Focal compression of the cervical spinal cord alone does not indicate high risk of neurological deterioration in patients with a diagnosis of mild degenerative cervical myelopathy.

Degenerative Cervical Myelopathy (DCM) is the functional derangement of the spinal cord resulting from vertebral column spondylotic degeneration. Typical neurological symptoms of DCM include gait imbalance, hand/arm numbness, and upper extremity dexterity loss. Greater spinal cord compression is believed to lead to a higher rate of neurological deterioration, although clinical experience suggests a more complex mechanism involving spinal canal diameter (SCD). In this study, we utilized machine learning clustering to understand the relationship between SCD and different patterns of cord compression (i.e. compression at one disc level, two disc levels, etc.) to identify patient groups at risk of neurological deterioration. 124 MRI scans from 51 non-operative DCM patients were assessed through manual scoring of cord compression and SCD measurements. Dimensionality reduction techniques and k-means clustering established patient groups that were then defined with their unique risk criteria. We found that the compression pattern is unimportant at SCD extremes (≤14.5 mm or > 15.75 mm). Otherwise, severe spinal cord compression at two disc levels increases deterioration likelihood. Notably, if SCD is normal and cord compression is not severe at multiple levels, deterioration likelihood is relatively reduced, even if the spinal cord is experiencing compression. We elucidated five patient groups with their associated risks of deterioration, according to both SCD range and cord compression pattern. Overall, SCD and focal cord compression alone do not reliably predict an increased risk of neurological deterioration. Instead, the specific combination of narrow SCD with multi-level focal cord compression increases the likelihood of neurological deterioration in mild DCM patients.

Advanced MRI metrics improve the prediction of baseline disease severity for individuals with degenerative cervical myelopathy.

BACKGROUND CONTEXT: Degenerative cervical myelopathy (DCM) is the most common form of atraumatic spinal cord injury globally. Degeneration of spinal discs, bony osteophyte growth and ligament pathology results in physical compression of the spinal cord contributing to damage of white matter tracts and grey matter cellular populations. This results in an insidious neurological and functional decline in patients which can lead to paralysis. Magnetic resonance imaging (MRI) confirms the diagnosis of DCM and is a prerequisite to surgical intervention, the only known treatment for this disorder. Unfortunately, there is a weak correlation between features of current commonly acquired MRI scans ("community MRI, cMRI") and the degree of disability experienced by a patient. PURPOSE: This study examines the predictive ability of current MRI sequences relative to "advanced MRI" (aMRI) metrics designed to detect evidence of spinal cord injury secondary to degenerative myelopathy. We hypothesize that the utilization of higher fidelity aMRI scans will increase the effectiveness of machine learning models predicting DCM severity and may ultimately lead to a more efficient protocol for identifying patients in need of surgical intervention. STUDY DESIGN/SETTING: Single institution analysis of imaging registry of patients with DCM. PATIENT SAMPLE: A total of 296 patients in the cMRI group and 228 patients in the aMRI group. OUTCOME MEASURES: Physiologic measures: accuracy of machine learning algorithms to detect severity of DCM assessed clinically based on the modified Japanese Orthopedic Association (mJOA) scale. METHODS: Patients enrolled in the Canadian Spine Outcomes Research Network registry with DCM were screened and 296 cervical spine MRIs acquired in cMRI were compared with 228 aMRI acquisitions. aMRI acquisitions consisted of diffusion tensor imaging, magnetization transfer, T2-weighted, and T2*-weighted images. The cMRI group consisted of only T2-weighted MRI scans. Various machine learning models were applied to both MRI groups to assess accuracy of prediction of baseline disease severity assessed clinically using the mJOA scale for cervical myelopathy. RESULTS: Through the utilization of Random Forest Classifiers, disease severity was predicted with 41.8% accuracy in cMRI scans and 73.3% in the aMRI scans. Across different predictive model variations tested, the aMRI scans consistently produced higher prediction accuracies compared to the cMRI counterparts. CONCLUSIONS: aMRI metrics perform better in machine learning models at predicting disease severity of patients with DCM. Continued work is needed to refine these models and address DCM severity class imbalance concerns, ultimately improving model confidence for clinical implementation.

