57829736
Functional and Structural Changes of the Blood-Nerve-Barrier in Diabetic Neuropathy
The incidence of diabetes mellitus is approaching global epidemic proportions and should be considered a major health-care problem of modern societies in the twenty-first century. Diabetic neuropathy is a common chronic complication of diabetes and, although an adequate glycemic control can reduce the frequency of diabetic neuropathy in type 1 diabetes, the majority of type 2 diabetic patients will develop this complication. The underlying cellular and molecular mechanisms are still poorly understood, preventing the development of effective treatment strategies. However, accumulating evidence suggests that breakdown of the blood-nerve barrier (BNB) plays a pivotal pathophysiological role in diabetic neuropathy. In the present review, we highlight the structural and functional significance of the BNB in health and disease, focusing on the pathological molecular events leading to BNB dysfunction in diabetic neuropathy. In addition, we discuss potential molecular targets involved in BNB homeostasis that may pave the way toward novel therapeutic strategies for treating diabetic neuropathy.
24247108
Prediabetes diagnosis and treatment: A review.
Prediabetes is an intermediate state of hyperglycemia with glycemic parameters above normal but below the diabetes threshold. While, the diagnostic criteria of prediabetes are not uniform across various international professional organizations, it remains a state of high risk for developing diabetes with yearly conversion rate of 5%-10%. Observational evidence suggests as association between prediabetes and complications of diabetes such early nephropathy, small fiber neuropathy, early retinopathy and risk of macrovascular disease. Several studies have shown efficacy of lifestyle interventions with regards to diabetes prevention with a relative risk reduction of 40%-70% in adults with prediabetes. While there is increasing evidence to prove the efficacy of pharmacotherapy in prevention of diabetes in adults with prediabetes, pharmaceutical treatment options other than metformin are associated with adverse effects that limit their use for prediabetes. There are no reports of systematic evaluation of health outcomes related to prediabetes in children. The effects of pharmacotherapy of prediabetes on growth and pubertal development in children remains unknown. Secondary intervention with pharmacotherapy with metformin is advocated for high-risk individuals but criteria for such consideration benefit of early intervention, long term cost effectiveness of such interventions and the end point of therapy remain unclear. Pharmacotherapy must be used with caution in children with prediabetes. Prediabetes is a condition defined as having blood glucose levels above normal but below the defined threshold of diabetes. It is considered to be an at risk state, with high chances of developing diabetes. While, prediabetes is commonly an asymptomatic condition, there is always presence of prediabetes before the onset of diabetes. The elevation of blood sugar is a continuum and hence prediabetes can not be considered an entirely benign condition. This aim of this review is to describe the challenges associated with diagnosis of prediabetes, the possible adverse medical outcomes associated with prediabetes and the treatment options and rationale for their use in context of prediabetes.
39771424
Painful diabetic neuropathy
Chronic painful diabetic neuropathy causes symptoms that can last for many years and severely impair the quality of life of affected patients. This review describes the epidemiology, pathophysiology and treatment of chronic neuropathic pain. Particular emphasis is placed on a comprehensive review of the management of painful symptoms through a detailed review of the published literature using a variety of databases particularly Medline and EMBASE.
207218086
New Horizons in Diabetic Neuropathy: Mechanisms, Bioenergetics, and Pain
Pre-diabetes and diabetes are a global epidemic, and the associated neuropathic complications create a substantial burden on both the afflicted patients and society as a whole. Given the enormity of the problem and the lack of effective therapies, there is a pressing need to understand the mechanisms underlying diabetic neuropathy (DN). In this review, we present the structural components of the peripheral nervous system that underlie its susceptibility to metabolic insults and then discuss the pathways that contribute to peripheral nerve injury in DN. We also discuss systems biology insights gleaned from the recent advances in biotechnology and bioinformatics, emerging ideas centered on the axon-Schwann cell relationship and associated bioenergetic crosstalk, and the rapid expansion of our knowledge of the mechanisms contributing to neuropathic pain in diabetes. These recent advances in our understanding of DN pathogenesis are paving the way for critical mechanism-based therapy development.
4909074
Transcriptional networks of progressive diabetic peripheral neuropathy in the db/db mouse model of type 2 diabetes: An inflammatory story
ABSTRACT Diabetic peripheral neuropathy is the most common complication of diabetes and a source of considerable morbidity. Numerous molecular pathways are linked to neuropathic progression, but it is unclear whether these pathways are altered throughout the course of disease. Moreover, the methods by which these molecular pathways are analyzed can produce significantly different results; as such it is often unclear whether previously published pathways are viable targets for novel therapeutic approaches. In the current study we examine changes in gene expression patterns in the sciatic nerve (SCN) and dorsal root ganglia (DRG) of db/db diabetic mice at 8, 16, and 24weeks of age using microarray analysis. Following the collection and verification of gene expression data, we utilized both self‐organizing map (SOM) analysis and differentially expressed gene (DEG) analysis to detect pathways that were altered at all time points. Though there was some variability between SOM and DEG analyses, we consistently detected altered immune pathways in both the SCN and DRG over the course of disease. To support these results, we further used multiplex analysis to assess protein changes in the SCN of diabetic mice; we found that multiple immune molecules were upregulated at both early and later stages of disease. In particular, we found that matrix metalloproteinase‐12 was highly upregulated in microarray and multiplex data sets suggesting it may play a role in disease progression. HIGHLIGHTSWe examined longitudinal transcriptomics patterns in SCN and DRG of db/db mice.Genes/pathways regulated across all stages of DPN are promising therapeutic targets.Immune‐associated pathways were increased in both SCN and DRG at all time points.IFN‐&ggr;, IL‐10, and MMP‐12 proteins were upregulated in SCN during early and late DPN.These data indicate specific immune system involvement at all stages of DPN.
44873749
Vascular factors and metabolic interactions in the pathogenesis of diabetic neuropathy
Abstract. Diabetes mellitus is a major cause of peripheral neuropathy, commonly manifested as distal symmetrical polyneuropathy. This review examines evidence for the importance of vascular factors and their metabolic substrate from human and animal studies. Diabetic neuropathy is associated with risk factors for macrovascular disease and with other microvascular complications such as poor metabolic control, dyslipidaemia, body mass index, smoking, microalbuminuria and retinopathy. Studies in human and animal models have shown reduced nerve perfusion and endoneurial hypoxia. Investigations on biopsy material from patients with mild to severe neuropathy show graded structural changes in nerve microvasculature including basement membrane thickening, pericyte degeneration and endothelial cell hyperplasia. Arterio-venous shunting also contributes to reduced endoneurial perfusion. These vascular changes strongly correlate with clinical defects and nerve pathology. Vasodilator treatment in patients and animals improves nerve function. Early vasa nervorum functional changes are caused by the metabolic insults of diabetes, the balance between vasodilation and vasoconstriction is altered. Vascular endothelium is particularly vulnerable, with deficits in the major endothelial vasodilators, nitric oxide, endothelium-derived hyperpolarising factor and prostacyclin. Hyperglycaemia and dyslipidaemia driven oxidative stress is a major contributor, enhanced by advanced glycation end product formation and polyol pathway activation. These are coupled to protein kinase C activation and ω-6 essential fatty acid dysmetabolism. Together, this complex of interacting metabolic factors accounts for endothelial dysfunction, reduced nerve perfusion and function. Thus, the evidence emphasises the importance of vascular dysfunction, driven by metabolic change, as a cause of diabetic neuropathy, and highlights potential therapeutic approaches. [Diabetologia (2001) 44: 1973–1988]
1035214
Advances in the epidemiology, pathogenesis and management of diabetic peripheral neuropathy
Diabetic peripheral neuropathy (DPN) affects up to 50% of patients with diabetes and is a major cause of morbidity and increased mortality. Its clinical manifestations include painful neuropathic symptoms and insensitivity, which increases the risk for burns, injuries and foot ulceration.
12517015
Alterations in mRNA Expression of Myelin Proteins in the Sciatic Nerves and Brains of Streptozotocin-induced Diabetic Rats
Diabetic neuropathy is the most common complication of diabetes. We examined the levels and the mRNA expression of myelin proteins in the sciatic nerves and the brains of streptozotocin-induced diabetic rats. The diabetic rats exhibited a decrease in body weight, elevation of the blood glucose level and a decrease in motor nerve conduction velocity at 2 weeks after streptozotocin injection. In the sciatic nerves of diabetic rats, the level of P0 protein and its mRNA expression were markedly reduced at 20 weeks after the injection. In the brains, the levels of proteolipid protein and myelin-associated glycoprotein and their mRNA expression were selectively decreased at 20 weeks after the injection. This affected expression of myelin proteins was found even when no histological abnormalities were detectable. Considering the functional significance of myelin proteins, this impairment of protein expression is possibly involved in the pathogenesis of diabetic neuropathy, including that in brain disorders.
2961276
Loss of the tight junction proteins occludin and zonula occludens-1 from cerebral vascular endothelium during neutrophil-induced blood–brain barrier breakdown in vivo

The tight junctions found between cerebral vascular endothelial cells form the basis of the blood-brain barrier. Breakdown of the blood-brain barrier is a feature of a variety of CNS pathologies that are characterized by extensive leucocyte recruitment, such as multiple sclerosis and stroke. The molecular mechanisms associated with opening of the blood-brain barrier and leucocyte recruitment in vivo are currently poorly understood. We have used an in vivo rat model to investigate the molecular response of the CNS endothelium to neutrophil adhesion and migration. Injection of interleukin-1 beta into the striatum of juvenile brains results in a neutrophil-dependent increase in vessel permeability at 4 h. Only a subset of blood vessels were associated with neutrophil recruitment. These particular vessels displayed an increase in phosphotyrosine staining, loss of the tight junctional proteins, occludin and zonula occludens-1, and apparent redistribution of the adherens junction protein vinculin. Examination of these vessels under the electron microscope indicated that the cell-cell adhesions in such vessels are morphologically different from normal junctions. This study provides the first direct evidence in vivo that leucocyte recruitment can trigger signal transduction cascades leading to junctional disorganization and blood-brain barrier breakdown. Our results have established an endothelial cell molecular profile associated with leucocyte-induced blood-brain barrier breakdown in vivo, and the relevance of different in vitro cell culture models may now be viewed more objectively.
