Perioperative care for lumbar microdiscectomy: a survey of Australasian neurosurgeons.

BACKGROUND: Lumbar microdiscectomy is the most commonly performed spine surgery procedure. Over time it has evolved to a minimally invasive procedure. Traditionally patients were advised to restrict activity following lumbar spine surgery. However, post-operative instructions are heterogeneous. The purpose of this report is to assess, by survey, the perioperative care practices of Australasian neurosurgeons in the minimally invasive era. METHODS: A survey was conducted by email invitation sent to all full members of the Neurosurgical Society of Australasia (NSA). This consisted of 11 multi-choice questions relating to operative indications, technique, and post-operative instructions for lumbar microdiscectomy answered by an electronically distributed anonymized online survey. RESULTS: The survey was sent to all Australasian Neurosurgeons. In total, 68 complete responses were received (28.9%). Most surgeons reported they would consider a period of either 4 to 8 weeks (42.7%) or 8 to 12 weeks (32.4%) as the minimum duration of radicular pain adequate to offer surgery. Unilateral muscle dissection with unilateral discectomy was practiced by 76.5%. Operative microscopy was the most commonly employed method of magnification (76.5%). The majority (55.9%) always refer patients to undergo inpatient physiotherapy. Sitting restrictions were advised by 38.3%. Lifting restrictions were advised by 83.8%. CONCLUSIONS: Australasian neurosurgical lumbar microdiscectomy perioperative care practices are generally consistent with international practices and demonstrate a similar degree of heterogeneity. Recommendation of post-operative activity restrictions by Australasian neurosurgeons is still common. This suggests a role for the investigation of the necessity of such restrictions in the era of minimally invasive spine surgery.

Mesenchymal progenitor cells primed with pentosan polysulfate promote lumbar intervertebral disc regeneration in an ovine model of microdiscectomy.

BACKGROUND CONTEXT: Neural compression associated with lumbar disc herniation is usually managed surgically by microdiscectomy. However, 10%-20% of patients re-present with debilitating back pain, and approximately 15% require further surgery. PURPOSE: Using an ovine model of microdiscectomy, the present study investigated the relative potential of pentosan polysulfate-primed mesenchymal progenitor cells (pMPCs) or MPC alone implanted into the lesion site to facilitate disc recovery. STUDY DESIGN: An ovine model of lumbar microdiscectomy was used to compare the relative outcomes of administering MPCs or pMPCs to the injury site postsurgery. METHODS: At baseline 3T magnetic resonance imaging (MRI) of 18 adult ewes was undertaken followed by annular microdiscectomy at two lumbar disc levels. Sheep were randomized into three groups (n=6). The injured controls received no further treatment. Defects of the treated groups were implanted with a collagen sponge and MPC (5×105 cells) or pMPC (5×105 cells). After 6 months, 3T MRI and standard radiography were performed. Spinal columns were dissected, individual lumbar discs were sectioned horizontally, and nucleus pulposus (NP) and annulus fibrosus (AF) regions were assessed morphologically and histologically. The NP and AF tissues were dissected into six regions and analyzed biochemically for their proteoglycans (PGs), collagen, and DNA content. RESULTS: Both the MPC- and pMPC-injected groups exhibited less reduction in disc height (p<.05) and lower Pfirrmann grades (p≤.001) compared with the untreated injury controls, but morphologic scores for the pMPC-injected discs were lower (p<.05). The PG content of the AF injury site region (AF1) of pMPC discs was higher than MPC and injury control AF1 (p<.05). At the AF1 and contralateral AF2 regions, the DNA content of pMPC discs was significantly lower than injured control discs and MPC-injected discs. Histologic and birefringent microscopy revealed increased structural organization and reduced degeneration in pMPC discs compared with MPC and the injured controls. CONCLUSIONS: In an ovine model 6 months after administration of pMPCs to the injury site disc PG content and matrix organization were improved relative to controls, suggesting pMPCs' potential as a postsurgical adjunct for limiting progression of disc degeneration after microdiscectomy.

Pentosan polysulfate binds to STRO-1+ mesenchymal progenitor cells, is internalized, and modifies gene expression: a novel approach of pre-programing stem cells for therapeutic application requiring their chondrogenesis.