Tryptase ß regulation of joint lubrication and inflammation via proteoglycan-4 in osteoarthritis.

PRG4 is an extracellular matrix protein that maintains homeostasis through its boundary lubricating and anti-inflammatory properties. Altered expression and function of PRG4 have been associated with joint inflammatory diseases, including osteoarthritis. Here we show that mast cell tryptase ß cleaves PRG4 in a dose- and time-dependent manner, which was confirmed by silver stain gel electrophoresis and mass spectrometry. Tryptase-treated PRG4 results in a reduction of lubrication. Compared to full-length, cleaved PRG4 further activates NF-κB expression in cells overexpressing TLR2, -4, and -5. In the destabilization of the medial meniscus model of osteoarthritis in rat, tryptase ß and PRG4 colocalize at the site of injury in knee cartilage and is associated with disease severity. When human primary synovial fibroblasts from male osteoarthritis patients or male healthy subjects treated with tryptase ß and/or PRG4 are subjected to a quantitative shotgun proteomics and proteome changes are characterized, it further supports the role of NF-κB activation. Here we show that tryptase ß as a modulator of joint lubrication in osteoarthritis via the cleavage of PRG4.

Prx1+ MPCs Accumulate in the Dura Mater of Wild-Type and p21-/- Mice Followed by a Specific Reduction in p21-/- Dural MPCs.

Epidural fat contains a population of mesenchymal progenitor cells (MPCs), and this study explores the behavior of these cells on the adjacent dura mater during growth and in response to injury in a p21 knockout mouse model. p21-/- mice are known to have increased cell proliferation and enhanced tissue regeneration post-injury. Therefore, it is hypothesized that the process by which epidural fat MPCs maintain the dura mater can be accelerated in p21-/- mice. Using a Prx1 lineage tracing mouse model, the epidural fat MPCs are found to increase in the dura mater over time in both C57BL/6 (p21+/+ ) and p21-/- mice; however, by 3 weeks post-tamoxifen induction, few MPCs are observed in p21-/- mice. These endogenous MPCs also localize to dural injuries in both mouse strains, with MPCs in p21-/- mice demonstrating increased proliferation. When epidural fat MPCs derived from p21-/- mice are transplanted into dural injuries in C57BL/6 mice, these MPCs are found in the injury site. It is demonstrated that epidural fat MPCs play a role in dural tissue maintenance and are able to directly contribute to dural injury repair. This suggests that these MPCs have the potential to treat injuries and/or pathologies in tissues surrounding the spinal cord.

IL-1Ra deficiency accelerates intervertebral disc degeneration in C57BL6J mice.

The expression of Interleukin-1ß (IL-1ß) and its antagonist and Interleukin-1 receptor antagonist (IL-1Ra) are correlated with greater human intervertebral disc (IVD) degeneration, suggesting that elevated IL-1ß activity promotes disc degeneration. Many in vitro studies support such a mechanistic relationship, whereas few in vivo investigations have been reported. The present study tests the effect of increased IL-1ß activity on intervertebral disc in mice with an IL-1Ra gene deletion. IL-1Ra-/- mice and wild-type (WT) C57Bl6J mice were examined at 3 and 12 months of age. Caudal IVD segments were evaluated for disc degeneration by histopathology, functional testing, and inflammatory gene expression relevant to IL-1ß pathways. To test differences in injury response, pinprick annular puncture was performed on IL-1Ra-/- and WT mice and evaluated similarly. IL-1Ra-/- IVDs had significantly worse histopathology at 3 months compared to WT controls, but not at 12 months. IL-1Ra-/- IVDs exhibited significantly more viscous mechanical properties than WT IVDs. qPCR revealed downregulation of inflammatory genes at 3 and 12 months in IL-1Ra-/- IVDs, with concomitant downregulation of anabolic and catabolic genes. Annular puncture yielded no appreciable differences between 2-week and 6-week post-injured WT and IL1-Ra-/- IVDs in histopathology or biomechanics, but inflammatory gene expression was sharply downregulated in IL-1Ra-/- mice at 2 weeks, returning by 6 weeks post injury. In the present study, IL-1Ra deletion resulted in increased IVD histopathology, inferior biomechanics, and transiently decreased pro-inflammatory cytokine gene expression. The histopathology of IL-1Ra-/- IVDs on a C57BL/6J background is less severe than a previous report of IL1Ra-/- on a BALB/c background, yet both strains exhibit IVD degeneration, reinforcing a mechanistic role of IL-1ß signaling in IVD pathobiology. Despite a pro-inflammatory environment, the annular puncture was no worse in IL-1Ra-/- mice, suggesting that response to injury involves pathways other than inflammation. Overall, this study supports the hypothesis that IL-1ß-driven inflammation is important in IVD degeneration.

