Neurovascular Ingrowth Research Articles

Tendinopathy in diabetes mellitus patients-Epidemiology, pathogenesis, and management.

Chronic tendinopathy is a frequent and disabling musculo-skeletal problem affecting the athletic and general populations. The affected tendon is presented with local tenderness, swelling, and pain which restrict the activity of the individual. Tendon degeneration reduces the mechanical strength and predisposes it to rupture. The pathogenic mechanisms of chronic tendinopathy are not fully understood and several major non-mutually exclusive hypotheses including activation of the hypoxia-apoptosis-pro-inflammatory cytokines cascade, neurovascular ingrowth, increased production of neuromediators, and erroneous stem cell differentiation have been proposed. Many intrinsic and extrinsic risk/causative factors can predispose to the development of tendinopathy. Among them, diabetes mellitus is an important risk/causative factor. This review aims to appraise the current literature on the epidemiology and pathology of tendinopathy in diabetic patients. Systematic reviews were done to summarize the literature on (a) the association between diabetes mellitus and tendinopathy/tendon tears, (b) the pathological changes in tendon under diabetic or hyperglycemic conditions, and (c) the effects of diabetes mellitus or hyperglycemia on the outcomes of tendon healing. The potential mechanisms of diabetes mellitus in causing and exacerbating tendinopathy with reference to the major non-mutually exclusive hypotheses of the pathogenic mechanisms of chronic tendinopathy as reported in the literature are also discussed. Potential strategies for the management of tendinopathy in diabetic patients are presented.

Structural and Functional Repair of the Annulus Fibrosus

Degenerated intervertebral discs (IVDs) are commonly implicated in low back pain and herniation is a direct cause of pain and disability. There are few minimally invasive treatments to treat and repair painful IVD degeneration including annulus fibrosus (AF) defects. Painful IVD injuries and degeneration are associated with structural disruption, chronic inflammation, and neurovascular ingrowth deep into the IVD and all of these conditions must be addressed to enhance function and inhibit painful conditions. Design goals for functional AF repair involve the restoration IVD height and motion segment biomechanics to intact conditions, sealing of defects in the AF, no (or minimal) risk of herniation under high physiological load conditions, and the ability to promote healing. Evaluation methods for AF repair strategies should progress from rapid screening studies to more advanced and rigorous evaluations in situ and eventually in vivo. Multiple hydrogel repair strategies are described including sealants and cell carriers. Fibrin crosslinked with geniplin (FibGen) shows promise for AF repair with good performance biomechanically and in organ culture. Disclosures: None

Deep Venous Thrombosis and Tendon Healing.

Tendon metabolism after acute Achilles tendon rupture (ATR) is associated with major complications related to immobilization, which results in reduced circulation, high risk of deep venous thrombosis (DVT), impaired healing and functional deficits.DVT has been demonstrated to occur in up to 50 % of the patients with ATR. Suffering from a DVT during tendon healing has been demonstrated as an independent predictive factor for impaired patient outcome at 1year after ATR, suggesting that specific interventions are warranted to prevent DVT. Since pharmacological DVT prophylaxis has low or no effect during lower leg immobilization it is speculated whether adjuvant treatment with intermittent pneumatic compression (IPC) applied during lower limb immobilization can reduce the incidence of DVT.IPC, which acts through mechanical, chemical and molecular mechanisms, has been demonstrated to enhance neuro-vascular ingrowth in a tendon repair model and stimulate collagen production leading to improved maximum force during healing.Recently, a prospective randomized trial compared adjuvant IPC applied under an orthosis versus plaster cast only in ATR patients. The study found at 2weeks post-operatively 21 % DVTs in the IPC-group compared to 37 % in the control group. Patients that received no IPC treatment exhibited an almost threefold increased odds for DVT, independently of age. Furthermore, using microdialysis technique, adjuvant IPC treatment was shown to increase the metabolic healing activity at 2weeks post-ATR.Tendon healing is impaired by reduced circulation and DVT. The demonstration that adjuvant IPC effectively reduced DVT incidence, and also is capable of enhancing the metabolic response suggests that IPC treatment may not only be a viable means of prophylaxis against DVT, but possibly also a method of promoting healing.