BACKGROUND: The pharmaceutical agent pentosan polysulfate (PPS) is known to induce proliferation and chondrogenesis of mesenchymal progenitor cells (MPCs) in vitro and in vivo. However, the mechanism(s) of action of PPS in mediating these effects remains unresolved. In the present report we address this issue by investigating the binding and uptake of PPS by MPCs and monitoring gene expression and proteoglycan biosynthesis before and after the cells had been exposed to limited concentrations of PPS and then re-established in culture in the absence of the drug (MPC priming). METHODS: Immuno-selected STRO-1+ mesenchymal progenitor stem cells (MPCs) were prepared from human bone marrow aspirates and established in culture. The kinetics of uptake, shedding, and internalization of PPS by MPCs was determined by monitoring the concentration-dependent loss of PPS media concentrations using an enzyme-linked immunosorbent assay (ELISA) and the uptake of fluorescein isothiocyanate (FITC)-labelled PPS by MPCs. The proliferation of MPCs, following pre-incubation and removal of PPS (priming), was assessed using the Wst-8 assay method, and proteoglycan synthesis was determined by the incorporation of 35SO4 into their sulphated glycosaminoglycans. The changes in expression of MPC-related cell surface antigens of non-primed and PPS-primed MPCs from three donors was determined using flow cytometry. RNA sequencing of RNA isolated from non-primed and PPS-primed MPCs from the same donors was undertaken to identify the genes altered by the PPS priming protocol. RESULTS: The kinetic studies indicated that, in culture, PPS rapidly binds to MPC surface receptors, followed by internalisation and localization within the nucleus of the cells. Following PPS-priming of MPCs and a further 48 h of culture, both cell proliferation and proteoglycan synthesis were enhanced. Reduced expression of MPC-related cell surface antigen expression was promoted by the PPS priming, and RNA sequencing analysis revealed changes in the expression of 42 genes. CONCLUSION: This study has shown that priming of MPCs with low concentrations of PPS enhanced chondrogenesis and MPC proliferation by modifying their characteristic basal gene and protein expression. These findings offer a novel approach to re-programming mesenchymal stem cells for clinical indications which require the repair or regeneration of cartilaginous tissues such as in osteoarthritis and degenerative disc disease.

Safety, tolerability, clinical, and joint structural outcomes of a single intra-articular injection of allogeneic mesenchymal precursor cells in patients following anterior cruciate ligament reconstruction: a controlled double-blind randomised trial.

BACKGROUND: Few clinical trials have investigated the safety and efficacy of mesenchymal stem cells for the management of post-traumatic osteoarthritis. The objectives of this pilot study were to determine the safety and tolerability and to explore the efficacy of a single intra-articular injection of allogeneic human mesenchymal precursor cells (MPCs) to improve clinical symptoms and retard joint structural deterioration over 24 months in patients following anterior cruciate ligament (ACL) reconstruction. METHODS: In this phase Ib/IIa, double-blind, active comparator clinical study, 17 patients aged 18-40 years with unilateral ACL reconstruction were randomized (2:1) to receive either a single intra-articular injection of 75 million allogeneic MPCs suspended in hyaluronan (HA) (MPC + HA group) (n = 11) or HA alone (n = 6). Patients were monitored for adverse events. Immunogenicity was evaluated by anti-HLA panel reactive antibodies (PRA) against class I and II HLAs determined by flow cytometry. Pain, function, and quality of life were assessed using the Knee Injury and Osteoarthritis Outcome Score (KOOS) and SF-36v2 scores. Joint space width was measured from radiographs, and tibial cartilage volume and bone area assessed from magnetic resonance imaging (MRI). RESULTS: Moderate arthralgia and swelling within 24 h following injection that subsided were observed in 4 out of 11 in the MPC + HA group and 0 out of 6 HA controls. No cell-related serious adverse effects were observed. Increases in class I PRA >10% were observed at week 4 in the MPC + HA group that decreased to baseline levels by week 104. Compared with the HA group, MPC + HA-treated patients showed greater improvements in KOOS pain, symptom, activities of daily living, and SF-36 bodily pain scores (p < 0.05). The MPC + HA group had reduced medial and lateral tibiofemoral joint space narrowing (p < 0.05), less tibial bone expansion (0.5% vs 4.0% over 26 weeks, p = 0.02), and a trend towards reduced tibial cartilage volume loss (0.7% vs -4.0% over 26 weeks, p = 0.10) than the HA controls. CONCLUSIONS: Intra-articular administration of a single allogeneic MPC injection following ACL reconstruction was safe, well tolerated, and may improve symptoms and structural outcomes. These findings suggest that MPCs warrant further investigations as they may modulate some of the pathological processes responsible for the development of post-traumatic osteoarthritis following ACL reconstruction. TRIAL REGISTRATION: ClinicalTrials.gov ( NCT01088191 ) registration date: March 11, 2010.