Proteoglycan 4 is present within the dura mater and produced by mesenchymal progenitor cells.

Mesenchymal progenitor cells (MPCs) have been recently identified in human and murine epidural fat and have been hypothesized to contribute to the maintenance/repair/regeneration of the dura mater. MPCs can secrete proteoglycan 4 (PRG4/lubricin), and this protein can regulate tissue homeostasis through bio-lubrication and immunomodulatory functions. MPC lineage tracing reporter mice (Hic1) and human epidural fat MPCs were used to determine if PRG4 is expressed by these cells in vivo. PRG4 expression co-localized with Hic1+ MPCs in the dura throughout skeletal maturity and was localized adjacent to sites of dural injury. When Hic1+ MPCs were ablated, PRG4 expression was retained in the dura, yet when Prx1+ MPCs were ablated, PRG4 expression was completely lost. A number of cellular processes were impacted in human epidural fat MPCs treated with rhPRG4, and human MPCs contributed to the formation of epidural fat, and dura tissues were xenotransplanted into mouse dural injuries. We have shown that human and mouse MPCs in the epidural/dura microenvironment produce PRG4 and can contribute to dura homeostasis/repair/regeneration. Overall, these results suggest that these MPCs have biological significance within the dural microenvironment and that the role of PRG4 needs to be further elucidated.

Proteoglycan 4 (PRG4) treatment enhances wound closure and tissue regeneration.

The wound healing response is one of most primitive and conserved physiological responses in the animal kingdom, as restoring tissue integrity/homeostasis can be the difference between life and death. Wound healing in mammals is mediated by immune cells and inflammatory signaling molecules that regulate tissue resident cells, including local progenitor cells, to mediate closure of the wound through formation of a scar. Proteoglycan 4 (PRG4), a protein found throughout the animal kingdom from fish to elephants, is best known as a glycoprotein that reduces friction between articulating surfaces (e.g. cartilage). Previously, PRG4 was also shown to regulate the inflammatory and fibrotic response. Based on this, we asked whether PRG4 plays a role in the wound healing response. Using an ear wound model, topical application of exogenous recombinant human (rh)PRG4 hastened wound closure and enhanced tissue regeneration. Our results also suggest that rhPRG4 may impact the fibrotic response, angiogenesis/blood flow to the injury site, macrophage inflammatory dynamics, recruitment of immune and increased proliferation of adult mesenchymal progenitor cells (MPCs) and promoting chondrogenic differentiation of MPCs to form the auricular cartilage scaffold of the injured ear. These results suggest that PRG4 has the potential to suppress scar formation while enhancing connective tissue regeneration post-injury by modulating aspects of each wound healing stage (blood clotting, inflammation, tissue generation and tissue remodeling). Therefore, we propose that rhPRG4 may represent a potential therapy to mitigate scar and improve wound healing.

Synovial mesenchymal progenitor derived aggrecan regulates cartilage homeostasis and endogenous repair capacity.