Annular puncture with tumor necrosis factor-alpha injection enhances painful behavior with disc degeneration in vivo

Background ContextPainfulintervertebral disc degeneration is extremely common and costly. Effective treatments are lacking because the nature of discogenic pain is complex with limited capacity to distinguish painful conditions from age-related changes in the spine. Hypothesized sources of discogenic pain include chronic inflammation, neurovascular ingrowth, and structural disruption. PurposeThis study aimed to investigate inflammation, pro-neurovascular growth factors, and structural disruption as sources of painful disc degeneration Study Design/SettingThis study used an in vivo study to address these hypothesized mechanisms with anterior intradiscal injections of tumor necrosis factor-alpha (TNFα), pro-neurovascular growth factors: nerve growth factor and vascular endothelial growth factor (NGF and VEGF), and saline with additional sham surgery and naïve controls. Depth of annular puncture was also evaluated for its effects on structural and painful degeneration. MethodsRat lumbar discs were punctured (shallow or deeper puncture) and intradiscally injected with saline, TNFα, or NGF and VEGF. Structural disc degeneration was assessed using X-ray, magnetic resonance imaging (MRI), and histology. The rat painful condition was evaluated using Von Frey hyperalgesia measurements, and substance P immunostaining in dorsal root ganglion (DRG) was performed to determine the source of pain. ResultsSaline injection increased painful responses with degenerative changes in disc height, MRI intensity, and morphologies of disc structure and cell. TNFα and NGF/VEGF accelerated painful behavior, and TNFα-injected animals had increased substance P in DRGs. Deeper punctures led to more severe disc degeneration. Multiple regression analysis showed that the painful behavior was correlated with disc height loss. ConclusionsWe concluded that rate and severity of structural disc degeneration was associated with the amount of annular disruption and puncture depth. The painful behavior was associated with disc height loss and discal inflammatory state, whereas pro-inflammatory cytokines might play a more important role in the level of pain, which might have resulted from enhanced DRG sensitization. These in vivo painful disc degeneration models with different severities of structural changes may be useful for investigating discogenic pain mechanisms and for screening therapies, although interpretations must note the differences between all surgically induced animal models and the human condition.

Soluble factors from the notochordal-rich intervertebral disc inhibit endothelial cell invasion and vessel formation in the presence and absence of pro-inflammatory cytokines

Chronic low back pain can be associated with the pathological ingrowth of blood vessels and nerves into intervertebral discs (IVDs). The notochord patterns the IVD during development and is a source of anti-angiogenic soluble factors such as Noggin and Chondroitin sulfate (CS). These factors may form the basis for a new minimally invasive strategy to target angiogenesis in the IVD. To examine the anti-angiogenic potential of soluble factors from notochordal cells (NCs) and candidates Noggin and CS under healthy culture conditions and in the presence of pro-inflammatory mediators. NC conditioned media (NCCM) was generated from porcine NC-rich nucleus pulposus tissue. To assess the effects of NCCM, CS and Noggin on angiogenesis, cell invasion and tubular formation assays were performed using human umbilical vein endothelial cells (HUVECs) ± tumor necrosis factor alpha (TNFα [10 ng/ml]). vascular endothelial growth factor (VEGF)-A, MMP-7, interleukin-6 (IL-6) and IL-8 mRNA levels were assessed using qRT-PCR. NCCM (10 & 100%), CS (10 and 100 μg) and Noggin (10 and 100 ng) significantly decreased cell invasion of HUVECs with and without TNFα. NCCM 10% and Noggin 10 ng inhibited tubular formation with and without TNFα and CS 100 μg inhibited tubules in Basal conditions whereas CS 10 μg inhibited tubules with TNFα. NCCM significantly decreased VEGF-A, MMP-7 and IL-6 mRNA levels in HUVECs with and without TNFα. CS and Noggin had no effects on gene expression. We provide the first evidence that soluble factors from NCs can inhibit angiogenesis by suppressing VEGF signaling. Notochordal-derived ligands are a promising minimally invasive strategy targeting neurovascular ingrowth and pain in the degenerated IVD.