Immunoselected STRO-3+ mesenchymal precursor cells reduce inflammation and improve clinical outcomes in a large animal model of monoarthritis.

BACKGROUND: The purpose of this study was to investigate the therapeutic efficacy of intravenously administered immunoselected STRO-3 + mesenchymal precursor cells (MPCs) on clinical scores, joint pathology and cytokine production in an ovine model of monoarthritis. METHODS: Monoarthritis was established in 16 adult merino sheep by administration of bovine type II collagen into the left hock joint following initial sensitization to this antigen. After 24 h, sheep were administered either 150 million allogeneic ovine MPCs (n = 8) or saline (n = 8) intravenously (IV). Lameness, joint swelling and pain were monitored and blood samples for leukocytes and cytokine levels were collected at intervals following arthritis induction. Animals were necropsied 14 days after arthritis induction and gross and histopathological evaluations were undertaken on tissues from the arthritic (left) and contralateral (right) joints. RESULTS: MPC-treated sheep demonstrated significantly reduced clinical signs of lameness, joint pain and swelling compared with saline controls. They also showed decreased cartilage erosions, synovial stromal cell activation and angiogenesis. This was accompanied by decreased infiltration of the synovial tissues by CD4+ lymphocytes and CD14+ monocytes/macrophages. Over the 3 days following joint arthropathy induction, the numbers of neutrophils circulating in the blood and plasma concentrations of activin A were significantly reduced in animals administered MPCs. CONCLUSIONS: The results of this study have demonstrated the capacity of IV-administered MPCs to mitigate the clinical signs and some of the inflammatory mediators responsible for joint tissue destruction in a large animal model of monoarthritis.

Reconstitution of degenerated ovine lumbar discs by STRO-3-positive allogeneic mesenchymal precursor cells combined with pentosan polysulfate.

OBJECTIVE Disc degeneration and associated low-back pain are major causes of suffering and disability. The authors examined the potential of mesenchymal precursor cells (MPCs), when formulated with pentosan polysulfate (PPS), to ameliorate disc degeneration in an ovine model. METHODS Twenty-four sheep had annular incisions made at L2-3, L3-4, and L4-5 to induce degeneration. Twelve weeks after injury, the nucleus pulposus of a degenerated disc in each animal was injected with ProFreeze and PPS formulated with either a low dose (0.1 million MPCs) or a high dose (0.5 million MPCs) of cells. The 2 adjacent injured discs in each spine were either injected with PPS and ProFreeze (PPS control) or not injected (nil-injected control). The adjacent noninjured L1-2 and L5-6 discs served as noninjured control discs. Disc height indices (DHIs) were obtained at baseline, before injection, and at planned death. After necropsy, 24 weeks after injection, the spines were subjected to MRI and morphological, histological, and biochemical analyses. RESULTS Twelve weeks after the annular injury, all the injured discs exhibited a significant reduction in mean DHI (low-dose group 17.19%; high-dose group 18.01% [p < 0.01]). Twenty-four weeks after injections, the discs injected with the low-dose MPC+PPS formulation recovered disc height, and their mean DHI was significantly greater than the DHI of PPS- and nil-injected discs (p < 0.001). Although the mean Pfirrmann MRI disc degeneration score for the low-dose MPC+PPS-injected discs was lower than that for the nil- and PPS-injected discs, the differences were not significant. The disc morphology scores for the nil- and PPS-injected discs were significantly higher than the normal control disc scores (p < 0.005), whereas the low-dose MPC+PPS-injected disc scores were not significantly different from those of the normal controls. The mean glycosaminoglycan content of the nuclei pulposus of the low-dose MPC+PPS-injected discs was significantly higher than that of the PPS-injected controls (p < 0.05) but was not significantly different from the normal control disc glycosaminoglycan levels. Histopathology degeneration frequency scores for the low-dose MPC+PPS-injected discs were lower than those for the PPS- and Nil-injected discs. The corresponding high-dose MPC+PPS-injected discs failed to show significant improvements in any outcome measure relative to the controls. CONCLUSIONS Intradiscal injections of a formulation composed of 0.1 million MPCs combined with PPS resulted in positive effects in reducing the progression of disc degeneration in an ovine model, as assessed by improvements in DHI and morphological, biochemical, and histopathological scores.

