Modified Mankin Scale Research Articles

Postoperative malrotation of humerus shaft fracture causes degeneration of rotator cuff and cartilage

We hypothesized that postoperative malrotation of humeral shaft fractures can alter the bio-mechanical environment of the shoulder; thus, rotator cuff and cartilage degeneration could be induced. Therefore, we designed an animal experiment to evaluate the impact of malrotation deformities after minimally invasive surgery for humeral fractures on the rotator cuff and cartilage, which has rarely been described in previous studies. Twenty-four New Zealand white rabbits were randomly divided into the sham control group (A), negative control group (B) and malrotated group (C). A sham operation with surgical exposure alone was performed in group A. Humeral shaft osteotomy was performed in Group B and C. In Group B, the fractures were fixed in situ with plate -screw system. While in Group C, iatrogenic rotational deformity was created after the proximal end of the fracture being internally rotated by 20 degrees and then subsequently fixed. The animals with bone healing were sacrificed for pathological and biochemical examination. In group C, the modified Mankin scale for cartilage pathology evaluation and the modified Movin scale for tendon both showed highest score among groups with statistical significance (P < 0.05); Disordered alignment and proportion of collagen I/III of rotator cuff were confirmed with picrosirius red staining; Transmission electron microscopy also showed ultrastructural tendon damage. Immunohistochemistry showed that both MMP-1 and MMP-13 expression were significantly higher in group C than groups A and B(P < 0.05). Minimally invasive techniques for humerus shaft fracture might be cosmetically advantageous, but the consequent postoperative malrotation could increase the risk of rotator cuff and cartilage degeneration. This conclusion is supported here by primary evidence from animal experiments.

Long non-coding RNA SNHG15 is a competing endogenous RNA of miR-141-3p that prevents osteoarthritis progression by upregulating BCL2L13 expression

Increasing evidence has demonstrated that the dysregulated expression of long noncoding RNAs (lncRNAs) has important roles in the progression of osteoarthritis (OA), but the function of the lncRNA SNHG15 remains unclear. In the present study, we observed that SNHG15 was downregulated in OA cartilage tissues and IL-1β-induced chondrocytes. The lower expression of SNHG15 was negatively associated with the observed modified Mankin scale scores, extracellular matrix (ECM) degradation and chondrocyte apoptosis. Downregulated expression of SNHG15 increased chondrocyte viability and decreased chondrocyte apoptosis and ECM degradation in vitro and reduced damage to articular cartilage in vivo. Mechanistically, we demonstrated that SNHG15 overexpression promotes the expression of BCL2L13 by sponging miR-141-3p. The higher expression of miR-141-3p was negatively correlated with SNHG15 and BCL2L13 levels in OA cartilage tissues, and a positive correlation was also shown between SNHG15 and BCL2L13 levels. Furthermore, ectopic expression of miR-141-3p or knockdown of BCL2L13 expression could both reduce the effects of SNHG15 on chondrocyte proliferation, apoptosis and ECM degradation. Collectively, these findings reveal that SNHG15 inhibits OA progression by acting as an miR-141-3p sponge to promote BCL2L13 expression, suggesting that knockdown of SNHG15 expression in chondrocytes can be a potential therapeutic strategy to ameliorate OA progression.

MicroRNA-21-5p as a novel therapeutic target for osteoarthritis.