Effect of Lumbar Intradiscal Injection of Tumor Necrosis Factor-Alpha and Nerve Growth Factor/Vascular Endothelial Growth Factor on Disc Degeneration, Pain Behavior and Neurovascular Ingrowth in an In Vivo Rat Model

Effect of Lumbar Intradiscal Injection of Tumor Necrosis Factor-Alpha and Nerve Growth Factor/Vascular Endothelial Growth Factor on Disc Degeneration, Pain Behavior and Neurovascular Ingrowth in an In Vivo Rat Model

Molecular Basis of Intervertebral Disc Degeneration and Herniations: What Are the Important Translational Questions?

Intervertebral disc degeneration is a common condition with few inexpensive and effective modes of treatment, but current investigations seek to clarify the underlying process and offer new treatment options. It will be important for physicians to understand the molecular basis for the pathology and how it translates to developing clinical treatments for disc degeneration. In this review, we sought to summarize for clinicians what is known about the molecular processes that causes disc degeneration. A healthy disc requires maintenance of a homeostatic environment, and when disrupted, a catabolic cascade of events occurs on a molecular level resulting in upregulation of proinflammatory cytokines, increased degradative enzymes, and a loss of matrix proteins. This promotes degenerative changes and occasional neurovascular ingrowth potentially contributing to the development of pain. Research demonstrates the molecular changes underlying the harmful effects of aging, smoking, and obesity seen clinically while demonstrating the variable influence of exercise. Finally, oral medications, supplements, biologic treatments, gene therapy, and stem cells hold great promise but require cautious application until their safety profiles are better outlined. Intervertebral disc degeneration occurs where there is a loss of homeostatic balance with a predominantly catabolic metabolic profile. A basic understanding of the molecular changes occurring in the degenerating disc is important for practicing clinicians because it may help them to inform patients to alter lifestyle choices, identify beneficial or harmful supplements, or offer new biologic, genetic, or stem cell therapies.

Notochordal Cell-Derived Therapies for Painful Disc Degeneration

Degenerated human intervertebral discs (IVDs) are commonly associated with low back pain conditions including stenosis, herniation, and axial back pain. There are few minimally invasive treatments available to treat painful IVD degeneration and axial back pain. Painful IVD degeneration is associated with structural disruption, chronic inflammation, and neurovascular ingrowth deep into the IVD and all of these conditions must be addressed to promote function and inhibit painful conditions in the spine. During development of the spine, the notochord produces factors including chondroitin sulfate that are important for creating the IVD structure as well as for patterning the spine to inhibit ingrowth of dorsal root ganglia (DRGs) axons into the IVD. Our overall concept is that we can use concepts and factors important in the developing spine to develop therapeutic candidates for painful IVD degeneration. In this study, we focus on improved understanding of notochordal cells, their maturation, and the secreted factors they produce that can be used for therapeutic effect. We focus on three studies. Study 1 describes the effects of loading on the notochordal cell phenotype. Study 2 identifies cell culture models to screen therapies for discogenic pain and to investigate which factors in notochordal cell conditioned medium are capable of inhibiting neurite growth. Study 3 investigates the capacity of notochordal cell conditioned medium to inhibit the neovascularization that may be a predecessor to neurite growth into the IVD. We use a variety of cell culture models using nucleus pulposus tissue from porcine IVDs and neural and vascular endothelial cells from human and rodent sources. Study 1 indicated that hydrostatic loading can induce maturation of notochordal cells from large vacuolated to small chondrocytic cells. This maturation was accompanied by some shifts in the quantity and quality of secreted factors. Study 2 determined that notochordal cell conditioned medium is capable of inhibiting neurite growth in two neural cell types: human neuroblastoma cells (SH-SY5Ys) and rat DRGs. Results indicated that the effect was dose-specific, and that CS is a necessary protein causing this inhibition. Study 3 determined that notochordal cell conditioned medium can inhibit neovascularization using cell human umbilical vascular endothelial cells. Importantly, notochordal cell conditioned medium was not cytotoxic to neural or endothelial cells, suggesting this could be used safely for back pain even if leakage occurred in the area around the IVD. However, future in vivo testing is required for further characterization of safety and efficacy profiles. Together, we present a novel paradigm for designing therapeutic interventions derived from trophic agents from notochordal cells and for screening the efficacy of those therapies. We conclude that notochordal cells secrete factors with structure and symptom modifying potential, and these may be useful for developing therapeutic agents with the capacity to restore IVD function and preventing discogenic pain. Acknowledgments This work was funded by NIAMS/NIH Grants (R01 AR064157 R01, R01 AR057397) and Grant S-13-50P of the AO Research Fund of the AO Foundation (D.P.) and Orthopaedic Research and Education Foundation (S.K.C.). Disclosure of Interest None declared