Cell-Based Therapies Used to Treat Lumbar Degenerative Disc Disease: A Systematic Review of Animal Studies and Human Clinical Trials.

Low back pain and degenerative disc disease are a significant cause of pain and disability worldwide. Advances in regenerative medicine and cell-based therapies, particularly the transplantation of mesenchymal stem cells and intervertebral disc chondrocytes, have led to the publication of numerous studies and clinical trials utilising these biological therapies to treat degenerative spinal conditions, often reporting favourable outcomes. Stem cell mediated disc regeneration may bridge the gap between the two current alternatives for patients with low back pain, often inadequate pain management at one end and invasive surgery at the other. Through cartilage formation and disc regeneration or via modification of pain pathways stem cells are well suited to enhance spinal surgery practice. This paper will systematically review the current status of basic science studies, preclinical and clinical trials utilising cell-based therapies to repair the degenerate intervertebral disc. The mechanism of action of transplanted cells, as well as the limitations of published studies, will be discussed.

Effect of mesenchymal precursor cells on the systemic inflammatory response and endothelial dysfunction in an ovine model of collagen-induced arthritis.

BACKGROUND AND AIM: Mesenchymal precursor cells (MPC) are reported to possess immunomodulatory properties that may prove beneficial in autoimmune and other inflammatory conditions. However, their mechanism of action is poorly understood. A collagen-induced arthritis model has been previously developed which demonstrates local joint inflammation and systemic inflammatory changes. These include not only increased levels of inflammatory markers, but also vascular endothelial cell dysfunction, characterised by reduced endothelium-dependent vasodilation. This study aimed to characterise the changes in systemic inflammatory markers and endothelial function following the intravenous administration of MPC, in the ovine model. METHODS: Arthritis was induced in sixteen adult sheep by administration of bovine type II collagen into the hock joint following initial sensitisation. After 24h, sheep were administered either 150 million allogeneic ovine MPCs intravenously, or saline only. Fibrinogen and serum amyloid-A were measured in plasma to assess systemic inflammation, along with pro-inflammatory and anti-inflammatory cytokines. Animals were necropsied two weeks following arthritis induction. Coronary and digital arterial segments were mounted in a Mulvaney-Halpern wire myograph. The relaxant response to endothelium-dependent and endothelium-independent vasodilators was used to assess endothelial dysfunction. RESULTS AND CONCLUSION: Arthritic sheep treated with MPC demonstrated a marked spike in plasma IL-10, 24h following MPC administration. They also showed significantly reduced plasma levels of the inflammatory markers, fibrinogen and serum amyloid A, and increased HDL. Coronary arteries from RA sheep treated with MPCs demonstrated a significantly greater maximal relaxation to bradykinin when compared to untreated RA sheep (253.6 ± 17.1% of pre-contracted tone vs. 182.3 ± 27.3% in controls), and digital arteries also demonstrated greater endothelium-dependent vasodilation. This study demonstrated that MPCs given intravenously are able to attenuate systemic inflammatory changes associated with a monoarthritis, including the development of endothelial dysfunction.

The role of stem cell therapies in degenerative lumbar spine disease: a review.

Degenerative conditions of the lumbar spine are extremely common. Ninety percent of people over the age of 60 years have degenerative change on imaging; however, only a small minority of people will require spine surgery (Hicks et al. Spine (Phila Pa 1976) 34(12):1301-1306, 2009). This minority, however, constitutes a core element of spinal surgery practice. Whilst the patient outcomes from spinal surgeries have improved in recent years, some patients will remain with pain and disability despite technically successful surgery. Advances in regenerative medicine and stem cell therapies, particularly the use of mesenchymal stem cells and allogeneic mesenchymal precursor cells, have led to numerous clinical trials utilising these cell-based therapies to treat degenerative spinal conditions. Through cartilage formation and disc regeneration, fusion enhancement or via modification of pain pathways, stem cells are well suited to enhance spinal surgery practice. This review will focus on the outcomes of lumbar spinal procedures and the role of stem cells in the treatment of degenerative lumbar conditions to enhance clinical practice. The current status of clinical trials utilising stem cell therapies will be discussed, providing clinicians with an overview of the various cell-based treatments likely to be available to patients in the near future.