Growing evidence indicates that microRNAs (miRNA) play a critical role in the pathogenesis of OA, and overexpressing or silencing miRNA expression in OA models can contribute to the development of miRNA-based therapeutics. The objective of this study was to determine whether intra-articular injection of miRNA can inhibit OA progression. The miRNA expression profile was determined in OA cartilage tissues and controls. Functional analysis of the miRNAs on extracellular matrix degradation was performed after miRNA mimic or inhibitor transfection. Luciferase reporter assays and western blotting were employed to determine miRNA targets. To investigate the functional mechanism of miR-21-5p in OA development, miR-21-5pfl/flCol2a1-CreER and wild-type mice were subject to surgical destabilization of the medial meniscus. Therapeutically, wild-type mice undergoing surgical destabilization of the medial meniscus were treated with intra-articular injection of agomir- and antagomir-21-5p. We found that expression of miR-21-5p was significantly up-regulated in OA cartilage tissues. The articular cartilage degradation of miR-21-5p conditional knockout mice was significantly alleviated compared with that of wild-type mice in spontaneous and destabilization of the medial meniscus models. Through gain-of-function and loss-of-function studies, miR-21-5p was shown to significantly affect matrix synthesis genes expression, and chondrocyte proliferation and apoptosis. Further, fibroblast growth factor 18 (FGF18) was identified as a target of miR-21-5p. Intra-articular injection of antagomir-21-5p significantly attenuated the severity of experimental OA. Clinically, FGF18 expression level was correlated with miR-21-5p expression and a modified Mankin scale. Our findings reveal a miRNA functional pathway important for OA development, highlighting miRNA-21-5p silencing as an attractive therapeutic regimen in future clinical trials involving patients with OA.

Long non-coding RNA HOTAIR promotes osteoarthritis progression via miR-17-5p/FUT2/\u03b2-catenin axis

Osteoarthritis (OA) is a chronic joint disease and hard to cure at present. Accumulating evidence suggests long noncoding RNA-HOTAIR (lncRNA-HOTAIR) plays important role in OA progression. However, the underlying molecular mechanism of HOTAIR in OA progression has not been well elucidated. In the present study, we identified that HOTAIR level was upregulated in OA cartilage tissues. High expression of HOTAIR was correlated with modified Mankin scale, extracellular matrix (ECM) degradation and chondrocytes apoptosis. The expression of miR-17-5p was down-regulated, while alpha-1, 2 fucosyltransferase 2 (FUT2) was increased in OA progression. Luciferase reporter and RNA immunoprecipitation (RIP) assays indicated that HOTAIR could directly bind to miR-17-5p and indirectly upregulate FUT2 level. Functional investigation revealed HOTAIR and FUT2 aggravated ECM degradation and chondrocytes apoptosis, and this effect could be reversed by miR-17-5p. Altered FUT2 modulated the activity of wnt/β-catenin pathway and HOTAIR/miR-17-5p also mediated wnt/β-catenin pathway through FUT2. Collectively, our findings indicated that HOTAIR/miR-17-5p/FUT2 axis contributed to OA progression via wnt/β-catenin pathway, which might provide novel insights into the function of lncRNA-driven in OA.

MicroRNA-218-5p as a Potential Target for the Treatment of Human Osteoarthritis

Emerging evidence suggests that dysregulated microRNAs (miRNAs) play a pivotal role in osteoarthritis (OA), but the role of specific miRNAs remains unclear. Accordingly, we identified OA-associated miRNAs and functional validation of results. Here, we demonstrate that miR-218-5p is significantly upregulated in moderate and severe OA and correlates with scores on a modified Mankin scale. Through gain-of-function and loss-of-function studies, miR-218-5p was shown to significantly affect matrix synthesis gene expression and chondrocyte proliferation and apoptosis. Using SW1353 and C28/I2 cells, PIK3C2A mRNA was identified as a target of miR-218-5p. Downregulation of miR-218-5p dramatically promoted expression of PIK3C2A and its downstream target proteins, such as Akt, mTOR, S6, and 4EBP1. More importantly, OA mice exposed to a miR-218-5p inhibitor were protected from cartilage degradation and had reduced proteoglycan loss and reduced loss of articular chondrocyte cellularity compared with control mice. miR-218-5p is a novel inducer of cartilage destruction via modulation of PI3K/Akt/mTOR signaling. Inhibition of endogenous miR-218-5p expression/activity appears to be an attractive approach to OA treatment.

Downregulation of miR-221-3p contributes to IL-1β-induced cartilage degradation by directly targeting the SDF1/CXCR4 signaling pathway.