Aggrecan is a critical component of the extracellular matrix of all cartilages. One of the early hallmarks of osteoarthritis (OA) is the loss of aggrecan from articular cartilage followed by degeneration of the tissue. Mesenchymal progenitor cell (MPC) populations in joints, including those in the synovium, have been hypothesized to play a role in the maintenance and/or repair of cartilage, however, the mechanism by which this may occur is unknown. In the current study, we have uncovered that aggrecan is secreted by synovial MPCs from healthy joints yet accumulates inside synovial MPCs within OA joints. Using human synovial biopsies and a rat model of OA, we established that this observation in aggrecan metabolism also occurs in vivo. Moreover, the loss of the "anti-proteinase" molecule alpha-2 macroglobulin (A2M) inhibits aggrecan secretion in OA synovial MPCs, whereas overexpressing A2M rescues the normal secretion of aggrecan. Using mice models of OA and cartilage repair, we have demonstrated that intra-articular injection of aggrecan into OA joints inhibits cartilage degeneration and stimulates cartilage repair respectively. Furthermore, when synovial MPCs overexpressing aggrecan were transplanted into injured joints, increased cartilage regeneration was observed vs. wild-type MPCs or MPCs with diminished aggrecan expression. Overall, these results suggest that aggrecan secreted from joint-associated MPCs may play a role in tissue homeostasis and repair of synovial joints.

Prx1 + and Hic1+ Mesenchymal Progenitors Are Present Within the Epidural Fat and Dura Mater and Participate in Dural Injury Repair.

Epidural fat is commonly discarded during spine surgery to increase the operational field. However, mesenchymal progenitor cells (MPCs) have now been identified in human epidural fat and within the murine dura mater. This led us to believe that epidural fat may regulate homeostasis and regeneration in the vertebral microenvironment. Using two MPC lineage tracing reporter mice (Prx1 and Hic1), not only have we found that epidural fat MPCs become incorporated in the dura mater over the course of normal skeletal maturation, but have also identified these cells as an endogenous source of repair and regeneration post-dural injury. Moreover, our results reveal a partial overlap between Prx1+ and Hic1+ populations, indicating a potential hierarchical relationship between the two MPC populations. This study effectively challenges the notion of epidural fat as an expendable tissue and mandates further research into its biological function and relevance.

The Dose-Response Effect of the Mast Cell Stabilizer Ketotifen Fumarate on Posttraumatic Joint Contracture: An in Vivo Study in a Rabbit Model.

Posttraumatic joint contracture is a debilitating complication following an acute fracture or intra-articular injury that can lead to loss of motion and an inability to complete activities of daily living. In prior studies using an established in vivo model, we found that ketotifen fumarate (KF), a mast cell stabilizer, was associated with a significant reduction in the severity of posttraumatic joint contracture. Our primary research question in the current study was to determine whether a dose-response relationship exists between KF and posttraumatic joint contracture reduction. METHODS: A standardized operative method to create posttraumatic joint contracture in a knee was performed on skeletally mature New Zealand White rabbits. The animals were randomly assigned to 1 of 5 groups (n = 10 per group): a nonoperative control group, an operative control group, or 1 of 3 experimental KF groups (0.01 mg/kg [the KF 0.01 group], 0.1 mg/kg [KF 0.1], or 5.0 mg/kg [KF 5.0]). Flexion contractures were measured following 8 weeks of knee immobilization using a hydraulic material-testing machine. The posterior knee joint capsules were then harvested for quantification of myofibroblast and mast cell numbers with immunohistochemistry analysis. RESULTS: Forty-five rabbits were used in the final analysis. Contracture severity was significantly reduced in the KF 0.1 group (p = 0.016) and the KF 5.0 group (p = 0.001) compared with the operative control group. When converted to a percent response, posttraumatic joint contracture reduction was 13%, 45%, and 63% for the KF 0.01, KF 0.1, and KF 5.0 groups, respectively. A half-maximal effective concentration (EC50) for KF of 0.22 mg/kg was established. There was also a decrease in myofibroblasts, mast cells, and substance P-containing nerve fiber counts with increasing doses of KF. CONCLUSIONS: Using a preclinical, rabbit in vivo model of posttraumatic joint contracture, increasing doses of KF were associated with decreasing biomechanical estimates of knee posttraumatic joint contracture as well as decreasing numbers of myofibroblasts, mast cells, and substance P-containing nerve fibers. CLINICAL RELEVANCE: KF has been used safely in humans for more than 40 years and, to our knowledge, is the first and only agent ready to be potentially translated into an effective treatment for posttraumatic joint contracture.