Mesenchymal progenitor cells combined with pentosan polysulfate mediating disc regeneration at the time of microdiscectomy: a preliminary study in an ovine model.

OBJECT: Following microdiscectomy, discs generally fail to undergo spontaneous regeneration and patients may experience chronic low-back pain and recurrent disc prolapse. In published studies, formulations of mesenchymal progenitor cells combined with pentosan polysulfate (MPCs+PPS) have been shown to regenerate disc tissue in animal models, suggesting that this approach may provide a useful adjunct to microdiscectomy. The goal of this preclinical laboratory study was to determine if the transplantation of MPCs+PPS, embedded in a gelatin/fibrin scaffold (SCAF), and transplanted into a defect created by microdiscectomy, could promote disc regeneration. METHODS: A standardized microdiscectomy procedure was performed in 18 ovine lumbar discs. The subsequent disc defects were randomized to receive either no treatment (NIL), SCAF only, or the MPC+PPS formulation added to SCAF (MPCs+PPS+SCAF). Necropsies were undertaken 6 months postoperatively and the spines analyzed radiologically (radiography and MRI), biochemically, and histologically. RESULTS: No adverse events occurred throughout the duration of the study. The MPC+PPS+SCAF group had significantly less reduction in disc height compared with SCAF-only and NIL groups (p < 0.05 and p < 0.01, respectively). Magnetic resonance imaging Pfirrmann scores in the MPC+PPS+SCAF group were significantly lower than those in the SCAF group (p = 0.0213). The chaotropic solvent extractability of proteoglycans from the nucleus pulposus of MPC+PPS+SCAF-treated discs was significantly higher than that from the SCAF-only discs (p = 0.0312), and using gel exclusion chromatography, extracts from MPC+PPS+SCAF-treated discs also contained a higher percentage of proteoglycan aggregates than the extracts from both other groups. Analysis of the histological sections showed that 66% (p > 0.05) of the MPC+PPS+SCAF-treated discs exhibited less degeneration than the NIL or SCAF discs. CONCLUSIONS: These findings demonstrate the capacity of MPCs in combination with PPS, when embedded in a gelatin sponge and sealed with fibrin glue in a microdiscectomy defect, to restore disc height, disc morphology, and nucleus pulposus proteoglycan content.

An injectable hydrogel incorporating mesenchymal precursor cells and pentosan polysulphate for intervertebral disc regeneration.

Intervertebral disc (IVD) degeneration is one of the leading causes of lower back pain and a major health problem worldwide. Current surgical treatments include excision or immobilisation, with neither approach resulting in the repair of the degenerative disc. As such, a tissue engineering-based approach in which stem cells, coupled with an advanced delivery system, could overcome this deficiency and lead to a therapy that encourages functional fibrocartilage generation in the IVD. In this study, we have developed an injectable hydrogel system based on enzymatically-crosslinked polyethylene glycol and hyaluronic acid. We examined the effects of adding pentosan polysulphate (PPS), a synthetic glycosaminoglycan-like factor that has previously been shown (in vitro and in vivo) to this gel system in order to induce chondrogenesis in mesenchymal precursor cells (MPCs) when added as a soluble factor, even in the absence of additional growth factors such as TGF-ß. We show that both the gelation rate and mechanical strength of the resulting hydrogels can be tuned in order to optimise the conditions required to produce gels with the desired combination of properties for an IVD scaffold. Human immunoselected STRO-1+ MPCs were then incorporated into the hydrogels. They were shown to retain good viability after both the initial formation of the gel and for longer-term culture periods in vitro. Furthermore, MPC/hydrogel composites formed cartilage-like tissue which was significantly enhanced by the incorporation of PPS into the hydrogels, particularly with respect to the deposition of type-II-collagen. Finally, using a wild-type rat subcutaneous implantation model, we examined the extent of any immune reaction and confirmed that this matrix is well tolerated by the host. Together these data provide evidence that such a system has significant potential as both a delivery vehicle for MPCs and as a matrix for fibrocartilage tissue engineering applications.