7506197
Impaired Hedgehog signalling-induced endothelial dysfunction is sufficient to induce neuropathy: implication in diabetes.
AIMS
Microangiopathy, i.e. endothelial dysfunction, has long been suggested to contribute to the development of diabetic neuropathy, although this has never been fully verified. In the present paper, we have identified the role of Hedgehog (Hh) signalling in endoneurial microvessel integrity and evaluated the impact of impaired Hh signalling in endothelial cells (ECs) on nerve function.


METHODS AND RESULTS
By using Desert Hedgehog (Dhh)-deficient mice, we have revealed, that in the absence of Dhh, endoneurial capillaries are abnormally dense and permeable. Furthermore, Smoothened (Smo) conditional KO mice clarified that this increased vessel permeability is specifically due to impaired Hh signalling in ECs and is associated with a down-regulation of Claudin5 (Cldn5). Moreover, impairment of Hh signalling in ECs was sufficient to induce hypoalgesia and neuropathic pain. Finally in Lepr(db/db) type 2 diabetic mice, the loss of Dhh expression observed in the nerve was shown to be associated with increased endoneurial capillary permeability and decreased Cldn5 expression. Conversely, systemic administration of the Smo agonist SAG increased Cldn5 expression, decreased endoneurial capillary permeability, and restored thermal algesia to diabetic mice, demonstrating that loss of Dhh expression is crucial in the development of diabetic neuropathy.


CONCLUSION
The present work demonstrates the critical role of Dhh in maintaining blood nerve barrier integrity and demonstrates for the first time that endothelial dysfunction is sufficient to induce neuropathy.
31539786
Basement membrane reduplication and pericyte degeneration precede development of diabetic polyneuropathy and are associated with its severity
In a recent paper, we showed that the number of endoneurial microvessels per square millimeter and the average luminal area and size distribution of these microvessels are not significantly different in sural nerves of patients with diabetes mellitus as compared to control subjects. Mural area, especially the component due to basement membrane reduplication and cellular debris, was unequivocally increased in diabetes mellitus. Because these latter changes are associated with a decrease in periendothelial cell area, we hypothesized that cellular degeneration, especially of pericytes, may account for basement membrane reduplication and increased frequency of cellular debris. In the present study, we showed that endoneurial microvessels undergo a statistically significant increase in basement membrane area, mural area, and frequency of cellular debris in diabetics without polyneuropathy and an even greater increase in diabetics with polyneuropathy. We also found that duration of diabetes mellitus was significantly associated with area occupied by reduplicated basement membrane and cellular debris, but not with mural and periendothelial area. None of the examined measurements was associated with age. Since the microvessel abnormalities we describe are already present before the development of polyneuropathy and increase with severity of polyneuropathy, it is likely that they reflect functional derangements of pericytes and microvessel function which precede and might be implicated in fiber degeneration.
22870488
Neurovascular dysfunction and neurodegeneration in dementia and Alzheimer's disease.
Vascular insults can initiate a cascade of molecular events leading to neurodegeneration, cognitive impairment, and dementia. Here, we review the cellular and molecular mechanisms in cerebral blood vessels and the pathophysiological events leading to cerebral blood flow dysregulation and disruption of the neurovascular unit and the blood-brain barrier, which all may contribute to the onset and progression of dementia and Alzheimer's disease (AD). Particularly, we examine the link between neurovascular dysfunction and neurodegeneration including the effects of AD genetic risk factors on cerebrovascular functions and clearance of Alzheimer's amyloid-β peptide toxin, and the impact of vascular risk factors, environment, and lifestyle on cerebral blood vessels, which in turn may affect synaptic, neuronal, and cognitive functions. Finally, we examine potential experimental treatments for dementia and AD based on the neurovascular model, and discuss some critical questions to be addressed by future studies. This article is part of a Special Issue entitled: Vascular Contributions to Cognitive Impairment and Dementia edited by M. Paul Murphy, Roderick A. Corriveau and Donna M. Wilcock.
3483915
Blood–brain barrier breakdown in Alzheimer disease and other neurodegenerative disorders
The blood–brain barrier (BBB) is a continuous endothelial membrane within brain microvessels that has sealed cell-to-cell contacts and is sheathed by mural vascular cells and perivascular astrocyte end-feet. The BBB protects neurons from factors present in the systemic circulation and maintains the highly regulated CNS internal milieu, which is required for proper synaptic and neuronal functioning. BBB disruption allows influx into the brain of neurotoxic blood-derived debris, cells and microbial pathogens and is associated with inflammatory and immune responses, which can initiate multiple pathways of neurodegeneration. This Review discusses neuroimaging studies in the living human brain and post-mortem tissue as well as biomarker studies demonstrating BBB breakdown in Alzheimer disease, Parkinson disease, Huntington disease, amyotrophic lateral sclerosis, multiple sclerosis, HIV-1-associated dementia and chronic traumatic encephalopathy. The pathogenic mechanisms by which BBB breakdown leads to neuronal injury, synaptic dysfunction, loss of neuronal connectivity and neurodegeneration are described. The importance of a healthy BBB for therapeutic drug delivery and the adverse effects of disease-initiated, pathological BBB breakdown in relation to brain delivery of neuropharmaceuticals are briefly discussed. Finally, future directions, gaps in the field and opportunities to control the course of neurological diseases by targeting the BBB are presented.
26002631
Barrier function in the peripheral and central nervous system—a review
The peripheral (PNS) and central nervous system (CNS) are delicate structures, highly sensitive to homeostatic changes—and crucial for basic vital functions. Thus, a selection of barriers ensures the protection of the nervous system from noxious blood-borne or surrounding stimuli. In this chapter, anatomy and functioning of the blood–nerve (BNB), the blood–brain (BBB), and the blood–spinal cord barriers (BSCB) are presented and the key tight junction (TJ) proteins described: claudin-1, claudin-3, claudin-5, claudin-11, claudin-12, claudin-19, occludin, Zona occludens-1 (ZO-1), and tricellulin are by now identified as relevant for nerval barriers. Different diseases can lead to or be accompanied by neural barrier disruption, and impairment of these barriers worsens pathology. Peripheral nerve injury and inflammatory polyneuropathy cause an increased permeability of BNB as well as BSCB, while, e.g., diseases of the CNS such as amyotrophic lateral sclerosis, multiple sclerosis, spinal cord injury, or Alzheimer’s disease can progress and worsen through barrier dysfunction. Moreover, the complex role and regulation of the BBB after ischemic stroke is described. On the other side, PNS and CNS barriers hamper the delivery of drugs in diseases when the barrier is intact, e.g., in certain neurodegenerative diseases or inflammatory pain. Understanding of the barrier - regulating processes has already lead to the discovery of new molecules as drug enhancers. In summary, the knowledge of all of these mechanisms might ultimately lead to the invention of drugs to control barrier function to help ameliorating or curing neurological diseases.
14753395
Structure and function of the blood–brain barrier
Neural signalling within the central nervous system (CNS) requires a highly controlled microenvironment. Cells at three key interfaces form barriers between the blood and the CNS: the blood-brain barrier (BBB), blood-CSF barrier and the arachnoid barrier. The BBB at the level of brain microvessel endothelium is the major site of blood-CNS exchange. The structure and function of the BBB is summarised, the physical barrier formed by the endothelial tight junctions, and the transport barrier resulting from membrane transporters and vesicular mechanisms. The roles of associated cells are outlined, especially the endfeet of astrocytic glial cells, and pericytes and microglia. The embryonic development of the BBB, and changes in pathology are described. The BBB is subject to short and long-term regulation, which may be disturbed in pathology. Any programme for drug discovery or delivery, to target or avoid the CNS, needs to consider the special features of the BBB.
9273181
Blood Brain Barrier: The Role of GAD Antibodies in Psychiatry.
OBJECTIVE
Goal of our case control study was to establish the presence of antibodies to glutamic acid decarboxylase (GAD) in patients with chronic psychotic disorders.


METHODS
Serum levels of GAD antibodies in 12 patients with chronic psychotic disorders (schizophrenia and schizoaffective disorders) and 10 age-matched healthy control subjects were evaluated utilizing enzyme linked immunosorbitent assay (ELISA).


RESULTS
Antibodies to GAD in patients with chronic psychotic disorders have a higher mean than nonpatient control individuals.


CONCLUSION
Our findings provide the first in-vivo evidence of positive GAD antibodies in chronic psychotic disorders and potentially may be used as a screening for these disorders.
40194749
Tight junction proteins: from barrier to tumorigenesis.
The tight junction is a multi-protein complex and is the apical most junctional complex in certain epithelial and endothelial cells. A great deal of attention has been devoted to the understanding of these proteins in contributing to the barrier function - that is, regulating the paracellular flux or permeability between adjacent cells. However, tight junction proteins are now recognized as having functions beyond the barrier. The focus of this review is to discuss the barrier function of the tight junction and to summarize the literature with a focus on the role of tight junction proteins in proliferation, transformation, and metastasis.
14085307
Size-selective loosening of the blood-brain barrier in claudin-5–deficient mice
Tight junctions are well-developed between adjacent endothelial cells of blood vessels in the central nervous system, and play a central role in establishing the blood-brain barrier (BBB). Claudin-5 is a major cell adhesion molecule of tight junctions in brain endothelial cells. To examine its possible involvement in the BBB, claudin-5–deficient mice were generated. In the brains of these mice, the development and morphology of blood vessels were not altered, showing no bleeding or edema. However, tracer experiments and magnetic resonance imaging revealed that in these mice, the BBB against small molecules (<800 D), but not larger molecules, was selectively affected. This unexpected finding (i.e., the size-selective loosening of the BBB) not only provides new insight into the basic molecular physiology of BBB but also opens a new way to deliver potential drugs across the BBB into the central nervous system.