Hyperemia in Plantar Fasciitis Determined by Power Doppler Ultrasound

Cross-sectional observational study. To investigate the presence of soft tissue hyperemia in plantar fasciitis with power Doppler ultrasound. Localized hyperemia is an established feature of tendinopathy, suggesting that neurovascular in-growth may contribute to tendon-associated pain in some patients. The presence of abnormal soft tissue vascularity can be assessed with Doppler ultrasound, and a positive finding can assist with targeted treatment plans. However, very little is known regarding the presence of hyperemia in plantar fasciitis and the ability of routine Doppler ultrasound to identify vascular in-growth in the plantar fascia near its proximal insertion. This observational study included 30 participants with plantar fasciitis unrelated to systemic disease and 30 age- and sex-matched controls. Ultrasound examination was performed with a 13- to 5-MHz linear transducer, and power Doppler images were assessed by 2 blinded investigators. Hyperemia of the plantar fascia was present in 8 of 30 participants with plantar fasciitis and in 2 of 30 controls. The between-group difference for hyperemia, using a 4-point scale, was statistically significant, with participants with plantar fasciitis showing increased Doppler ultrasound signal compared to controls (Mann-Whitney U, P = .03). However, the majority of participants with plantar fasciitis with evidence of hyperemia demonstrated very mild color changes, and only 3 were found to have moderate or marked hyperemia. Mild hyperemia can occur with plantar fasciitis, but most individuals will not exhibit greater soft tissue vascularity when assessed with routine Doppler ultrasound. Clinicians treating plantar fasciitis should not consider a positive Doppler signal as essential for diagnosis of the condition but, rather, as a feature that may help to refine the treatment plan for an individual patient.

Notochordal Cell–Derived Therapeutic Strategies for Discogenic Back Pain

An understanding of the processes that occur during development of the intervertebral disk can help inform therapeutic strategies for discogenic pain. This article reviews the literature to identify candidates that are found in or derived from the notochord or notochordal cells and evaluates the theory that such factors could be isolated and used as biologics to target the structural disruption, inflammation, and neurovascular ingrowth often associated with discogenic back pain. A systematic review using PubMed was performed with a primary search using keywords “(notochordal OR notochord) And (nerves OR blood vessels OR SHH OR chondroitin sulfate OR notch OR CTGF) NOT chordoma.” Secondary searches involved keywords associated with the intervertebral disk and pain. Several potential therapeutic candidates from the notochord and their possible targets were identified. Studies are needed to further identify candidates, explore mechanisms for effect, and to validate the theory that these candidates can promote structural restoration and limit or inhibit neurovascular ingrowth using in vivo studies.