Current and future applications for stem cell therapies in spine surgery.

Spinal surgery involves the bone-cartilage-neural interface. It is a field of surgery that is rapidly changing and evolving; not only through the development of novel techniques, approaches and devices but also through evidence from large clinical trials assessing indications, efficacy and outcomes. The use of biologics in spine surgery has now become widespread. Biologics in the form of autologous or allogeneic stem cells or progenitor cells are not yet in routine clinical use in spine surgery. However it is likely that they will have a significant role in the future, since increasing numbers of preclinical and clinical studies have demonstrated the safety and efficacy of progenitor cells to treat a variety of spinal conditions. Such studies have paved the way to larger clinical trials. Cell therapies encompass a wide range of stem cell and progenitor cell types. Stem cells subtypes differ in their lineage potential often being described as pluripotent or multipotent, some of which have potential application in therapies to treat diseases of the spine having the ability to differentiate into tissues including bone and cartilage and to secrete factors that promote matrix repair and regeneration. Furthermore, studies have shown that some cells, particularly mesenchymal stromal cells, modulate oxidative stress and secrete cytokines and growth factors that have immunomodulatory, antiinflammatory, angiogenic and antiapoptotic effects. It is these combined characteristics that make cell based therapies prime candidates for advancing current techniques in spine surgery and for providing new strategies directed at targeting the underlying causes of spinal diseases and disorders to promote repair and regeneration. This review will explore the characteristics of various stem cells and other progenitor cells derived from different sources. The authors are not suggesting that all these cells are necessarily suitable clinically. The review will thus focus on their application to both current and potentially future areas of spine surgery based on results of the available evidence and clinical trials. This review will not address spinal cord injury.

Lateral surgical approach to lumbar intervertebral discs in an ovine model.

The sheep is becoming increasingly used as a large animal model for preclinical spine surgery studies. Access to the ovine lumbar intervertebral discs has traditionally been via an anterior or anterolateral approach, which requires larger wound incisions and, at times, significant abdominal retraction. We present a new minimally invasive operative technique for a far-lateral approach to the ovine lumbar spine that allows for smaller incisions, excellent visualisation of intervertebral discs, and minimal abdominal retraction and is well tolerated by animals with minimal morbidity.

Immunoselected STRO-3+ mesenchymal precursor cells and restoration of the extracellular matrix of degenerate intervertebral discs.

OBJECT: Chronic low-back pain of discal origin is linked strongly to disc degeneration. Current nonsurgical treatments are palliative and fail to restore the disc extracellular matrix. In this study the authors examined the capacity of ovine mesenchymal precursor cells (MPCs) to restore the extracellular matrix of degenerate discs in an ovine model. METHODS: Three adjacent lumbar discs of 24 adult male sheep were injected intradiscally with chondroitinase-ABC (cABC) to initiate disc degeneration. The remaining lumbar discs were used as normal controls. Three months after cABC injection, the L3-4 discs of all animals were injected with either a high dose (4 × 10(6) cells, in 12 sheep) or low dose (0.5 × 10(6) cells, in 12 sheep) of MPCs suspended in hyaluronic acid (HA). The adjacent L4-5 degenerate discs remained untreated; the L5-6 discs were injected with HA only. The animals were euthanized at 3 or 6 months after MPC injections (6 sheep from each group at each time point), and histological sections of the lumbar discs were prepared. Radiographs and MR images were obtained prior to cABC injection (baseline), 3 months after cABC injection (pretreatment), and just prior to necropsy (posttreatment). RESULTS: Injection of cABC decreased the disc height index (DHI) of target discs by 45%-50%, confirming degeneration. Some recovery in DHI was observed 6 months after treatment in all cABC-injected discs, but the DHI increased to within baseline control values only in the MPC-injected discs. This improvement was accompanied by a reduction in MRI degeneration scores. The histopathology scores observed at 3 months posttreatment for the high-dose MPC-injected discs and at 6 months posttreatment for the low-dose MPC-injected discs were significantly different from those of the noninjected and HA-injected discs (p <0.001) but not from the control disc scores. CONCLUSIONS: On the basis of the findings of this study, the authors conclude that the injection of MPCs into degenerate intervertebral discs can contribute to the regeneration of a new extracellular matrix.

