real-time RT-PCR showed that the gene expression of both collagen II and aggrecan were significantly up-regulated in the hADSCs/MS group. At 12 weeks after injection, the hADSCs/MS group also exhibited less OA than the other two groups. Combined use of hADSCs and TGF-β3 MS promoted articular cartilage regeneration in rabbit OA models.     

Jellyfish mucin may have potential disease-modifying effects on osteoarthritis

Background  

We aimed to study the effects of intra-articular injection of jellyfish mucin (qniumucin) on articular cartilage degeneration in a model of osteoarthritis (OA) created in rabbit knees by resection of the anterior cruciate ligament. Qniumucin was extracted from Aurelia aurita (moon jellyfish) and Stomolophus nomurai (Nomura's jellyfish) and purified by ion exchange chromatography. The OA model used 36 knees in 18 Japanese white rabbits. Purified qniumucin extracts from S. nomurai or A. aurita were used at 1 mg/ml. Rabbits were divided into four groups: a control (C) group injected with saline; a hyaluronic acid (HA)-only group (H group); two qniumucin-only groups (M groups); and two qniumucin + HA groups (MH groups). One milligram of each solution was injected intra-articularly once a week for 5 consecutive weeks, starting from 4 weeks after surgery. Ten weeks after surgery, the articular cartilage was evaluated macroscopically and histologically.     

MMP13 is a critical target gene during the progression of osteoarthritis

Introduction

Osteoarthritis (OA) is a degenerative joint disease affecting a large population of people. The mechanism of this highly prevalent disease is not fully understood. Currently there is no effective disease-modifying treatment for OA. The purpose of this study was two-fold: 1) to investigate the role of MMP13 in the development of OA; and 2) to evaluate the efficacy of the MMP13 inhibitor CL82198 as a pharmacologic treatment for preventing OA progression.

Methods

To investigate the role of the endogenous Mmp13 gene in OA development, tamoxifen was administered to two-week-old Col2CreER;Mmp13^fx/fx (Mmp13^Col2ER) and Cre-negative control mice for five days. OA was induced by meniscal-ligamentous injury (MLI) when the mice were 10 weeks old and MLI or sham-operated joints were harvested 4, 8, 12, or 16 weeks after surgery. To evaluate the efficacy of CL82198, MLI surgery was performed on 10-week-old wild type mice. CL82198 or saline was administered to the mice daily beginning immediately after the surgery for up to 16 weeks. The joint tissues collected from both experiments were evaluated by cartilage grading, histology/histomorphometry, immunohistochemistry (IHC), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The ability of CL82198 to inhibit MMP13 activity in vitro was confirmed by ELISA.

Results

The OA progression was decelerated in Mmp13^Col2ER mice 8, 12, and 16 weeks post-surgery. Cartilage grading by blinded observers confirmed decreased articular cartilage degeneration in Mmp13^Col2ER mice at 8, 12 and 16 weeks compared to Cre-negative mice. Histomorphometric analysis demonstrated that Mmp13^Col2ER mice had a higher articular cartilage area and thickness at 12 and 16 weeks post-surgery compared to the control mice. Results of IHC revealed greater type II collagen and proteoglycan expression in Mmp13^Col2ER mice. Chondrocyte apoptosis, as determined by TUNEL staining, was higher in control mice compared to Mmp13^Col2ER mice. CL82198 inhibited MMP13 activity in conditioned media from vehicle (> 85%) or bone morphogenetic protein 2 (BMP2)-treated (> 90%) primary murine sternal chondrocytes. Intraperitoneal injection of CL82198 decelerated MLI-induced OA progression, increased type II collagen and proteoglycan levels, and inhibited chondrocyte apoptosis compared to saline treatment as determined by OA grading, histology, histomorphometry, IHC, and TUNEL staining, respectively.

Conclusions

Mmp13 is critical for OA progression and pharmacologic inhibition of MMP13 is an effective strategy to decelerate articular cartilage loss in a murine model of injury-induced knee OA.     