19925658
Localization of claudin-3 in tight junctions of the blood-brain barrier is selectively lost during experimental autoimmune encephalomyelitis and human glioblastoma multiforme
In the central nervous system (CNS) complex endothelial tight junctions (TJs) form a restrictive paracellular diffusion barrier, the blood-brain barrier (BBB). During inflammation, BBB properties are frequently lost, resulting in brain edema. To investigate whether BBB leakiness correlates with molecular changes at BBB TJs, we performed immunofluorescence stainings for TJ molecules in a mouse model of experimental autoimmune encephalomyelitis (EAE) and in human tissue with glioblastoma multiforme (GBM). In TJs of healthy CNS vessels in both mouse and man we detected occludin, ZO-1, claudin-5 and claudin-3. In EAE brain and spinal cord sections we observed the selective loss of claudin-3 immunostaining from TJs of venules surrounded by inflammatory cuffs, whereas the localization of the other TJ proteins remained unchanged. In addition, selective loss of claudin-3 immunostaining was also observed in altered cerebral microvessels of human GBM. Our data demonstrate the selective loss of claudin-3 from BBB TJs under pathological conditions such as EAE or GBM when the integrity of the BBB is compromised, and therefore suggest that claudin-3 is a central component determining the integrity of BBB TJs in vivo.
17320173
Tight Junction Proteins ZO-1, Occludin, and Claudins in Developing and Adult Human Perineurium
In peripheral nerves, groups of Schwann cell–axon units are isolated from the adjacent tissues by the perineurium, which creates a diffusion barrier responsible for the maintenance of endoneurial homeostasis. The perineurium is formed by concentric layers of overlapping, polygonal perineurial cells that form tight junctions at their interdigitating cell borders. In this study, employing indirect immunofluorescence and immunoelectron microscopy, we demonstrate that claudin-1 and -3, ZO-1, and occludin, but not claudin-2, -4, and -5, are expressed in the perineurium of adult human peripheral nerve. We also describe the expression of occludin, ZO-1, claudin-1, -3, and -5 in the developing human perineurium, showing that the expressions of claudin-1 and -3, ZO-1, and occludin follow similar spatial developmental expression patterns but follow different timetables in achieving their respective adult distributions. Specifically, claudin-1 is already largely restricted to perineurium-derived structures at 11 fetal weeks, whereas claudin-3 and occludin are weakly expressed in the perineurial structures at this age and acquire a well-defined perineurial distribution only between 22 and 35 fetal weeks. ZO-1 appears to acquire its mature profile even later during the third trimester. The results of the present and previous studies show that the perineurial diffusion barrier matures relatively late during human peripheral nerve development.
23752159
Advanced glycation end-products induce basement membrane hypertrophy in endoneurial microvessels and disrupt the blood–nerve barrier by stimulating the release of TGF-β and vascular endothelial growth factor (VEGF) by pericytes
Aims/hypothesisThe breakdown of the blood–nerve barrier (BNB) is considered to be a key step in diabetic neuropathy. Although basement membrane hypertrophy and breakdown of the BNB are characteristic features of diabetic neuropathy, the underlying pathogenesis remains unclear. The purpose of the present study was to identify the possible mechanisms responsible for inducing the hypertrophy of basement membrane and the disruption of the BNB after exposure to AGEs.MethodsThe newly established human peripheral nerve microvascular endothelial cell (PnMEC) and pericyte cell lines were used to elucidate which cell types constituting the BNB regulate the basement membrane and to investigate the effect of AGEs on the basement membrane of the BNB using western blot analysis.ResultsFibronectin, collagen type IV and tissue inhibitor of metalloproteinase (TIMP-1) protein were produced mainly by peripheral nerve pericytes, indicating that the basement membrane of the BNB is regulated mainly by these cells. AGEs reduced the production of claudin-5 in PnMECs by increasing autocrine signalling through vascular endothelial growth factor (VEGF) secreted by the PnMECs themselves. Furthermore, AGEs increased the amount of fibronectin, collagen type IV and TIMP-1 in pericytes through a similar upregulation of autocrine VEGF and transforming growth factor (TGF)-β released by pericytes.Conclusions/interpretationThese results indicate that pericytes may be the main regulators of the basement membrane at the BNB. AGEs induce basement membrane hypertrophy and disrupt the BNB by increasing autocrine VEGF and TGF-β signalling by pericytes under diabetic conditions.
3286791
The Human Blood-Nerve Barrier Transcriptome
The blood-nerve barrier (BNB), formed by tight junction-forming microvessels within peripheral nerve endoneurium, exists to regulate its internal microenvironment essential for effective axonal signal transduction. Relatively little is known about the unique human BNB molecular composition. Such knowledge is crucial to comprehend the relationships between the systemic circulation and peripheral nerves in health, adaptations to intrinsic or extrinsic perturbations and alterations that may result in disease. We performed RNA-sequencing on cultured early- and late-passage adult primary human endoneurial endothelial cells and laser-capture microdissected endoneurial microvessels from four cryopreserved normal adult human sural nerves referenced to the Genome Reference Consortium Human Reference 37 genome browser, using predefined criteria guided by known transcript or protein expression in vitro and in situ. We identified 12881 common transcripts associated by 125 independent biological networks, defined as the normal adult BNB transcriptome, including a comprehensive array of transporters and specialized intercellular junctional complex components. These identified transcripts and their interacting networks provide insights into peripheral nerve microvascular morphogenesis, restrictive barrier formation, influx and efflux transporters with relevance to understanding peripheral nerve homeostasis and pharmacology, including targeted drug delivery and the mediators of leukocyte trafficking in peripheral nerves during normal immunosurveillance.
6359867
The Spared Nerve Injury (SNI) Model of Induced Mechanical Allodynia in Mice
Peripheral neuropathic pain is a severe chronic pain condition which may result from trauma to sensory nerves in the peripheral nervous system. The spared nerve injury (SNI) model induces symptoms of neuropathic pain such as mechanical allodynia i.e. pain due to tactile stimuli that do not normally provoke a painful response [1]. The SNI mouse model involves ligation of two of the three branches of the sciatic nerve (the tibial nerve and the common peroneal nerve), while the sural nerve is left intact [2]. The lesion results in marked hypersensitivity in the lateral area of the paw, which is innervated by the spared sural nerve. The non-operated side of the mouse can be used as a control. The advantages of the SNI model are the robustness of the response and that it doesn’t require expert microsurgical skills. The threshold for mechanical pain response is determined by testing with von Frey filaments of increasing bending force, which are repetitively pressed against the lateral area of the paw [3], [4]. A positive pain reaction is defined as sudden paw withdrawal, flinching and/or paw licking induced by the filament. A positive response in three out of five repetitive stimuli is defined as the pain threshold. As demonstrated in the video protocol, C57BL/6 mice experience profound allodynia as early as the day following surgery and maintain this for several weeks.
12205090
Blood-nerve barrier dysfunction contributes to the generation of neuropathic pain and allows targeting of injured nerves for pain relief
Summary Nerve injury leads to a persistent disruption of the blood‐nerve barrier, which allows the entrance of blood‐borne pronociceptive molecules and cells, contributing directly to hypersensitivity. ABSTRACT The blood‐nerve barrier (BNB) is a selectively permeable barrier that creates an immunologically and biochemically privileged space for peripheral axons and supporting cells. The breakdown of the BNB allows access of blood‐borne (hematogenous) cells and molecules to the endoneurium to engage in the local inflammatory cascade. This process was examined in a mouse model of trauma‐associated neuropathic pain. The impact of nerve injury–triggered opening of the BNB in the development of chronic pain behavior was investigated. Partial ligation of the sciatic nerve led to a long‐lasting disruption of the BNB distal to the site of injury. Vascular endothelial growth factor (VEGF) was expressed by resident macrophages after nerve injury. Intraneural injection of VEGF decreased mechanical thresholds while opening the BNB. Serum from nerve‐injured or lipopolysaccharide‐treated animals elicited mechanical allodynia in naive animals, when allowed to bypass the BNB by intraneural injection. Intraneural injection of fibrinogen, a clotting protein in plasma that was found to deposit in the nerve after nerve injury, also produced a decrease in mechanical thresholds when introduced into naive nerves. These results demonstrate that blood‐borne molecules may play a role in the generation of neuropathic pain, suggesting that pain may be driven from infection or injury, at a distance from the nervous system. Furthermore, the breakdown of the BNB in neuropathic conditions was exploited to permit the entry of analgesic molecules that typically cannot pass the BNB, such as ProToxin‐II, a BNB‐impermeable Nav1.7 inhibitor. Therapeutics utilizing this mechanism could have selective access to injured nerves over healthy tissues.
20037471
Peripheral nerve pericytes modify the blood–nerve barrier function and tight junctional molecules through the secretion of various soluble factors
The objectives of this study were to establish pure blood–nerve barrier (BNB) and blood–brain barrier (BBB)‐derived pericyte cell lines of human origin and to investigate their unique properties as barrier‐forming cells. Brain and peripheral nerve pericyte cell lines were established via transfection with retrovirus vectors incorporating human temperature‐sensitive SV40 T antigen (tsA58) and telomerase. These cell lines expressed several pericyte markers such as α‐smooth muscle actin, NG2, platelet‐derived growth factor receptor β, whereas they did not express endothelial cell markers such as vWF and PECAM. In addition, the inulin clearance was significantly lowered in peripheral nerve microvascular endothelial cells (PnMECs) through the up‐regulation of claudin‐5 by soluble factors released from brain or peripheral nerve pericytes. In particular, bFGF secreted from peripheral nerve pericytes strengthened the barrier function of the BNB by increasing the expression of claudin‐5. Peripheral nerve pericytes may regulate the barrier function of the BNB, because the BNB does not contain cells equivalent to astrocytes which regulate the BBB function. Furthermore, these cell lines expressed several neurotrophic factors such as NGF, BDNF, and GDNF. The secretion of these growth factors from peripheral nerve pericytes might facilitate axonal regeneration in peripheral neuropathy. Investigation of the characteristics of peripheral nerve pericytes may provide novel strategies for modifying BNB functions and promoting peripheral nerve regeneration. J. Cell. Physiol. 226: 255–266, 2010. © 2010 Wiley‐Liss, Inc.