Osteoarthritis (OA) is characterized by degradation of chondrocyte extracellular matrix (ECM). Accumulating evidence suggests that microRNAs (miRNAs) are associated with OA, but little is known of their function in chondrocyte ECM degradation. The objective of this study was to investigate the expression and function of miRNAs in OA. miRNA expression profile was determined in OA cartilage tissues and controls, employing Solexa sequencing and reverse transcription quantitative PCR (RT-qPCR). According to a modified Mankin scale, cartilage degradation was evaluated. Functional analysis of the miRNAs on chondrocyte ECM degradation was performed after miRNA transfection and IL-1β treatment. Luciferase reporter assays and western blotting were employed to determine miRNA targets. Expression of miR-221-3p was downregulated in OA cartilage tissues, which was significantly correlated with a modified Mankin scale. Through gain-of-function and loss-of-function studies, miR-221-3p was shown to significantly affect matrix synthesis gene expression and chondrocyte proliferation and apoptosis. Using SW1353 and C28I2 cells, SDF1 was identified as a target of miR-221-3p. SDF1 overexpression resulted in increased expression of catabolic genes such as MMP-13 and ADAMTS-5 in response to IL-1β, but these effects were moderated by miR-221-3p. SDF1 treatment antagonized this effect, while knockdown of SDF1 by shSDF1 induced inhibitory effects on the expression of CXCR4 and its main target genes, similar to miR-221-3p. The results indicate that upregulation of miR-221-3p could prevent IL-1β-induced ECM degradation in chondrocytes. Targeting the SDF1/CXCR4 signaling pathway may be used as a therapeutic approach for OA. miR-221-3p is downregulated in human cartilage tissues. miR-221-3p levels are associated with cartilage degeneration grade. miR-221-3p upregulation prevents IL-1β-induced ECM degradation in chondrocytes. Protection of ECM degradation by miR-223-3p occurs via SDF1/CXCR4 signaling. miR-221-3p is identified as a novel potential therapeutic target for osteoarthritis. KEY MESSAGES: miR-221-3p is downregulated in human cartilage tissues. miR-221-3p levels are associated with cartilage degeneration grade. miR-221-3p upregulation prevents IL-1β-induced ECM degradation in chondrocytes. Protection of ECM degradation by miR-223-3p occurs via SDF1/CXCR4 signaling. miR-221-3p is identified as a novel potential therapeutic target for osteoarthritis.

Section of the anterior cruciate ligament in the rabbit as animal model for osteoarthritis progression.

The purpose of this study was to determine whether instability of knee is a risk factor in the progression of osteoarthritis (OA). Twenty-four mature New Zealand White rabbits were randomly divided into four groups. The control group received 0.3ml saline in the first, fourth and seventh days in the right knee, while the other three groups received the same dosage 4% papain and its activator 0.03ML-cystein. The P3w group knees were harvested at three weeks after the last papain injection, the P6w group knees received a sham surgery at three weeks and were harvested at six weeks after the last papain injection, while the P+ACLT group knees received ACL transection at three weeks and were harvested at six weeks after the last papain injection. Cartilage degradation of femoral condyles and tibial plateaus were evaluated by X-rays, macroscopy, light microscopic and transmission electron microscopy (TEM). According to X-rays grade scale, macroscopic grade scale, light microscopic modified Mankin scale and TEM, in the P3w knees, cartilage degeneration of femoral condyles and tibial plateaus were significantly severe compared to those of the control group (P < 0.05), but the differences were not apparent in comparison with the P6w knees (P > 0.05). However, in P+ACLT knees, cartilage degeneration of femoral condyles and tibial plateaus appeared more severe in comparison with P6w knees, and the difference was significant (P < 0.05). Instability of knee plays a significant role in increasing the severity of cartilage degradation in rabbit knees and should be considered as a risk factor in OA knee progression. Our data may suggest that reconstruction of knee stability may prevent or delay the progression of OA.

Correlation of CD₄⁺ CD₂₅⁺ Foxp₃⁺ Treg with the recovery of joint function after total knee replacement in rats with osteoarthritis.