重组人中期因子midkine对大鼠膝关节软骨部分损伤的修复作用

目的:通过外源注射不同剂量的重组人中期因子midkine(rhMK),研究其对大鼠膝关节软骨部分损伤的修复作用。方法:雄性SD大鼠双侧膝关节建立软骨部分损伤的动物模型,术后24小时分别向关节腔内注射生理盐水或rhMK (20μg/kg、60μg/kg、180μg/kg)。于术后8周将大鼠全部处死,取材进行组织学观察,从而确定最佳注射剂量;在药代动力学研究中,按最佳注射剂量向正常大鼠膝关节腔内注射rhMK,分别于注射后1小时、1天、3天、6天、9天、12天和15天处死大鼠,检测膝关节软骨组织中rhMK的含量。结果:不同剂量的重组蛋白对膝关节软骨部分损伤均有不同程度的修复作用,其中180μg/kg的剂量效果最佳;以180μg/kg的剂量向正常大鼠膝关节腔内注射rhMK后,经过Kinetica5.0药代动力学软件拟合后,计算得rhMK在软骨组织中的消除相半衰期为8.69天。结论:rhMK对大鼠膝关节软骨部分损伤有明显的修复作用,最佳注射剂量为180μg/kg,最佳注射时间间隔为8天。     

Proteoglycan 4 downregulation in a sheep meniscectomy model of early osteoarthritis

Osteoarthritis is a disease of multifactorial aetiology characterised by progressive breakdown of articular cartilage. In the early stages of the disease, changes become apparent in the superficial zone of articular cartilage, including fibrillation and fissuring. Normally, a monolayer of lubricating molecules is adsorbed on the surface of cartilage and contributes to the minimal friction and wear properties of synovial joints. Proteoglycan 4 is the lubricating glycoprotein believed to be primarily responsible for this boundary lubrication. Here we have used an established ovine meniscectomy model of osteoarthritis, in which typical degenerative changes are observed in the operated knee joints at three months after surgery, to evaluate alterations in proteoglycan 4 expression and localisation in the early phases of the disease. In normal control joints, proteoglycan 4 was immunolocalised in the superficial zone of cartilage, particularly in those regions of the knee joint covered by a meniscus. After the onset of early osteoarthritis, we demonstrated a loss of cellular proteoglycan 4 immunostaining in degenerative articular cartilage, accompanied by a significant (p < 0.01) decrease in corresponding mRNA levels. Early loss of proteoglycan 4 from the cartilage surface in association with a decrease in its expression by superficial-zone chondrocytes might have a role in the pathogenesis of osteoarthritis.     

Alterative effects of an oral alginate extract on experimental rabbit osteoarthritis

Background

Osteoarthritis (OA) is a common joint disease that causes disabilities in elderly. However, few agents with high efficacy and low side effects have been developed to treat OA. In this study, we evaluated the effects of the alginate extract named CTX in OA cell and rabbit models.

Results

CTX was formulated by hydrolyzing sodium alginate polymers with alginate lyase and then mixing with pectin. HPLC was used to analyze the CTX content. Human chondrosarcoma SW1353 cells treated with interleukin-1β were used as OA model cells to investigate the effects of CTX on chondrocyte inflammation and anabolism. CTX at concentrations up to 1000 μg/ml exerted low cytotoxicity. It inhibited the gene expression of proinflammatory matrix metalloproteinases (MMPs) including MMP1, MMP3 and MMP13 in a dose-dependent manner and increased the mRNA level of aggrecan, the major proteoglycan in articular cartilage, at 1000 μg/ml. Thirteen-week-old New Zealand White rabbits underwent a surgical anterior cruciate ligament transection and were orally treated with normal saline, glucosamine or CTX for up to 7 weeks. Examinations of the rabbit femur and tibia samples demonstrated that the rabbits taking oral CTX at a dosage of 30 mg/kg/day suffered lesser degrees of articular stiffness and histological cartilage damage than the control rabbits.

Conclusions

The gene expression profiles in the cell and the examinations done on the rabbit cartilage suggest that the alginate extract CTX is a pharmaco-therapeutic agent applicable for OA therapy.