Epidural fat mesenchymal stem cells: Important microenvironmental regulators in health, disease, and regeneration: Do EF-MSCs play a role in dural homeostasis/maintenance?

Mesenchymal stem cells (MSCs) are present in fat tissues throughout the body, yet little is known regarding their biological role within epidural fat. We hypothesize that debridement of epidural fat and/or subsequent loss of MSCs within this tissue, disrupts homeostasis in the vertebral environment resulting in increased inflammation, fibrosis, and decreased neovascularization leading to poorer functional outcomes post-injury/operatively. Clinically, epidural fat is commonly considered a space-filling tissue with limited functionality and therefore typically discarded during surgery. However, the presence of MSCs within epidural fat suggests that itis more biologically active than historically believed and may contribute to the regulation of homeostasis and regeneration in the dural environment. While the current literature supports our hypothesis, it will require additional experimentation to determine if epidural fat is an endogenous driver of repair and regeneration and if so, this tissue should be minimally perturbed from its original location in the spinal canal. Also see the video abstract here https://youtu.be/MIol_IWK1os.

Effect of oral anticoagulants on hemostatic and thromboembolic complications in hip fracture: A systematic review and meta-analysis.

BACKGROUND: Hip fracture patients on oral anticoagulants (OACs) experience increased time-to-surgery and higher mortality compared to non-anticoagulated patients. However, it is unclear whether pre-injury OAC status and its associated operative delay are associated with worsening of peri-operative hemostasis or an increased risk of postoperative thromboembolism. METHODS: We performed a systematic review to identify studies that directly compared hemostatic and thromboembolic outcomes among hip fracture patients on an OAC prior to admission with those not on anticoagulants. Random effects meta-analyses were used to pool all outcomes of interest (estimated blood loss, transfusion requirements, and postoperative thromboembolism). RESULTS: Twenty-one studies involving 21 417 patients were included. Estimated blood loss was higher among patients presenting with OACs compared to those not anticoagulated (mean difference 31.0 mL, 95% confidence interval [CI] 6.2-55.7). Anticoagulated patients also had a 1.3-fold higher risk of receiving red blood cell transfusions (odds ratio [OR] 1.34, 95% CI 1.20-1.51); however, rates of postoperative thromboembolism were similar regardless of anticoagulation status (OR 0.96, 95% CI 0.40-2.79 for venous thromboembolism; OR 0.58, 95% CI 0.25-1.36 for arterial thromboembolism). No subgroup effect was found based on anticoagulant type or degree of surgical delay. CONCLUSION: Hip fracture patients on OACs experience increased surgical blood loss and higher risk of red blood cell transfusions. However, the degree of surgical delay did not mitigate this risk, and there was no difference in postoperative thromboembolism. The impact of appropriate, timely OAC reversal on blood conservation and expedited surgery in anticoagulated hip fracture patients warrants urgent evaluation.

Human serum mast cell tryptase levels in elbow fractures or dislocations and its association with injury severity.

Mast cells contain an abundance of tryptase, and preclinical models have shown elevated serum mast cell tryptase (SMCT) in the setting of posttraumatic joint contractures. Therefore, SMCT emerged as a potential biomarker to help recognize patients with more severe injuries and a higher likelihood of developing contractures. The objective of this study is to assess SMCT levels in participants with varying severity of elbow fractures and/or dislocations. A prospective cohort including 13 participants with more severe injuries that required an operation and 28 participants with less severe injuries managed nonoperatively were evaluated. A control group of eight individuals without elbow injuries was also evaluated. The SMCT levels were measured using an enzyme-linked immunosorbent assay kit specific for human mast cell tryptase. A one-way analysis of variance and Tukey's Honest Significance test was used to assess for statistical significance among and between the three groups. The average time from injury to the collection of the blood samples was 4 ± 2 days. Highly significant differences were identified between the operative, nonoperative, and control groups (P = .0005). In the operative group, SMCT levels were significantly higher than the nonoperative group (P = .0005) and the control group (P = .009), suggesting a correlation between SMCT levels and injury severity. There was no statistically significant difference in SMCT levels between the nonoperative and control groups. The SMCT levels were elevated in participants with acute elbow injuries requiring operative intervention, suggesting that SMCT levels were higher in injuries regarded as more severe.