In this study, we observed changes in CD4(+) CD25(+) Foxp3(+) Treg expression in rats with osteoarthritis (OA) to explore the role that CD4(+) CD25(+) Foxp3(+) Treg plays in the decline in the condition of OA rats. Thirty rats were randomly divided into 2 groups equally and OA was induced in rats in the model group by injection of papain and l-cysteine into the right knee joint. Cartilage lesions were scored by the modified Mankin scale; pulmonary function was assessed by spirometry; interleukin (IL)-17 and IL-4 levels were evaluated by the enzyme-linked immunosorbent assay; and the levels of CD4(+) CD25(+) Foxp3(+) Treg in peripheral blood were measured by flow cytometry. The left knee joints of the model rats appeared palpable swelling and osteophytes, while the body weight, heart and lung function of these rats decreased. The serum IL-4 level was lower, whereas the serum IL-17 level was higher in the model group (P < 0.05). The peripheral blood CD4(+) CD25(+) Foxp3(+) Treg of CD4(+)T cells was significantly lower. Correlation of the changes in the levels of IL-4, IL-17, and Treg suggests that the underlying mechanism may be a reduction of the regulatory effect of Treg. The specific mechanism still requires further study.

Effect of long-term voluntary exercise on the induction of osteoarthritis by a very high-fat diet in aged mice

Purpose: Short-term voluntary exercise protects against cartilage proteoglycan loss in young mice fed a very high-fat diet. Exercise also reduces the clustering of metabolic inflammatory markers and improves glucose metabolism without weight loss, suggesting a role for systemic metabolic factors in mediating obesity-associated knee OA. We hypothesized that despite an increase in joint loading, long-term exercise would protect against knee OA in aged mice fed a very high-fat diet by reducing age and diet-associated increases in weight and glucose intolerance. Methods: Male C57BL/6J mice were fed either a control fat (CF; 10% kcal fat; n=21) or a very high fat (HF; 60% kcal fat; n=22) diet starting at 6 wks of age. At 26 wks, mice from each diet (n=9) were single-housed in cages with a running wheel until the end of the study at 52 wks of age. We compared joint loading between CF and HF fed mice using a custom force-instrumented running wheel. Body composition was quantified by DEXA and dissection of fat pads. Glucose tolerance testing was conducted at 24 and 48 wks. We evaluated OA pathology by MicroCT and histomorphometry. MicroCT was used to evaluate subchondral bone thickness and relative trabecular bone volume (BV/TV) in the medial and lateral tibial compartments. Stained sagittal knee sections were graded for cartilage OA severity using a modified Mankin scale (0-24), and anterior and posterior medial tibial osteophytes were graded using a semi-quantitative scale (0-3) by two blinded graders. Results: Prior to exercise, body weight and body fat were increased 71% and 63%, respectively, with HF feeding. Voluntary running distance (7.1 vs. 2.7 km/day), speed (59 vs. 33 cm/s) and peak limb force (115 vs. 92 %bodyweight) were lower in HF fed animals (p<0.05). Exercise did not significantly decrease body weight or epididymal fat pad mass in HF animals, although weight and fat mass were reduced with exercise in CF animals (Fig. 1A, B). Glucose tolerance area under the curve (AUC) was greater in exercised but not sedentary HF fed animals compared to activity-matched dietary controls (Fig 1C). A HF diet increased cartilage OA scores in both activity groups (Fig. 2A, p=0.007), although there was no effect of exercise. There was, however, a trend for decreased osteophyte formation in exercised animals regardless of diet (Fig. 2B, p=0.05). Conclusions: Contrary to our hypothesis, long-term voluntary wheel running exercise did not protect against HF diet-induced obesity, glucose intolerance, or OA severity in aged mice. Unlike short-term exercise in young HF-fed mice, mice that have already developed substantial diet-induced obesity show a decreased propensity for long-term voluntary wheel running exercise, which may contribute to the minimal activity-dependent changes with HF feeding in this study. Increased OA severity with long-term HF feeding was primarily due to increased cartilage degradation and proteoglycan loss and not due to increased osteophyte formation. Future work is needed to determine if systemic inflammatory markers are reduced with exercise in CF or HF fed animals to better understand the relationship between systemic metabolic inflammation and OA pathology.Figure 2Effect of a HF diet (filled bars) and long-term exercise on OA severity in 52-wk old mice. Bars sharing the same letters are not significantly different from each other (p>0.05).View Large Image Figure ViewerDownload Hi-res image Download (PPT)