3776855
Changes in plasma cytokines associated with peripheral nerve injury
Plasma levels of interleukins 1, 2, 4 and 6 and tumor necrosis factor (TNF) were measured from 0 to 30 days in rats after a unilateral crush of the sciatic nerve at the level of the sciatic notch and after sham operations without nerve crush. Interleukin-6 was observed to peak and return to baseline levels within 24 h and remained at baseline for the duration of the experiment. An initial sharp rise in interleukin-1 and TNF was observed in all animals 1-2 days after the operation. A transient increase in interleukin-1 and TNF was also observed only in nerve-injured animals between 10 and 14 days after injury. A large increase in interleukin-2 appeared only in nerve-injured animals beginning at 11 days after injury and remained elevated for the remaining study period. No alterations in plasma interleukin-4 were observed at any time point. The experiments provide preliminary evidence for significant trauma-induced alterations in plasma cytokines which could provide a basis for some of the diffuse responses of peripheral neurons to trauma. The biphasic nature changes in plasma cytokines suggest that the immune system may participate in tissue reactions involved in the recovery from nerve injury.
22300095
Altered Blood‐Nerve Barrier Permeability to Small Molecules in Experimental Diabetes Mellitus
Permeability-surface area products (PA) were determined with a quantitative in vivo injection technique at the blood-nerve barrier of tibial nerve, and at the blood-brain barrier, in control and streptozotocin-induced diabetic rats. The PA product for [14C]mannitol at the blood-nerve barrier was increased by 100% in diabetic animals, 3.12 ± 0.15 ± 10-5 mls-1·g-1, compared with controls, 1.61 ± 0.10 ± 10-5 ml·s-1·g-1. In contrast, PA for [14C]mannitol at the blood-brain barrier was unaltered in the diabetic animals. Following intravenous injection, no leakage of microperoxidase across the perineurium or endoneurial vessels of diabetic rats could be demonstrated by morphological techniques. Nerve blood-space, as determined with intravenous [3H]inulin, and blood-nerve barrier surface area as determined by morphometric methods, did not differ in diabetic when compared to control animals. Thus, the calculated permeability coefficient for [14C]mannitol at the blood-nerve barrier was about 100% greater in diabetic nerve compared to control nerves. The increased permeability was accompanied by a 7% increase in nerve-water content and a 32% decrease in motor-nerve conduction velocity. The results demonstrate a specific vulnerability of nerve as compared to brain in an animal model of diabetes mellitus. Chronically altered permeability to small water-soluble molecules reduces the protective effect of salt impermeability at the blood-nerve barrier against nerve edema, and may be an important pathogenic mechanism in diabetic neuropathy.
7345903
Increase in albumin, IgG, and IgM blood-nerve barrier indices in human diabetic neuropathy.
The albumin (Alb), IgG, and IgM concentrations in the endoneurium of fascicular sural nerve biopsy samples were evaluated in controls (n = 9 or 10), diabetic patients without neuropathy (n = 6), and diabetic patients with polyneuropathy (n = 17 or 18). These values were significantly increased in diabetic patients with and without neuropathy when expressed both per endoneurial dry weight or endoneurial total protein compared to biopsy samples from healthy controls. When these concentrations, expressed per endoneurial total protein, were related to plasma concentrations similarly expressed, the resulting blood-nerve barrier (BNB) indices were significantly increased for Alb (6.1 times; P less than 0.00001), IgG (4.9 times; P = 0.00037), and IgM (2.7 times; P = 0.015). The diabetic patients without neuropathy (defined as having an index of pathology of greater than 0.65; a measure of the severity of the pathological abnormality based on morphological criteria) also had significant increases in two of these BNB indices that were intermediate between the diabetic neuropathy patients and controls (Alb, 3.9 times controls; P = 0.00002: IgG, 4.6 times controls; P = 0.00016: IgM, 1.8 times controls; not significant). No correlations were observed between the endoneurial concentrations of these plasma proteins or the BNB indices and the index of pathology, suggesting that these increases in endoneurial plasma proteins precede the pathologic alterations. The increased values for the diabetics in the absence of pathological abnormalities may prove useful in predicting neuropathic complications. The ratio of the IgG-BNB index to the Alb-BNB index was decreased 19%, and the ratio of the IgM-BNB index to the Alb-BNB index was decreased 56% in diabetic neuropathy patients compared to controls. Although the IgG and IgM concentrations are increased in the diabetic endoneurium, the Alb increase is greater and a mechanism other than size indiscriminate extravasation of plasma proteins, therefore, is suggested. Morphometric assessment of the endoneurial compartments, which would be expected to contain these plasma proteins, suggests that they are not altered in diabetic neuropathy; hence, it is hypothesized that the observed increase in endoneurial concentration of these plasma proteins results from altered transport through the endothelial or perineurial barrier, which supports an underlying vascular mechanism in the development of diabetic polyneuropathy.
9925749
Homeostatic regulation of the endoneurial microenvironment during development, aging and in response to trauma, disease and toxic insult
The endoneurial microenvironment, delimited by the endothelium of endoneurial vessels and a multi-layered ensheathing perineurium, is a specialized milieu intérieur within which axons, associated Schwann cells and other resident cells of peripheral nerves function. The endothelium and perineurium restricts as well as regulates exchange of material between the endoneurial microenvironment and the surrounding extracellular space and thus is more appropriately described as a blood–nerve interface (BNI) rather than a blood–nerve barrier (BNB). Input to and output from the endoneurial microenvironment occurs via blood–nerve exchange and convective endoneurial fluid flow driven by a proximo-distal hydrostatic pressure gradient. The independent regulation of the endothelial and perineurial components of the BNI during development, aging and in response to trauma is consistent with homeostatic regulation of the endoneurial microenvironment. Pathophysiological alterations of the endoneurium in experimental allergic neuritis (EAN), and diabetic and lead neuropathy are considered to be perturbations of endoneurial homeostasis. The interactions of Schwann cells, axons, macrophages, and mast cells via cell–cell and cell–matrix signaling regulate the permeability of this interface. A greater knowledge of the dynamic nature of tight junctions and the factors that induce and/or modulate these key elements of the BNI will increase our understanding of peripheral nerve disorders as well as stimulate the development of therapeutic strategies to treat these disorders.
34428286
FUNCTIONAL ASPECTS AND PATHOGENETIC CONSIDERATIONS OF THE NEUROPATHY IN THE SPONTANEOUSLY DIABETIC BB‐WISTAR RAT
Sima A.A.F. & Hay K. (1981) Neuropathology and Applied Neurobiology 7, 341–350
17250618
Glycated albumin is a better glycemic indicator than glycated hemoglobin values in hemodialysis patients with diabetes: effect of anemia and erythropoietin injection.
The significance of glycated albumin (GA), compared with casual plasma glucose (PG) and glycated hemoglobin (HbA(1c)), was evaluated as an indicator of the glycemic control state in hemodialysis (HD) patients with diabetes. The mean PG, GA, and HbA(1c) levels were 164.5 +/- 55.7 mg/dl, 22.5 +/- 7.5%, and 5.85 +/- 1.26%, respectively, in HD patients with diabetes (n = 538), which were increased by 51.5, 31.6, and 17.7%, respectively, compared with HD patients without diabetes (n = 828). HbA(1c) levels were significantly lower than simultaneous PG and GA values in those patients in comparison with the relationship among the three parameters in patients who had diabetes without renal dysfunction (n = 365), as reflected by the significantly more shallow slope of regression line between HbA(1c) and PG or GA. A significant negative correlation was found between GA and serum albumin (r = -0.131, P = 0.002) in HD patients with diabetes, whereas HbA(1c) correlated positively and negatively with hemoglobin (r = 0.090, P = 0.036) and weekly dose of erythropoietin injection (r = -0.159, P < 0.001), respectively. Although PG and GA did not differ significantly between HD patients with diabetes and with and without erythropoietin injection, HbA(1c) levels were significantly higher in patients without erythropoietin. Categorization of glycemic control into arbitrary quartile by HbA(1c) level led to better glycemic control in a significantly higher proportions of HD patients with diabetes than those assessed by GA. Multiple regression analysis demonstrated that the weekly dose of erythropoietin, in addition to PG, emerged as an independent factor associated with HbA(1c) in HD patients with diabetes, although PG but not albumin was an independent factor associated with GA. In summary, it is suggested that GA provides a significantly better measure to estimate glycemic control in HD patients with diabetes and that the assessment of glycemic control by HbA(1c) in these patients might lead to underestimation likely as a result of the increasing proportion of young erythrocyte by the use of erythropoietin.
21079533
Increased permeability across the blood-nerve barrier of albumin glycated in vitro and in vivo from patients with diabetic polyneuropathy.
The blood-nerve transfer of human plasma albumin glycated with D-glucose was investigated by measuring the permeability coefficient-surface area product (PS) of the blood-nerve barrier to radioiodinated albumin in normal adult rat sciatic nerve. Human albumin (ALB) from normal individuals, freshly isolated by CM-Affi-Gel Blue affinity chromatography, was glycated in vitro for 1, 3, 10, 19, and 30 weeks. Glycated ALB (gALB) was separated from the nonglycated form by boronate-affinity chromatography. The efficiency of this separation was assessed by chromatography of ALB glycated with [14C]glucose and by rechromatography of isolated ALB and gALB after radioiodination. The gALB was also shown to have a higher molecular weight and be completely separated from ALB after SDS/pore gradient electrophoresis in a Tris borate/EDTA buffer. After 1 week of glycation, the gALB PS was 2.2-fold greater than the ALB PS (0.724 +/- 0.063 x 10(-6) vs. 0.328 +/- 0.053 x 10(-6) ml.g-1.s-1; mean +/- SD; P less than 0.0001) and it increased with the time of glycation reaching a maximum value of 16.2-fold greater at 30 weeks (4.656 +/- 1.117 x 10(-6) vs. 0.288 +/- 0.042 x 10(-6) ml.g-1.s-1; mean +/- SD; P less than 0.0001). No change was observed in the residual endoneurial plasma volume. In addition, the PS of gALB isolated from patients with diabetic polyneuropathy was significantly increased (P less than 0.0001) compared to the PS for ALB isolated from the same patients. It is hypothesized that the increased permeability of gALB and presumably other glycated serum components across the blood-nerve barrier, as well as the observed quantitative increase in ALB, IgG, and IgM in sural nerve biopsies from patients with diabetic polyneuropathy contribute to the development of diabetic polyneuropathy over a prolonged period of time by mechanisms that might involve osmotic changes in the nerve microenvironment, direct toxic effects of glycated macromolecules on cells within the endoneurium, or nerve damage by classical immunological mechanisms due to trapping of glycated immunoglobulins within nerve.