Pentosan polysulfate increases affinity between ADAMTS-5 and TIMP-3 through formation of an electrostatically driven trimolecular complex.

The semi-synthetic sulfated polysaccharide PPS (pentosan polysulfate) increases affinity between the aggrecan-degrading ADAMTSs (adamalysins with thrombospondin motifs) and their endogenous inhibitor, TIMP (tissue inhibitor of metalloproteinases)-3. In the present study we demonstrate that PPS mediates the formation of a high-affinity trimolecular complex with ADAMTS-5 and TIMP-3. A TIMP-3 mutant that lacks extracellular-matrix-binding ability was insensitive to this affinity increase, and truncated forms of ADAMTS-5 that lack the Sp (spacer) domain had reduced PPS-binding ability and sensitivity to the affinity increase. PPS molecules composed of 11 or more saccharide units were 100-fold more effective than those of eight saccharide units, indicating the involvement of extended or multiple protein-interaction sites. The formation of a high-affinity trimolecular complex was completely abolished in the presence of 0.4 M NaCl. These results suggest that PPS enhances the affinity between ADAMTS-5 and TIMP-3 by forming electrostatically driven trimolecular complexes under physiological conditions.

A comparison of mesenchymal precursor cells and amnion epithelial cells for enhancing cervical interbody fusion in an ovine model.

BACKGROUND: Rapid, reliable fusion is the goal in anterior cervical diskectomy and fusion. Iliac crest autograft has a high rate of donor-site morbidity. Alternatives such as bone graft substitutes lack osteoinductivity, and recombinant bone morphogenetic proteins risk life-threatening complications. Both allogeneic mesenchymal precursor cells (MPCs) and amnion derived epithelial cells (AECs) have osteogenic potential. OBJECTIVE: To compare for the first time the capacity of MPCs and AECs to promote osteogenesis in an ovine model. METHODS: Five groups of 2-year-old ewes were subjected to C3-4 anterior cervical diskectomy and fusion with a Fidji interbody cage packed with iliac crest autograft alone (group A; n = 6), hydroxyapatite-tricalcium phosphate Mastergraft granules (HA/TCP) alone (group B; n = 6), HA/TCP containing 5 million MPCs (group C; n = 6), or HA/TCP containing 5 million AECs (group D; n = 5); group E was made up of age-matched nonoperative controls (n = 6). At 3 months, animals were euthanized and quantitative multislice computed tomography, functional radiography, biomechanics, histology, and histomorphometry were performed. RESULTS: No procedure- or cell-related adverse events were observed. There was significantly more fusion in the MPC group (C) than in group A, B, or D. Computed tomography scan at 3 months revealed that 5 of 6 MPC-treated animals (83%) had continuous bony bridging compared with 0 of 5 AEC-treated and only 1 of 6 autograft- and 2 of 6 HA/TCP-treated animals (P = .01). CONCLUSION: Implantation of allogeneic MPCs in combination with HA/TCP within an interbody spacer facilitates interbody fusion after diskectomy. The earlier, more robust fusion observed with MPCs relative to autograft and HA/TCP bone substitute indicates that this approach may offer a therapeutic benefit.

Cervical interbody fusion is enhanced by allogeneic mesenchymal precursor cells in an ovine model.

STUDY DESIGN: An experimental study using a sheep cervical spine interbody fusion model. OBJECTIVE: To compare allogeneic mesenchymal precursor cells combined with hydroxyapatite and tricalcium phosphate (HA/TCP) with HA/TCP alone or iliac crest autograft (AG) for cervical interbody fusion. SUMMARY OF BACKGROUND DATA: We investigated the effect of mesenchymal precursor cells on cervical fusion because of the shortcomings of using iliac crest (donor site morbidity), bone substitute (poor osteoinductive properties), and bone morphogenic proteins (serious complications). METHODS: Thirty ewes were divided randomly into four groups of six having C3-C4 anterior cervical discectomy and fusion using a Fidji cage packed with, AG, HA/TCP, HA/TCP containing 5 million MPCs, and HA/TCP containing 10 million MPCs. MPCs were derived from a single batch of immuno-selected and culture-expanded MPCs isolated from bone marrow of out-bred sheep. The fifth group were nonoperated controls. Safety, fusion parameters, and biomechanics were assessed. RESULTS: No cell-related adverse events were observed. No significant differences were found between the five or 10 million MPC groups. Evaluation of fusion by CT scan at 3 months showed that 9 of 12 (75%) MPC-treated animals had continuous bony bridging compared with only 1 of 6 AG and 2 of 6 HA/TCP (P = 0.019 and P = 0.044, respectively). By quantitative CT, density of new bone in MPC-treated animals was 121% higher than in HA/TCP (P = 0.017) and 128% higher than in AG (P < 0.0001). Functional radiology at 3 months revealed that MPC-treated animals had significantly reduced macromotion at C3/4 compared with AG and HA/TCP groups combined (P = 0.007). CONCLUSION: Implantation of allogeneic MPCs when combined with HA/TCP and an interbody spacer facilitates new bone formation after discectomy without any cell-related complications. The earlier and dense new bone formation observed with MPCs relative to autograft and HA/TCP alone suggest that this approach may offer therapeutic benefit.