The Effects of Low-Intensity Pulsed Ultrasound and Nanomagnet Applications on the Expressions of MMP-13 and MAPKs in Rabbit Knee Osteoarthritis

An animal model of Osteoarthritis (OA) was established to observe the influences of low-intensity pulsed ultrasound (LIPUS) and nano magnet application (NMA) on Collagenase 3 (MMP-13) expression and the activation status of mitogen activated protein kinases (MAPKs) in rabbit. 24 experimental rabbits from New Zealand were randomly divided into four groups: LIPUS, NMA, LIPUS + NMA group, and control group. The experimental rabbit OA model was established in the right knee joint of rabbits received ACLT operation. Rabbits in LIPUS group received LIPUS treatment and rabbits in NMA group were given NMA treatment. In LIPUS + NMA group, both treatments were applied on experimental rabbits everyday. However, the rabbits in control group only underwent ACLT operation. Four weeks later all rabbits were killed and changes of histopathology in rabbit articular cartilage were assessed and evaluated using Mankin method (Modified Mankin Scale). The protein expressions of MMP-13 and MAPKs were estimated using Western Blot. The results showed that both LIPUS and NMA treatments could significantly decrease the Mankin scores and suppress the expression level of MMP-13. However, there were some inverse results of MAPKs expression in these two applications and imply their treatment mechanisms of OA were different from each other.

Modification of osteoarthritis in the guinea pig with pulsed low-intensity ultrasound treatment

The Hartley guinea pig develops articular cartilage degeneration similar to that seen in idiopathic human osteoarthritis (OA). We investigated whether the application of pulsed low-intensity ultrasound (PLIUS) to the Hartley guinea pig joint would prevent or attenuate the progression of this degenerative process. Treatment of male Hartley guinea pigs was initiated at the onset of degeneration (8 weeks of age) to assess the ability of PLIUS to prevent OA, or at a later age (12 months) to assess the degree to which PLIUS acted to attenuate the progression of established disease. PLIUS (30 mW/cm(2)) was applied to stifle joints for 20 min/day over periods ranging from 3 to 10 months, with contralateral limbs serving as controls. Joint cartilage histology was graded according to a modified Mankin scale to evaluate treatment effect. Immunohistochemical staining for interleukin-1 receptor antagonist (IL-1ra), matrix metalloproteinase (MMP)-3, MMP-13, and transforming growth factor (TGF)-beta1 was performed on the cartilage to evaluate patterns of expression of these proteins. PLIUS did not fully prevent cartilage degeneration in the prevention groups, but diminished the severity of the disease, with the treated joints showing markedly decreased surface irregularities and a much smaller degree of loss of matrix staining as compared to controls. PLIUS also attenuated disease progression in the groups with established disease, although to a somewhat lesser extent as compared to the prevention groups. Immunohistochemical staining demonstrated a markedly decreased degree of TGF-beta1 production in the PLIUS-treated joints. This indicates less active endogenous repair, consistent with the marked reduction in cartilage degradation. PLIUS exhibits the ability to attenuate the progression of cartilage degeneration in an animal model of idiopathic human OA. The effect was greater in the treatment of early, rather than established, degeneration.

Joint degeneration following closed intraarticular fracture in the mouse knee: A model of posttraumatic arthritis