CCL2 But Not CCR2 Is Required for Spontaneous Articular Cartilage Regeneration Post-Injury.

The role of the inflammatory response in articular cartilage degeneration and/or repair is often debated. Chemokine networks play a critical role in directing the recruitment of immune cells to sites of injury and have been shown to regulate cell behavior. In this study, we investigated the role of the CCL2/CCR2 signaling axis in cartilage regeneration and degeneration. CCL2-/- , CCR2-/- , CCL2-/- CCR2-/- , and control (C57) mice were subjected to full-thickness cartilage defect (FTCD) injuries (n = 9/group) within the femoral groove. Cartilage regeneration at 4 and 12 weeks post-FTCD was assessed using a 14-point histological scoring scale. Mesenchymal stem cells (MSCs) (Sca-1+ , CD140a+ ), macrophages (M1:CD38+ , M2:CD206+ , and M0:F4/80+ ) and proliferating cells (Ki67+ ) were quantified within joints using immunofluorescence. The multi-lineage differentiation capacity of Sca1+ MSCs was determined for all mouse strains. ACL transection (ACL-x) was employed to determine if CCL2-/- CCR2-/- mice were protected against osteoarthritis (OA) (n = 6/group). Absence of CCR2, but not CCL2 nor both (CCL2 and CCR2), enhanced spontaneous articular cartilage regeneration by 4 weeks post-FTCD. Furthermore, increased chondrogenesis was observed in MSCs derived from CCR2-/- mice. CCL2 deficiency promoted MSC homing to the adjacent synovium and FTCD at both 4 and 12 weeks post-injury; with no MSCs present at the surface of the FTCD in the remaining strains. Lower OA scores were observed in CCL2-/- CCR2-/- mice at 12 weeks post-ACL-x compared with C57 mice. Our findings demonstrate an inhibitory role for CCR2 in cartilage regeneration after injury, while CCL2 is required for regeneration, acting through a CCR2 independent mechanism. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:2561-2574, 2019.

Effect of spinal decompression on back pain in lumbar spinal stenosis: a Canadian Spine Outcomes Research Network (CSORN) study.

BACKGROUND CONTEXT: Surgical decompression is usually offered for improvement of neurogenic claudication in patients with symptomatic lumbar canal stenosis. These patients often have associated low back pain (LBP) and little is known about the effect of decompression on this symptom. PURPOSE: The goal of the present study is to specifically quantify the improvement in LBP following surgical decompression for lumbar canal stenosis and to identify factors associated with changes in LBP in this population. STUDY DESIGN: This is a multicenter, retrospective review of consecutive spine surgery patients enrolled by the Canadian Spine Outcomes and Research Network. PATIENT SAMPLE: Consecutive patients who underwent surgical treatment for symptomatic lumbar spine stenosis without instability between 2014 and 2017. OUTCOME MEASURES: Change in LBP on the Numeric Rating Scale (NRS). METHODS: Patient-reported outcomes were collected at baseline and at 3, 12, and 24 months after surgery. The primary outcome was change in LBP on the NRS. Multivariable logistic regression was used to model the relationship between the outcome and potential factors associated with achieving minimal clinical important difference in back pain using a backward selection procedure. RESULTS: In all, 1,221 patients were included in the analysis. Mean age was 64 years and 58% were males. Baseline back pain scores were available in 1,133 patients and follow-up evaluations were available in 968/1,133 (85%) patients at 3 months, 649/903 (72%) patients at 12 months, and 331/454 (73%) at 24 months. LBP significantly improved 3 months after surgery and the improvement was sustained at 24 months (p<.001). We found that 74% of patients reached the minimal clinical important difference in back pain. Predictive factors for sustained improvement (12 and 24 months) in LBP after surgical intervention were absence of narcotic usage or compensation claims and increased severity of LBP before surgery (high NRS). CONCLUSIONS: Alleviation of clinically significant LBP was observed at 3 months after lumbar decompression surgery for neurogenic claudication and was maintained at 12 and 24 months after surgery in the majority of patients.