207067563
The neuroinflammatory role of Schwann cells in disease
Peripheral neuropathies are associated with a variety of clinical symptoms ranging from motor and sensory symptoms to autonomic dysfunction. The primary disease causes for peripheral nerve disorders are also very heterogeneous, including genetic causes, inflammation mediated damage and physical trauma. A common theme in these neuropathies is the important contribution of the immune system; leading either to a deterioration or an amelioration of the disease. Immune responses are typically mediated by immune cells such as antigen-presenting cells, macrophages or T-cells. However, also non-immune cells such as microglia in the central nervous system or Schwann cells in the peripheral nervous system might play a key role in innate and adaptive immune responses. Just like microglia, Schwann cells express a plethora of pattern recognition receptors that allows them to recognize exogenous as well as endogenous danger signals. Upon activation, Schwann cells initiate and regulate local immune responses by presenting antigens and by secreting cytokines and chemokines, which will further attract immune cells to the site of injury. By interacting with immune cells they contribute in shaping immune responses that can lead to inflammatory neuropathies. In hereditary neuropathies, the immune system has also been shown to aggravate the disease phenotype. Besides, a neuroprotective role for the immune system has been recognized that becomes mainly prominent in cases of acute nerve injury. The present review focuses on the recently recognized immune competent role of Schwann cells and its involvement in peripheral neuropathies.
22439449
Non-neural-specific T lymphocytes can orchestrate inflammatory peripheral neuropathy.
Neural-specific T lymphocytes are held to play a pathological role in inflammatory peripheral nerve disorders such as the Guillain-Barré syndrome (GBS) and chronic inflammatory demyelinating polyneuropathy (CIDP). Here, non-neural-specific T-cell-mediated inflammation was studied in peripheral nerves in Lewis rats by systemic transfer of ovalbumin-specific activated T cells followed by intraneural injection of ovalbumin. Rapid endoneurial perivenular infiltration of alpha beta T cells and ED1+ macrophages occurred with ovalbumin injection following transfer of 2 x 10(6) T cells. This cellular infiltration and accumulation produced marked increases in blood-nerve barrier (BNB) permeability. In contrast, control casein injections produced neither significant cell accumulation nor BNB permeability changes. Transfer of a higher number of T cells (5 x 10(6)) induced severe Wallerian degeneration and nerve conduction failure in ovalbumin injected nerves. Fewer T cells (5 x 10(5)) induced conduction block and mild demyelination which were markedly augmented by systemic cotransfer of anti-myelin immunoglobulin. This study demonstrates that activated T cells of non-neural specificity can accumulate in peripheral nerve, produce dramatic changes in BNB permeability and with intravenous anti-myelin antibody orchestrate primary demyelination or axonal degeneration in a dose-dependent fashion.
30512871
Early alterations of Hedgehog signaling pathway in vascular endothelial cells after peripheral nerve injury elicit blood-nerve barrier disruption, nerve inflammation, and neuropathic pain development
Abstract Changes in the nerve's microenvironment and local inflammation resulting from peripheral nerve injury participate in nerve sensitization and neuropathic pain development. Taking part in these early changes, disruption of the blood–nerve barrier (BNB) allows for infiltration of immunocytes and promotes the neuroinflammation. However, molecular mechanisms engaged in vascular endothelial cells (VEC) dysfunction and BNB alterations remain unclear. In vivo, BNB permeability was assessed following chronic constriction injury (CCI) of the rat sciatic nerve (ScN) and differential expression of markers of VEC functional state, inflammation, and intracellular signaling was followed from 3 hours to 2 months postinjury. Several mechanisms potentially involved in functional alterations of VEC were evaluated in vitro using human VEC (hCMEC/D3), then confronted to in vivo physiopathological conditions. CCI of the ScN led to a rapid disruption of endoneurial vascular barrier that was correlated to a decreased production of endothelial tight-junction proteins and an early and sustained alteration of Hedgehog (Hh) signaling pathway. In vitro, activation of Toll-like receptor 4 in VEC downregulated the components of Hh pathway and altered the endothelial functional state. Inhibition of Hh signaling in the ScN of naive rats mimicked the biochemical and functional alterations observed after CCI and was, on its own, sufficient to evoke local neuroinflammation and sustained mechanical allodynia. Alteration of the Hh signaling pathway in VEC associated with peripheral nerve injury, is involved in BNB disruption and local inflammation, and could thus participate in the early changes leading to the peripheral nerve sensitization and, ultimately, neuropathic pain development.
5247252
Inflammation in the pathogenesis of microvascular complications in diabetes
Diabetes and hyperglycemia create a proinflammatory microenvironment that progresses to microvascular complications such as nephropathy, retinopathy, and neuropathy. Diet-induced insulin resistance is a potential initiator of this change in type 2 diabetes which can increase adipokines and generate a chronic low-grade inflammatory state. Advanced glycation end-products and its receptor, glycation end-products AGE receptor axis, reactive oxygen species, and hypoxia can also interact to worsen complications. Numerous efforts have gained way to understanding the mechanisms of these modulators and attenuation of the inflammatory response, however, effective treatments have still not emerged. The complexity of inflammatory signaling may suggest a need for multi-targeted therapy. This review presents recent findings aimed at new treatment strategies.
207068628
BTBR ob/ob mice as a novel diabetic neuropathy model: Neurological characterization and gene expression analyses
Given the lack of treatments for diabetic neuropathy (DN), a common diabetic complication, accurate disease models are necessary. Characterization of the leptin-deficient BTBR ob/ob mouse, a type 2 diabetes model, demonstrated that the mice develop robust diabetes coincident with severe neuropathic features, including nerve conduction deficits and intraepidermal nerve fiber loss by 9 and 13 weeks of age, respectively, supporting its use as a DN model. To gain insight into DN mechanisms, we performed microarray analysis on sciatic nerve from BTBR ob/ob mice, identifying 1503 and 642 differentially expressed genes associated with diabetes at 5 and 13 weeks, respectively. Further analyses identified overrepresentation of inflammation and immune-related functions in BTBR ob/ob mice, which interestingly were more highly represented at 5 weeks, an observation that may suggest a contributory role in DN onset. To complement the gene expression analysis, we demonstrated that protein levels of select cytokines were significantly upregulated at 13 weeks in BTBR ob/ob mouse sciatic nerve. Furthermore, we compared our array data to that from an established DN model, the C57BKS db/db mouse, which reflected a common dysregulation of inflammatory and immune-related pathways. Together, our data demonstrate that BTBR ob/ob mice develop rapid and robust DN associated with dysregulated inflammation and immune-related processes.
33229024
Toll-like receptor 4 and tumor necrosis factor-alpha as diagnostic biomarkers for diabetic peripheral neuropathy
Diabetic peripheral neuropathy (DPN) is one of the most common complications of diabetes, but currently no protein biomarkers have been introduced into clinical diagnosis, especially among early-stage diabetic patients. Our previous study in animal model showed Toll-like receptor (TLR) 4 and its downstream signaling molecules were associated with DPN. To assess the diagnostic values of TLR4, TNF-α, and IL-6 as biomarkers, here we detected their expressions in peripheral blood from normal controls, type 2 diabetic and DPN subjects. Both TLR4 mRNA and protein expressions increased significantly in DPN compare with both diabetic and control subjects. The protein levels of TNF-α and IL-6 were also raised significantly and correlated with TLR4 expression. Receiver operating characteristics (ROC) analysis suggested TLR4 and TNF-α had great potential advantages to predict the progression of neuropathy, the risks of DPN were increased in subjects with higher TLR4 (odds ratio: 5.27; 95% CI: 1.02-26.40) and TNF-α (odds ratio: 12.67; 95% CI: 2.35-68.22). These findings demonstrated TLR4 and TNF-α could be potential sensitive diagnostic biomarkers for DPN in both general population and type 2 diabetic subjects.
20588294
Monocytes/Macrophages control resolution of transient inflammatory pain.
UNLABELLED
Insights into mechanisms governing resolution of inflammatory pain are of great importance for many chronic pain-associated diseases. Here we investigate the role of macrophages/monocytes and the anti-inflammatory cytokine interleukin-10 (IL-10) in the resolution of transient inflammatory pain. Depletion of mice from peripheral monocytes/macrophages delayed resolution of intraplantar IL-1β- and carrageenan-induced inflammatory hyperalgesia from 1 to 3 days to >1 week. Intrathecal administration of a neutralizing IL-10 antibody also markedly delayed resolution of IL-1β- and carrageenan-induced inflammatory hyperalgesia. Recently, we showed that IL-1β- and carrageenan-induced hyperalgesia is significantly prolonged in LysM-GRK2(+/-) mice, which have reduced levels of G-protein-coupled receptor kinase 2 (GRK2) in LysM(+) myeloid cells. Here we show that adoptive transfer of wild-type, but not of GRK2(+/-), bone marrow-derived monocytes normalizes the resolution of IL-1β-induced hyperalgesia in LysM-GRK2(+/-) mice. Adoptive transfer of IL-10(-/-) bone marrow-derived monocytes failed to normalize the duration of IL-1β-induced hyperalgesia in LysM-GRK2(+/-) mice. Mechanistically, we show that GRK2(+/-) macrophages produce less IL-10 in vitro. In addition, intrathecal IL-10 administration attenuated IL-1β-induced hyperalgesia in LysM-GRK2(+/-) mice, whereas it had no effect in wild-type mice. Our data uncover a key role for monocytes/macrophages in promoting resolution of inflammatory hyperalgesia via a mechanism dependent on IL-10 signaling in dorsal root ganglia.


PERSPECTIVE
We show that IL-10-producing monocytes/macrophages promote resolution of transient inflammatory hyperalgesia. Additionally, we show that reduced monocyte/macrophage GRK2 impairs resolution of hyperalgesia and reduces IL-10 production. We propose that low GRK2 expression and/or impaired IL-10 production by monocytes/macrophages represent peripheral biomarkers for the risk of developing chronic pain after inflammation.