Potential applications for using stem cells in spine surgery.

While the use of biologics as adjuncts for spine surgery is growing annually stem cells have yet to be approved for this clinical application. Stem cells have the unique ability to differentiate into a variety of musculoskeletal tissues including bone or cartilage. Moreover they have been shown to secrete growth factors that promote matrix repair and regeneration and can down regulate inflammation and immune cell functions. It is these combined activities that make stem cells attractive candidates for advancing current techniques in spine surgery and possibly mitigating those pathologies responsible for tissue degeneration and failure thereby minimising the need for surgical intervention at a later date. This review focuses on the characteristics of progenitor cells from different sources and explores their potential as adjuncts for both current and future applications in spine surgery. Where possible we draw on the experimental outcomes from our own preclinical studies using adult mesenchymal progenitor stem cells, as well as related studies by others to support our contention that stem cell based therapies will play a significant role in spine surgery in the future.

Cervical motion preservation using mesenchymal progenitor cells and pentosan polysulfate, a novel chondrogenic agent: preliminary study in an ovine model.

OBJECT: There is an unmet need for a procedure that could generate a biological disc substitute while at the same time preserving the normal surgical practice of achieving anterior cervical decompression. The objective of the present study was to test the hypothesis that adult allogeneic mesenchymal progenitor cells (MPCs) formulated with a chondrogenic agent could synthesize a cartilaginous matrix when implanted into a biodegradable carrier and cage, and over time, might serve as a dynamic interbody spacer following anterior cervical discectomy (ACD). METHODS: Eighteen ewes were divided randomly into 3 groups of 6 animals. Each animal was subjected to C3-4 and C4-5 ACD followed by implantation of bioresorbable interbody cages and graft containment plates. The cage was packed with 1 of 3 implants. In Group A, the implant was Gelfoam sponge only. In Group B, the implant consisted of Gelfoam sponge with 1 million MPCs only. In Group C, the implant was Gelfoam sponge with 1 million MPCs formulated with the chondrogenic agent pentosan polysulfate (PPS). In each animal the cartilaginous endplates were retained intact at 1 level, and perforated in a standardized manner at the other level. Allogeneic ovine MPCs were derived from a single batch of immunoselected and culture-expanded MPCs isolated from bone marrow of outbred sheep (mixed stock). Radiological and histological measures were used to assess cartilage formation and the presence or absence of new bone formation. RESULTS: The MPCs with or without PPS were safe and well-tolerated in the ovine cervical spine. There was no significant difference between groups in the radiographic or histological outcome measures, regardless of whether endplates were perforated or retained intact. According to CT scans obtained at 3 months after the operation, new bone formation within the interbody space was observed in the Gelfoam only group (Group A) in 9 (75%) of 12 interbody spaces, and 11 (92%) of 12 animals in the MPC cohort (Group B) had new bone formation within the interbody space. Significantly, in the MPC & PPS group (Group C), there were only 1 (8%) of 12 levels with new bone formation (p = 0.0009 vs Group A; p = 0.0001 vs Group B). According to histological results, there was significantly more cartilaginous tissue within the interbody cages of Group C (MPC & PPS) compared with both the control group (Group A; p = 0.003) and the MPC Group (p = 0.017). CONCLUSIONS: This study demonstrated the feasibility of using MPCs in combination with PPS to produce cartilaginous tissue to replace the intervertebral disc following ACD. This biological approach may offer a means preserving spinal motion and offers an alternative to fusion to artificial prostheses.