17-DMAG regulates p21 expression to induce chondrogenesis in vitro and in vivo.

Cartilage degeneration after injury affects a significant percentage of the population, including those that will go on to develop osteoarthritis (OA). Like humans, most mammals, including mice, are incapable of regenerating injured cartilage. Interestingly, it has previously been shown that p21 (Cdkn1a) knockout (p21-/-) mice demonstrate auricular (ear) cartilage regeneration. However, the loss of p21 expression is highly correlated with the development of numerous types of cancer and autoimmune diseases, limiting the therapeutic translation of these findings. Therefore, in this study, we employed a screening approach to identify an inhibitor (17-DMAG) that negatively regulates the expression of p21. We also validated that this compound can induce chondrogenesis in vitro (in adult mesenchymal stem cells) and in vivo (auricular cartilage injury model). Furthermore, our results suggest that 17-DMAG can induce the proliferation of terminally differentiated chondrocytes (in vitro and in vivo), while maintaining their chondrogenic phenotype. This study provides new insights into the regulation of chondrogenesis that might ultimately lead to new therapies for cartilage injury and/or OA.

Microglial pannexin-1 channel activation is a spinal determinant of joint pain.

Chronic joint pain such as mechanical allodynia is the most debilitating symptom of arthritis, yet effective therapies are lacking. We identify the pannexin-1 (Panx1) channel as a therapeutic target for alleviating mechanical allodynia, a cardinal sign of arthritis. In rats, joint pain caused by intra-articular injection of monosodium iodoacetate (MIA) was associated with spinal adenosine 5'-triphosphate (ATP) release and a microglia-specific up-regulation of P2X7 receptors (P2X7Rs). Blockade of P2X7R or ablation of spinal microglia prevented and reversed mechanical allodynia. P2X7Rs drive Panx1 channel activation, and in rats with mechanical allodynia, Panx1 function was increased in spinal microglia. Specifically, microglial Panx1-mediated release of the proinflammatory cytokine interleukin-1ß (IL-1ß) induced mechanical allodynia in the MIA-injected hindlimb. Intrathecal administration of the Panx1-blocking peptide 10panx suppressed the aberrant discharge of spinal laminae I-II neurons evoked by innocuous mechanical hindpaw stimulation in arthritic rats. Furthermore, mice with a microglia-specific genetic deletion of Panx1 were protected from developing mechanical allodynia. Treatment with probenecid, a clinically used broad-spectrum Panx1 blocker, resulted in a striking attenuation of MIA-induced mechanical allodynia and normalized responses in the dynamic weight-bearing test, without affecting acute nociception. Probenecid reversal of mechanical allodynia was also observed in rats 13 weeks after anterior cruciate ligament transection, a model of posttraumatic osteoarthritis. Thus, Panx1-targeted therapy is a new mechanistic approach for alleviating joint pain.

Serum Mast Cell Tryptase as a Marker of Posttraumatic Joint Contracture in a Rabbit Model.

OBJECTIVES: Mast cells have been identified as key mediators of posttraumatic joint contracture, and stabilizing medications (ketotifen) have been shown to decrease contracture severity. Serum mast cell tryptase (SMCT) levels are used clinically to monitor mast cell-mediated conditions. The goals of this study were to determine if SMCT levels are elevated in the setting of joint contracture, if they can be decreased in association with ketotifen therapy, and if they correlate with contracture severity. METHODS: This study used a previously developed rabbit model in which 39 animals were divided into 4 groups: operatively created joint contracture (ORC, n = 13), operatively created contracture treated with ketotifen at 2 doses (KF0.5, n = 9; KF1.0, n = 9), and healthy rabbits (NC, n = 8). Range of motion measures were performed at 8 weeks after the surgery. Serum samples were collected on postoperative days 1, 3, 5, 7, 21, 35, and 49. SMCT levels were measured using a rabbit-specific enzyme-linked immunosorbent assay. RESULTS: Levels of SMCT were highest in the operatively created joint contracture group and were significantly greater compared with both ketotifen groups (P < 0.001). Levels were highest at postoperative day 1 with a trend to decrease over time. A positive correlation between SMCT levels and contracture severity was observed in all operative groups (P < 0.05). CONCLUSIONS: Levels of SMCT are elevated in the setting of joint contracture, decreased in association with ketotifen therapy, and positively correlated with contracture severity. This is the first study to establish a relationship between SMCT and joint injury. Measurement of SMCT may be valuable in identifying those at risk of posttraumatic joint contracture.