23233577
Diabetes mellitus as a prothrombotic condition
Diabetes mellitus (DM) is characterized by fasting hyperglycaemia and a high risk of atherothrombotic disorders affecting the coronary, cerebral and peripheral arterial trees. The risk of myocardial infarction (MI) is 3‐5 fold higher in Type 2 DM and a DM subject with no history of MI has the same risk as a non‐DM subject with a past history of MI. In total around 70% of deaths are vascular with poorer outcomes to both acute events and cardiological interventions. It was proposed that clustering of vascular risk factors (hyperinsulinaemia, dysglycaemia, dyslipidaemia and hypertension) around insulin resistance (IR) accounted for the increase in risk with Type 2 DM. The importance of this became apparent with the recognition that risk clustering occurs in normoglycaemic and impaired glucose tolerance (IGT) subjects with IR, in total around 25% of the population in addition to long‐standing Type 1 subjects with renal disease. Evidence indicates that thrombotic risk clustering also occurs in association with IR, suppression of fibrinolysis due to elevated concentrations of the fibrinolytic inhibitor, plasminogen activator inhibitor‐1 (PAI‐1) is invariable with IR and there is evidence that this is regulated by the effects of triglyceride on the PAI‐1 gene promoter. Other studies indicated that prothrombotic risk (coagulation factors VII, XII and fibrinogen) also associates with the IR syndrome. The development of endothelial cell dysfunction with suppression of nitric oxide and prostacyclin synthesis, combined with platelet resistance to the anti‐aggregatory effects of these hormones leads to loss of control over platelet activation. In addition, hyperglycaemia and glycation have marked effects on fibrin structure function, generating a clot which has a denser structure, resistant to fibrinolysis. The combination of increased circulating coagulation zymogens, inhibition of fibrinolysis, changes in fibrin structure/function and alterations in platelet reactivity creates a thrombotic risk clustering which underpins the development of cardiovascular disease.
20758297
Microangiopathy in human diabetic neuropathy: relationship between capillary abnormalities and the severity of neuropathy
SummaryClinical, electrophysiological and ultrastractural morphometric observations were made in 5 diabetic non-neuropathic patients, 5 diabetic patients with mild neuropathy and 11 diabetic patients with severe neuropathy. Capillary abnormalities were assessed in simultaneous nerve, muscle and skin biopsies and compared with results from 6 age-matched, non-diabetic control subjects.Nerve capillaries demonstrated markedly greater pathology than skin and muscle capillaries. Endoneurial capillary density was significantly reduced in severely neuropathic diabetic patients (p<0.01) when compared with control subjects. Capillary basement membrane (p<0.002), endothelial cell (p<0.003) and total diffusion barrier (endothelial cell, pericyte, basement membrane) (p<0.001) thickness were significantly increased, and oxygen diffusing capacity was significantly reduced (p<0.001) in the nerves of patients with severe diabetic neuropathy when compared to control subjects. Endothelial cell profile number and luminal perimeter were significantly increased in asymptomatic (p<0.01), (p<0.05) and severely neuropathic (p<0.001), (p<0.05) diabetic patients respectively. However, endothelial cell outer perimeter, a measure of capillary size, showed no significant increase in diabetic patients when compared with control subjects. An association was observed between neurophysiological and neuropathological measures of neuropathic severity. There was no significant correlation between the duration of diabetes and HbA1 levels with capillary pathology or with neuropathic severity. Very few abnormalities of muscle and skin correlated with neuropathic severity. However, all measures of nerve capillary pathology correlated significantly with neurophysiological and neuropathological measures of neuropathic severity.
7030600
Sural nerve pathology in diabetic patients with minimal but progressive neuropathy
Aims/hypothesisThe early pathological features of human diabetic neuropathy are not clearly defined. Therefore we quantified nerve fibre and microvascular pathology in sural nerve biopsies from diabetic patients with minimal neuropathy.MethodsTwelve diabetic patients underwent detailed assessment of neuropathy and fascicular sural nerve biopsy at baseline, with repeat assessment of neuropathy 8.7±0.6 years later.ResultsAt baseline, neuropathic symptoms, neurological deficits, quantitative sensory testing, cardiac autonomic function and peripheral nerve electrophysiology showed minimal abnormality, which deteriorated at follow-up. Myelinated fibre density, fibre and axonal area, and g-ratio were normal but teased fibre studies showed paranodal abnormalities (p<0.001), segmental demyelination (p<0.01) and remyelination (p<0.01) without axonal degeneration. Unassociated Schwann cell profile density (p<0.04) and unmyelinated axon density (p<0.001) were increased and axon diameter was decreased (p<0.007). Endoneurial capillaries demonstrated basement membrane thickening (p<0.006), endothelial cell hyperplasia (p<0.004) and a reduction in luminal area (p<0.007).Conclusions/interpretationThe early pathological features of human diabetic neuropathy include an abnormality of the myelinated fibre Schwann cell and unmyelinated fibre degeneration with regeneration. These changes are accompanied by a significant endoneurial microangiopathy.
9751917
Endoneurial capillary abnormalities presage deterioration of glucose tolerance and accompany peripheral neuropathy in man.
To explore whether microangiopathy is associated with disturbed glucose tolerance and peripheral neuropathy, we assessed endoneurial capillary morphology in sural nerve biopsies from men with diabetes, impaired glucose tolerance (IGT), and normal glucose tolerance (NGT). Baseline morphology was related to glucose tolerance and neuropathy at baseline and at follow-up 6 years later. Capillary density (in number per millimeters squared) at baseline was higher in subjects with diabetes (n = 10) compared with those with NGT (n = 5) at follow-up (median [interquartile range]) (86.0 [24.3] vs. 54.9 [17.1]; P = 0.0200) and in those progressing from IGT to diabetes (n = 4) compared with those with persistent IGT (n = 4) (86.7 [25.2] vs. 54.1 [14.6]; P = 0.0433). The capillary luminal area (in micrometers squared) was lower in subjects with NGT progressing to IGT (n = 2) or subjects with IGT progressing to diabetes (n = 3) compared with subjects with constant NGT (n = 6) or constant IGT (n = 4) (11.9 [2.4] vs. 20.8 [7.8]; P = 0.0201). The capillary basement membrane area (in micrometers squared) was increased in patients with peripheral neuropathy (n = 10) compared with those without (n = 7) (114.6 [68.8] vs. 75.3 [28.7]; P = 0.0084). In conclusion, increased capillary density was associated with current or future diabetes, decreased capillary luminal area with future deterioration in glucose tolerance, and increased basement membrane area with peripheral neuropathy.
9294785
Sural nerve oxygen tension in diabetes.
Peripheral nerve oxygen tensions were assessed in vivo by using microelectrodes to measure endoneurial oxygen tension in exposed sural nerve. In 11 diabetic patients with chronic sensorimotor neuropathy the mean endoneurial oxygen tension was 39.7 (SD 10.2) mm Hg. In all but one patient compared with none of four non-neuropathic subjects the mean nerve oxygen tensions were below dorsal foot vein values. This unphysiological state may have a role in the aetiology of diabetic neuropathy.
24052865
Aldose reductase inhibition, nerve perfusion, oxygenation and function in streptozotocin-diabetic rats: Dose-response considerations and independence from a myo-inositol mechanism
SummaryWe examined the effects of aldose reductase inhibition on nerve biochemistry and function, blood flow and endoneurial oxygenation in experimental diabetes mellitus. After 1 month untreated diabetes in rats, treatment with the novel sulphonylnitromethane aldose reductase inhibitor, ZENECA ZD5522, prevented a progressive increase in sciatic nerve resistance to hypoxic conduction failure (p<0.05). Motor conduction velocity deficits after 4 months untreated diabetes were rapidly returned to normal within 12 days (p<0.0001) by ZD5522 treatment. Following 2-months untreated diabetes, examination of 1 month ZD5522 treatment dose-response relationships for correction of nerve sorbitol and fructose accumulations and reduction in myo-inositol concentration, sciatic motor and saphenous sensory conduction velocity and sciatic blood flow by laser-Doppler flowmetry revealed poor agreement between nerve function and biochemical indices. In addition, polyol accumulation differed between sciatic and saphenous nerves, the latter showing ten-fold lower sorbitol concentrations. Laser-Doppler blood flow was 60% decreased by untreated diabetes (p<0.001) and there was a strong correlation between ZD5522-mediated increases in blood flow and conduction velocity (p<0.0001). Measurement of nutritive endoneurial blood flow by microelectrode polarography and hydrogen clearance showed 44% and 45% deficits for 1 and 2 months untreated diabetes (p<0.001) that were prevented by ponalrestat and ZD5522 treatments, respectively. In contrast, 2 months myo-inositol treatment from diabetes induction did not prevent reduction in blood flow or sciatic motor conduction velocity. A 37% reduction in endoneurial oxygen tension after 2 months diabetes (p<0.001) was completely prevented by ZD5522 treatment (p< 0.001). The data show that a very high degree of polyol pathway blockade is necessary to correct nerve functional deficits and that aldose reductase inhibitors have a neurovascular action that does not depend on restoration of nerve myo-inositol.