Increased Expression of Netrin-1 and Its Deleted in Colorectal Cancer Receptor in Human Diseased Lumbar Intervertebral Disc Compared With Autopsy Control

The expression of netrin-1 and its deleted in colorectal cancer (DCC) receptor was investigated in human lumbar discs using immunohistochemistry. To investigate the expression of netrin-1 and DCC receptor in human diseased and healthy lumbar intervertebral discs (IVDs) and to clarify the correlation between netrin-1 expression and the degree of neurovascular ingrowth. Previous studies have shown neurovascular ingrowth into the inner regions of degenerated IVD and suggested that the ingrowth may contribute to discogenic low back pain. Netrin-1 is an axon guidance molecule that regulates axons seeking their final targets and has been identified as involved in various pathological conditions, so is its DCC receptor. However, the role of netrin-1 in diseased IVDs remains unknown. Thirty-five diseased IVD specimens were collected from 34 patients with different lumbar diseases during posterior lumbar interbody fusion. Eight normal discs were obtained at autopsy as control. Using polyclonal or monoclonal antibody, the disc slides were immmunostained to detect the expression and distribution of netrin-1, the DCC, the neuronal marker (neurofilament), and the vascular endothelial cell marker (CD34). Netrin-1 and DCC immunopositive cells distributed substantially from the annulus fibrosus to the nucleus pulposus (NP), and the immunopositivity was detected in the disc cells, endothelial cells and granulation tissue cells in the diseased discs. The percentage of netrin-1 positive disc cells of the NP was more than that of the annulus fibrosus. The expression of netrin-1 and DCC was weak in the normal discs. A significant positive correlation between the percentage of netrin-1 immunopositive disc cells and neurovascular scores was found. The increased expression of netrin-1 and DCC in diseased IVDs compared with controls suggested that they might play an important role in the process of neurovascular ingrowth.

Intermittent pneumatic compression enhances neurovascular ingrowth and tissue proliferation during connective tissue healing: A study in the rat

Intermittent pneumatic compression (IPC) is a treatment method to decrease venous stasis and stimulate blood flow. Recently, it was hypothesized that IPC may exert positive effects on tissue healing, a process highly dependent upon adequate circulation. In this study, we investigated the effects of daily 1-h IPC treatment during 2 and 4 weeks post-rat Achilles tendon rupture. The tendons were subjectively and semiquantitatively analyzed for collagen organization, fibroblast density, angiogenesis, and the occurrence of sensory neuropeptides, substance P (SP) and calcitonine gene related peptide (CGRP), as well as for a nerve regeneration marker, growth associated protein 43 (GAP-43). After 2 weeks of treatment, fibroblast density increased by 53% (p = 0.0004), vessel density by 64% (p = 0.022), and the occurrence of SP by 110% (p = 0.047) and CGRP by 47% (p = 0.0163) compared to untreated controls. Following 4 weeks of treatment, both the occurrence of sensory neuropeptides and the vessel density remained significantly higher (p < 0.05), whereas fibroblast density returned to normal. However, at 4 weeks the treated tendons displayed a higher degree of organized parallel collagen fibers, a sign of increased maturation. Daily IPC treatment improves neurovascular ingrowth and fibroblast proliferation in the healing tendon and may accelerate the repair process.

Neovascularisation in chronic tendon injuries detected with colour Doppler ultrasound in horse and man: implications for research and treatment

Recent research on chronic painful Achilles tendons in humans using ultrasonography and immunohistochemistry, has demonstrated an association between neurovascular ingrowth and tendon pain. In horses, chronic debilitating tendon conditions are well-known to be very difficult to treat, and the background to impaired function and pain is not scientifically clarified. In a collaborative research project between the Sports Medicine Unit in Umeå and Strömsholm Equine Hospital, grey-scale ultrasonography (US) and colour Doppler (CD) examination were performed in ten horses with chronic tendon injuries (>3 months) and a control group of six healthy and asymptomatic horses. In all symptomatic tendons, but not in any of the tendons in the control group, neovessels were seen in the area with structural tendon changes. The neovessels found in the horse tendons looked similar to what has recently been presented in human Achilles tendons. These findings motivate evaluation of the same treatment, a sclerosing injection that was demonstrated recently to give promising results in the treatment of chronic Achilles tendon injuries (tendinosis) in humans.