Compromised Neurotrophic and Angiogenic Regenerative Capability during Tendon Healing in a Rat Model of Type-II Diabetes.

Metabolic diseases such as diabetes mellitus type-II (DM-II) may increase the risk of suffering painful connective tissue disorders and tendon ruptures. The pathomechanisms, however, by which diabetes adversely affects connective tissue matrix metabolism and regeneration, still need better definition. Our aim was to study the effect of DM-II on expressional changes of neuro- and angiotrophic mediators and receptors in intact and healing Achilles tendon. The right Achilles tendon was transected in 5 male DM-II Goto-Kakizaki (GK) and 4 age-matched Wistar control rats. The left Achilles tendons were left intact. At week 2 post-injury, NGF, BDNF, TSP, and receptors TrkA, TrkB and Nk1 gene expression was studied by quantitative RT-PCR (qRT-PCR) and their protein distribution by immunohistochemistry in intact and injured tendons. The expression of tendon-related markers, Scleraxis (SCX) and Tenomodulin (TNMD), was evaluated by qRT-PCR in intact and injured tendons. Injured tendons of diabetic GK rats exhibited significantly down-regulated Ngf and Tsp1 mRNA and corresponding protein levels, and down-regulated Trka gene expression compared to injured Wistar controls. Intact tendons of DM-II GK rats displayed reduced mRNA levels for Ngf, Tsp1 and Trkb compared to corresponding intact non-diabetic tendons. Up-regulated Scx and Tnmd gene expression was observed in injured tendons of normal and diabetic GK rats compared to intact Wistar controls. However, these molecules were not up-regulated in injured DM-II GK rats compared to their corresponding controls. Our results suggest that DM-II has detrimental effects on neuro- and angiotrophic pathways, and such effects may reflect the compromised repair seen in diabetic Achilles tendon. Thus, novel approaches for regeneration of injured, including tendinopathic, and surgically repaired diabetic tendons may include therapeutic molecular modulation of neurotrophic pathways such as NGF and its receptors.

Tendon Innervation.

The regulation of tendon metabolism including the responses to loading is far from being well understood. During the last decade, however, accumulating data show that tendon innervation in addition to afferent functions, via efferent pathways has a regulatory role in tendon homeostasis via a wide range of neuromediators, which coordinate metabolic and neuro-inflammatory pathways.Innervation of intact healthy tendons is localized in the surrounding structures, i.e paratenon, endotenon and epitenon, whereas the tendon proper is practically devoid of neuronal supply. This anatomical finding reflects that the tendon metabolism is regulated from the tendon envelope, i.e. interfascicular matrix (see Chap. 1 ).Tendon innervation after injury and during repair, however, is found as extensive nerve ingrowth into the tendon proper, followed by a time-dependent emergence of different neuronal mediators, which amplify and fine-tune inflammatory and metabolic pathways in tendon regeneration. After healing nerve fibers retract to the tendon envelope.In tendinopathy innervation has been identified to consist of excessive and protracted nerve ingrowth in the tendon proper, suggesting pro-inflammatory, nociceptive and hypertrophic (degenerative) tissue responses.In metabolic disorders such as eg. diabetes impaired tendon healing has been established to be related to dysregulation of neuronal growth factors.Targeted approaches to the peripheral nervous system including neuronal mediators and their receptors may prove to be effective therapies for painful, degenerative and traumatic tendon disorders.