24205848
A new minimally invasive technique to show nerve ischaemia in diabetic neuropathy
Aims/hypothesis. Experimental studies have shown that abnormalities of nerve microcirculation are important factors in the pathogenesis of diabetic neuropathy but there have been few clinical studies. We have applied microlightguide spectrophotometry to measure intravascular oxygen saturation (HbO2%) and blood flow in human sural nerve. Methods. We studied ten patients with mild-moderate sensory motor diabetic neuropathy, nine patients without neuropathy and nine control subjects. We took 300 measurements of oxygen saturation under direct visual control through a 1.9 mm rigid endoscope over three regions of the nerve. Spectrophotometric measurements of nerve fluorescence were taken after an intravenous injection of sodium fluorescein and the rate of increase in nerve fluorescence (rise time) was used as an indicator of nerve blood flow. Results. Nerve oxygen saturation was reduced in patients with neuropathy compared with control subjects (67.1 ± 2.2 % vs 76.7 ± 2.1 %, p = 0.006). Fluorescein rise time was prolonged in patients with neuropathy compared with the control group (48.5 ± 7.0 s vs 14.0 ± 3.1 s, p = 0.001) suggesting impaired nerve blood flow. There was a correlation between rise time, nerve oxygen saturation, glycaemic control and sural nerve sensory conduction velocity (p < 0.01). Conclusion/interpretation. The combination of microlight-guide spectrophotometry and micro-endoscopy provides a valuable minimally invasive technique for clinical investigation of nerve microcirculation. We have shown reduced nerve oxygenation and impaired blood flow in diabetic neuropathy and these findings strongly support a central role of microvascular disease in the pathogenesis of diabetic neuropathy. [Diabetologia (1999) 42: 737–742]
9996814
Peripheral Nerve Injury Induces Persistent Vascular Dysfunction and Endoneurial Hypoxia, Contributing to the Genesis of Neuropathic Pain
Nerve injury is associated with microvascular disturbance; however, the role of the vascular system has not been well characterized in the context of neuropathic pain. Furthermore, ischemia is thought to play a role in a number of neuropathic pain conditions, and yet the role of hypoxia has also not been characterized in neuropathic pain conditions. In this study, we observed the presence of persistent endoneurial hypoxia in a mouse model of traumatic peripheral nerve injury, causing painful mononeuropathy. We attribute the ongoing hypoxia to microvascular dysfunction, endoneurial fibrosis, and increased metabolic requirements within the injured nerve. Increased lactate levels were observed in injured nerves, as well as increased oxygen consumption and extracellular acidification rates, suggesting that anaerobic glycolysis is required to maintain cellular ATP levels. Hypoxia causes a reduction in levels of the Na+/K+ ATPase ion transporter in both cultured primary dorsal root ganglion neurons and injured peripheral nerve. A reduction of Na+/K+ ATPase ion transporter levels likely contributes to the hyperexcitability of injured nerves. Physiological antagonism of hypoxia with hyperbaric oxygen alleviated mechanical allodynia in nerve-injured animals. These results suggest that hypoxia and the Na+/K+ ATPase ion transporter may be a novel mechanistic target for the treatment of neuropathic pain. In addition, the findings support the possibility of using hypoxia activated pro-drugs to localize treatments for neuropathic pain and nerve injury to injured nerves.
23824707
Production of vascular endothelial growth factor by murine macrophages: regulation by hypoxia, lactate, and the inducible nitric oxide synthase pathway.
Murine thioglycolate-induced peritoneal macrophages (MPMs) and the murine RAW264.7 macrophage-like cell line (RAW cells) constitutively produce vascular endothelial growth factor (VEGF). VEGF production is increased under hypoxic conditions or after cell activation with interferon-gamma (IFNgamma) and endotoxin (lipopolysaccharide, LPS). In contrast, tumor necrosis factor-alpha is produced only by IFNgamma/LPS-activated cells. Lactate (25 mmol/L) does not increase VEGF production by these cells. However, hypoxia, lactate, and IFNgamma/LPS-activated MPMs express angiogenic activity, whereas normoxic, nonactivated MPMs do not. Lack of angiogenic activity is not due to an antiangiogenic factor(s) in the medium of these cells. Angiogenic activity produced by hypoxia and lactate-treated MPMs is neutralized by anti-VEGF antibody, which also neutralizes most of the angiogenic activity produced by IFNgamma/LPS-activated MPMs. The inducible nitric oxide synthase inhibitors Ng-nitro-L-arginine-methyl ester (1.5 mmol/L) and aminoguanidine (1 mmol/L) block production of angiogenic activity by MPMs and RAW cells. In RAW cells, Ng-nitro-L-arginine-methyl ester and AG block IFNgamma/LPS-activated, but not constitutive, VEGF production, whereas in MPMs, neither constitutive nor IFNgamma/LPS-activated VEGF synthesis is affected. Synthesis of tumor necrosis factor-alpha is also unaffected. In contrast to normoxic, nonactivated MPMs, inducible nitric oxide synthase-inhibited, IFNgamma/LPS-activated MPMs produce an antiangiogenic factor(s). We conclude that VEGF is a major contributor to macrophage-derived angiogenic activity, and that activation by hypoxia, lactate, or IFNgamma/LPS switches macrophage-derived VEGF from a nonangiogenic to an angiogenic state. This switch may involve a posttranslational modification of VEGF, possibly by the process of ADP-ribosylation. ADP-ribosylation by MPM cytosolic extracts or by cholera toxin switches rVEGF165 from an angiogenic to a nonangiogenic state. In IFNgamma/LPS-activated MPMs, the inducible nitric oxide synthase-dependent pathway also regulates the expression of an antiangiogenic factor(s) that antagonizes the bioactivity of VEGF and provides an additional regulatory pathway controlling the angiogenic phenotype of macrophages.
34395300
Hypoxia and hypoxia signaling in tissue repair and fibrosis.
Following injury, vascular damage results in the loss of perfusion and consequent low oxygen tension (hypoxia) which may be exacerbated by a rapid influx of inflammatory and mesenchymal cells with high metabolic demands for oxygen. Changes in systemic and cellular oxygen concentrations induce tightly regulated response pathways that attempt to restore oxygen supply to cells and modulate cell function in hypoxic conditions. Most of these responses occur through the induction of the transcription factor hypoxia-inducible factor-1 (HIF-1) which regulates many processes needed for tissue repair during ischemia in the damaged tissue. HIF-1 transcriptionally upregulates expression of metabolic proteins (GLUT-1), adhesion proteins (integrins), soluble growth factors (TGF-β and VEGF), and extracellular matrix components (type I collagen and fibronectin), which enhance the repair process. For these reasons, HIF-1 is viewed as a positive regulator of wound healing and a potential regulator of organ repair and tissue fibrosis. Understanding the complex role of hypoxia in the loss of function in scarring tissues and biology of chronic wound, and organ repair will aid in the development of pharmaceutical agents that can redress the detrimental outcomes often seen in repair and scarring.
3583349
Role of Hypoxia Inducible Factor 1 in Hyperglycemia-Exacerbated Blood-Brain Barrier Disruption in Ischemic Stroke
Diabetes is a major stroke risk factor and is associated with poor functional recovery after stroke. Accumulating evidence indicates that the worsened outcomes may be due to hyperglycemia-induced cerebral vascular complications, especially disruption of the blood-brain barrier (BBB). The present study tested a hypothesis that the activation of hypoxia inducible factor-1 (HIF-1) was involved in hyperglycemia-aggravated BBB disruption in an ischemic stroke model. Non-diabetic control and Streptozotocin-induced type I diabetic mice were subjected to 90min transient middle cerebral artery occlusion (MCAO) followed by reperfusion. Our results demonstrated that hyperglycemia induced higher expression of HIF-1α and vascular endothelial growth factor (VEGF) in brain microvessels after MCAO/reperfusion. Diabetic mice showed exacerbated BBB damage and tight junction disruption, increased infarct volume as well as worsened neurological deficits. Furthermore, suppressing HIF-1 activity by specific knock-out endothelial HIF-1α ameliorated BBB leakage and brain infarction in diabetic animals. Moreover, glycemic control by insulin abolished HIF-1α up-regulation in diabetic animals and reduced BBB permeability and brain infarction. These findings strongly indicate that HIF-1 plays an important role in hyperglycemia-induced exacerbation of BBB disruption in ischemic stroke. Endothelial HIF-1 inhibition warrants further investigation as a therapeutic target for the treatment of stroke patients with diabetes.
25607286
PTEN/mTOR and axon regeneration
How axon regeneration is controlled in both PNS and CNS remains elusive. Mechanistic studies of axon growth during development and axon regeneration after injury reveal the PTEN dependent molecular mechanism as a commonality. This pathway could impact the processes occurring in the neuronal soma, such as mTOR-regulated protein translation, and in the axons, such as cytoskeleton assembly. In this review, we will discuss the current understanding of the involvement of these processes in the regulation of axon growth and the potential implication in promoting axon regeneration after injury.
5032379
Regeneration of diabetic axons is enhanced by selective knockdown of the PTEN gene.
Diabetes mellitus renders both widespread and localized irreversible damage to peripheral axons while imposing critical limitations on their ability to regenerate. A major failure of regenerative capacity thereby imposes a 'double hit' in diabetic patients who frequently develop focal neuropathies such as carpal tunnel syndrome in addition to generalized diffuse polyneuropathy. The mechanisms of diabetic neuron regenerative failure have been speculative and few approaches have offered therapeutic opportunities. In this work we identify an unexpected but major role for PTEN upregulation in diabetic peripheral neurons in attenuating axon regrowth. In chronic diabetic neuropathy models in mice, we identified significant PTEN upregulation in peripheral sensory neurons of messenger RNA and protein compared to littermate controls. In vitro, sensory neurons from these mice responded to PTEN knockdown with substantial rises in neurite outgrowth and branching. To test regenerative plasticity in a chronic diabetic model with established neuropathy, we superimposed an additional focal sciatic nerve crush injury and assessed morphological, electrophysiological and behavioural recovery. Knockdown of PTEN in dorsal root ganglia ipsilateral to the side of injury was achieved using a unique form of non-viral short interfering RNA delivery to the ipsilateral nerve injury site and paw. In comparison with scrambled sequence control short interfering RNA, PTEN short interfering RNA improved several facets of regeneration: recovery of compound muscle action potentials, reflecting numbers of reconnected motor axons to endplates, conduction velocities of both motor and sensory axons, reflecting their maturation during regrowth, numbers and calibre of regenerating myelinated axons distal to the injury site, reinnervation of the skin by unmyelinated epidermal axons and recovery of mechanical sensation. Collectively, these findings identify a novel therapeutic approach, potentially applicable to other neurological conditions requiring specific forms of molecular knockdown, and also identify a unique target, PTEN, to treat diabetic neuroregenerative failure.