Posttraumatic arthritis is one of the most frequent causes of disability following joint trauma. The objective of this study was to develop a model of a closed articular fracture in the mouse knee joint to quantify the temporal sequence of joint degeneration in a model of posttraumatic arthritis. Closed intraarticular fractures were created in the tibial plateau of adult mice (C57BL/6) using a computer-controlled materials testing system and a custom-built indenter tip. Tibial plateau fractures were classified and imaged over time using high-resolution digital radiography. Animals were sacrificed at 2, 4, 8, and 50 weeks following fracture, and the experimental and contralateral control limbs were harvested for histology and micro-computed tomography (microCT) analysis. By radiographic analysis, tibial plateau fractures closely resembled clinical fractures. More complex and comminuted fractures correlated to significantly higher fracture energies. Histologic analysis demonstrated progressive joint degeneration as measured by a modified Mankin scale, with fibrillation and loss of proteoglycan in the articular cartilage. Subchondral bone thickening was also observed in experimental joints. The induction of a closed intraarticular fracture of the mouse tibial plateau generated a reproducible and clinically relevant joint injury that progressed to osteoarthritis-like changes by histologic and microCT evaluations. The ability to induce joint degeneration without an osteotomy or open arthrotomy provides a valuable new model for studying the natural sequelae of posttraumatic arthritis. Notably, the use of a murine model will facilitate the use of genetically modified animals for the investigation of specific genes implicated in the pathology of posttraumatic arthritis.

Histomorphometric analysis of articular cartilage, zone of calcified cartilage, and subchondral bone plate in femoral heads from clinically normal dogs and dogs with moderate or severe osteoarthritis

To quantify and compare the microscopic changes in articular cartilage (AC), zone of calcified cartilage (ZCC), and subchondral bone plate in femoral heads from clinically normal dogs and dogs with moderate or severe osteoarthritis. Femoral heads from clinically normal dogs (n = 16) and dogs with moderate (24) or severe (14) osteoarthritis. Femoral heads were allocated to 3 categories (normal, moderate, or severe osteoarthritis) on the basis of radiographic findings, macroscopic findings, and histologic grade determined by use of a modified Mankin scale. Equally spaced 2-mm sections were cut in each femoral head in a coronal or transverse plane. Thickness of the AC, ZCC, and subchondral bone plate was recorded. Mean thickness of AC was significantly greater in samples with moderate and severe osteoarthritis than those considered normal. Mean thickness of the ZCC was significantly greater in samples with moderate and severe osteoarthritis than those considered normal. Mean thickness of the subchondral bone plate in samples with severe osteoarthritis was significantly greater than those with moderate osteoarthritis and those considered normal. A significant decrease in AC thickness was detected in the proximomedial area of femoral heads with severe osteoarthritis, compared with those considered normal. A cause and effect association between thickening of subchondral structures and thinning and loss of the overlying AC was not detected. Changes in AC were associated with changes in the subchondral bone plate, which is compatible with the theory of adaptation in response to altered load distribution.

Radiofrequency (electrosurgical) ablation of articular cartilage: a study in sheep

The objective of this study was to examine the effect of a bipolar ablation probe on experimentally roughened articular cartilage and compare it with the traditional mechanical shaving technique using the knee joint of sheep. Twenty-eight skeletally mature ewes were divided randomly into two groups: one group was treated with a rotating shaving device and another group was treated using the bipolar ablation probe (Bipolar Arthroscopic Probe; Electroscope, Inc, Boulder, CO). Animals were killed at 0, 6, 12, and 24 weeks, and histological sections of the experimental limbs were compared with sections of the opposite limb using a modified Mankin scale. The following variables were used to determine scores: surface (0-6), cells (0-4), hypocellularity (0-3), matrix staining (transitional zone [0-4], radiate zone [0-4], and focal empty lacunae or hypereosinophilic cells (0-3). Differences in scores for all response variables were calculated as treated limb minus sham limb. Response variables were formed: score >0 recoded as 1 (favorable response treated better than sham), score of 0 recoded as 2 (neutral response no differences), and score <0 recoded as 3 (unfavorable response treated worse than sham). Bipolar ablative probe-treated limbs had 14.29% favorable responses and 35.71% favorable or neutral responses, whereas shave-treated limbs had 0% favorable and only 7.14% favorable or neutral responses. For all variables, bipolar ablative probe-treated limbs had more favorable responses. The less severe histological change in the bipolar ablative probe-treated joints compared with the shave-treated joints suggests that bipolar ablation of articular cartilage may be a better treatment for chondromalacia than the usual shaving methods of debridement. Further, there were no pathological changes in the subchondral bone. Arthroscopy 1998 Sep;14(6):585-91