38293062
Hypoxia‐inducible factor 1 mediates increased expression of NADPH oxidase‐2 in response to intermittent hypoxia
Sleep‐disordered breathing with recurrent apnea is associated with intermittent hypoxia (IH). Cardiovascular morbidities caused by IH are triggered by increased generation of reactive oxygen species (ROS) by pro‐oxidant enzymes, especially NADPH oxidase‐2 (Nox2). Previous studies showed that (i) IH activates hypoxia‐inducible factor 1 (HIF‐1) in a ROS‐dependent manner and (ii) HIF‐1 is required for IH‐induced ROS generation, indicating the existence of a feed‐forward mechanism. In the present study, using multiple pharmacological and genetic approaches, we investigated whether IH‐induced expression of Nox2 is mediated by HIF‐1 in the central and peripheral nervous system of mice as well as in cultured cells. IH increased Nox2 mRNA, protein, and enzyme activity in PC12 pheochromocytoma cells as well as in wild‐type mouse embryonic fibroblasts (MEFs). This effect was abolished or attenuated by blocking HIF‐1 activity through RNA interference or pharmacologic inhibition (digoxin or YC‐1) or by genetic knockout of HIF‐1α in MEFs. Increasing HIF‐1α expression by treating PC 12 cells with the iron chelator deferoxamine for 20 h or by transfecting them with HIF‐1alpha expression vector increased Nox2 expression and enzyme activity. Exposure of wild‐type mice to IH (8 h/day for 10 days) up‐regulated Nox2 mRNA expression in brain cortex, brain stem, and carotid body but not in cerebellum. IH did not induce Nox2 expression in cortex, brainstem, carotid body, or cerebellum of Hif1a+/− mice, which do not manifest increased ROS or cardiovascular morbidities in response to IH. These results establish a pathogenic mechanism linking HIF‐1, ROS generation, and cardiovascular pathology in response to IH. J. Cell. Physiol. 226: 2925–2933, 2011. © 2011 Wiley‐Liss, Inc.
32667928
Involvement of ROS in BBB dysfunction
The blood–brain barrier (BBB) forms a protective barrier around the brain, with the important function of maintaining brain homeostasis. Pathways thought to initiate BBB dysfunction include the kinin system, excitotoxicity, neutrophil recruitment, mitochondrial alterations and macrophage/microglial activation, all of which converge on the same point—reactive oxygen species (ROS). Interestingly, ROS also provide a common trigger for many downstream pathways that directly mediate BBB compromise such as oxidative damage, tight junction (TJ) modification and matrix metalloproteinases (MMP) activation. These observations suggest that ROS are key mediators of BBB breakdown and implicate antioxidants as potential neuroprotectants in conditions like stroke and traumatic brain injury (TBI). This review explores some of the pathways both upstream and downstream of ROS that have been implicated in increased BBB permeability and discusses the role of ROS and antioxidants in neuropathology.
3698993
Effects of antioxidants on nerve and vascular dysfunction in experimental diabetes.
Reactive oxygen species (ROS) are elevated by metabolic changes in diabetes, including autoxidation and increased advanced glycation. Endogenous protection by the glutathione redox cycle is also compromised by the competing NADPH requirement of elevated polyol pathway flux. Antioxidant treatment strategies prevent or reverse nerve conduction velocity (NCV) deficits in diabetic rats. These include lipophilic scavengers such as butylated hydroxytoluene, probucol and vitamin E, more hydrophilic agents like alpha-lipoic acid and acetyl cysteine, and transition metal chelators that inhibit autoxidation. In the long-term, elevated ROS cause cumulative damage to neurons and Schwann cells, however, they also have a deleterious effect on nerve blood flow in the short term. This causes endoneurial hypoxia, which is responsible for early NCV deficits. Antioxidant treatment corrects the blood flow deficit and promotes normal endoneurial oxygenation. ROS cause antioxidant-preventable vascular endothelium abnormalities, neutralizing nitric oxide mediated vasodilation and increasing reactivity to vasoconstrictors. Unsaturated fatty acids are a major target for ROS and essential fatty acid metabolism is impaired by diabetes. Gamma-linolenic acid stimulates vasodilator prostanoid production, and there are marked synergistic interactions between gamma-linolenic acid and antioxidants. This has encouraged the development of novel drugs such as ascorbyl-gamma-linolenic acid and gamma-linolenic acid-lipoic acid with enhanced therapeutic potential.
21056964
Endothelial/Pericyte Interactions
Interactions between endothelial cells and mural cells (pericytes and vascular smooth muscle cells) in the blood vessel wall have recently come into focus as central processes in the regulation of vascular formation, stabilization, remodeling, and function. Failure of the interactions between the 2 cell types, as seen in numerous genetic mouse models, results in severe and often lethal cardiovascular defects. Abnormal interactions between the 2 cell types are also implicated in a number of human pathological conditions, including tumor angiogenesis, diabetic microangiopathy, ectopic tissue calcification, and stroke and dementia syndrome CADASIL. In the present review, we summarize current knowledge concerning the identity, characteristics, diversity, ontogeny, and plasticity of pericytes. We focus on the advancement in recent years of the understanding of intercellular communication between endothelial and mural cells with a focus on transforming growth factor α, angiopoietins, platelet-derived growth factor, spingosine-1-phosphate, and Notch ligands and their respective receptors. We finally highlight recent important data contributing to the understanding of the role of pericytes in tumor angiogenesis, diabetic retinopathy, and hereditary lymphedema.
46999655
The Role of Advanced Glycation End Products in Diabetic Vascular Complications
In cases of chronic hyperglycemia, advanced glycation end-products (AGEs) are actively produced and accumulated in the circulating blood and various tissues. AGEs also accelerate the expression of receptors for AGEs, and they play an important role in the development of diabetic vascular complications through various mechanisms. Active interventions for glucose and related risk factors may help improve the clinical course of patients by reducing AGEs. This review summarizes recent updates on AGEs that have a significant impact on diabetic vascular complications.
9022300
Isolation and properties of endothelial cells forming the blood-nerve barrier.
The blood-nerve barrier (BNB) is one of the functional barriers sheltering the nervous system from circulating blood. It is very important to understand the cellular properties of endothelial cells of endoneurial origin because these cells constitute the bulk of the BNB. This chapter describes a standard protocol for isolating the endothelial cells forming the BNB. In addition, methods for confirming some of the barrier properties of isolated endothelial cells are also described.
25743781
Establishment and characterization of human peripheral nerve microvascular endothelial cell lines: a new in vitro blood-nerve barrier (BNB) model.
The blood-nerve barrier (BNB) is a highly specialized unit that maintains the microenvironments of the peripheral nervous system. Since the breakdown of the BNB has been considered a key step in autoimmune neuropathies such as Guillain-Barré syndrome and chronic inflammatory demyelinating polyraduculoneuropathy, it is important to understand the cellular properties of the peripheral nerve microvascular endothelial cells (PnMECs) which constitute the BNB. For this purpose, we established an immortalized cell line derived from human PnMECs. The human PnMECs were transduced with retroviral vectors encoding the temperature-sensitive SV40 large T antigen and human telomerase. This cell line, termed FH-BNB, showed a spindle fiber-shaped morphology, expression of von Willebrand factor and uptake of acetylated low density lipoprotein. These cells expressed tight junction proteins including occludin, claudin-5, ZO-1 and ZO-2 at the cell-cell boundaries. P-glycoprotein and GLUT-1 were also detected by a Western blot analysis and the cells exhibited the functional expression of p-glycoprotein. In addition, transendothelial electrical resistance experiments and paracellular permeabilities of sodium fluorescein and fluorescein isothiocyanate-labeled dextran of molecular weight 4 kDa across these cells demonstrated that FH-BNBs had functional tight junctions. These results indicated that FH-BNBs had highly specialized barrier properties and they might therefore be a useful tool to analyze the pathophysiology of various neuropathies.
9262376
Perineurial defect and peripheral opioid analgesia in inflammation
Opioid receptors have been demonstrated on sensory nerves in both inflamed and normal subcutaneous tissue but locally applied opioid agonists produce analgesia in inflamed tissue only. Inflammation confers a disruption of the perineurial barrier that can also be induced deliberately by hyperosmolar solutions. The present study examines at which stage of Freund's adjuvant-induced inflammation peripheral opioid analgesic effects become manifest and whether a perineurial defect contributes to the appearance of such effects. To this end we have monitored the temporal evolution of inflammatory signs (swelling, temperature, hyperalgesia) and of peripheral antinociceptive effects (by the paw pressure test) of mu-, delta-, and kappa-selective opioids. Using horseradish peroxidase histochemistry, the perineurial barrier was assessed in normal and inflamed tissue and following its artificial disruption by hyperosmolar saline and mannitol in vivo. Finally, we sought to elicit analgesia in normal tissue by the concomitant application of mannitol and receptor-selective opioids or by an extremely lipophilic opioid agonist (fentanyl). We found that peripheral opioid antinociception and perineurial leakage occur simultaneously at a very early stage (within 12 hr) of the inflammatory reaction and that both can be mimicked by the administration of hyperosmolar solutions in normal tissue. Fentanyl produced peripheral antinociception in noninflamed tissue that was potentiated by mannitol or inflammation. Our findings demonstrate that the perineurium is a crucial determinant for peripheral opioid analgesia and that the efficacy of locally applied hydrophilic or lipophilic neuromodulatory compounds can be improved dramatically by the concomitant modulation of perineurial permeability.(ABSTRACT TRUNCATED AT 250 WORDS)
52006
Transient opening of the perineurial barrier for analgesic drug delivery
Selective targeting of sensory or nociceptive neurons in peripheral nerves remains a clinically desirable goal. Delivery of promising analgesic drugs is often impeded by the perineurium, which functions as a diffusion barrier attributable to tight junctions. We used perineurial injection of hypertonic saline as a tool to open the perineurial barrier transiently in rats and elucidated the molecular action principle in mechanistic detail: Hypertonic saline acts via metalloproteinase 9 (MMP9). The noncatalytic hemopexin domain of MMP9 binds to the low-density lipoprotein receptor-related protein-1, triggers phosphorylation of extracellular signal-regulated kinase 1/2, and induces down-regulation of the barrier-forming tight junction protein claudin-1. Perisciatic injection of any component of this pathway, including MMP9 hemopexin domain or claudin-1 siRNA, enables an opioid peptide ([D-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin) and a selective sodium channel (NaV1.7)-blocking toxin (ProToxin-II) to exert antinociceptive effects without motor impairment. The latter, as well as the classic TTX, blocked compound action potentials in isolated nerves only after disruption of the perineurial barrier, which, in return, allowed endoneurally released calcitonin gene-related peptide to pass through the nerve sheaths. Our data establish the function and regulation of claudin-1 in the perineurium as the major sealing component, which could be modulated to facilitate drug delivery or, potentially, reseal the barrier under pathological conditions.