Neurovascular development and links to disease

The developing central nervous system (CNS) is vascularized via ingression of blood vessels from the outside as the neural tissue expands. This angiogenic process occurs without perturbing CNS architecture due to exquisite cross-talk between the neural compartment and invading blood vessels. Subsequently, this intimate relationship also promotes the formation of the neurovascular unit that underlies the blood–brain barrier and regulates blood flow to match brain activity. This review provides a historical perspective on research into CNS blood vessel growth and patterning, discusses current models used to study CNS angiogenesis, and provides an overview of the cellular and molecular mechanisms that promote blood vessel growth and maturation. Finally, we highlight the significance of these mechanisms for two different types of neurovascular CNS disease.     

Monocyte chemoattractant protein-1 and the blood–brain barrier

The blood–brain barrier (BBB) is a dynamic structure that maintains the homeostasis of the brain and thus proper neurological functions. BBB compromise has been found in many pathological conditions, including neuroinflammation. Monocyte chemoattractant protein-1 (MCP1), a chemokine that is transiently and significantly up-regulated during inflammation, is able to disrupt the integrity of BBB and modulate the progression of various diseases, including excitotoxic injury and hemorrhage. In this review, we first introduce the biochemistry and biology of MCP1, and then summarize the effects of MCP1 on BBB integrity as well as individual BBB components.     

Cyclooxygenase,lipoxygenase and tumor angiogenesis

Arachidonic acid metabolism through cyclooxygenase (COX) and lipoxygenase (LOX) pathways generates various biologically active lipids that play important roles in inflammation, thrombosis and tumor progression. Angiogenesis, the formation of new capillary vessels from preexisting ones, underpins a number of physiological processes and participates in the development of several pathological conditions such as arthritis, cancer and various eye diseases. The formation of new capillary vessels is a multistep process that involves endothelial cell proliferation, migration and tube formation. In the present review, we survey the literature on the regulation of angiogenesis by arachidonate metabolites, especially those from the COX and 12-LOX pathways in the context of tumor growth, and put forward some unanswered but important questions for future studies.     

Transient receptor potential canonical channels in angiogenesis and axon guidance

Wiring of vascular and neural networks requires precise guidance of growing blood vessels and axons, respectively, to reach their targets during development. Both of the processes share common molecular signaling pathways. Transient receptor potential canonical (TRPC) channels are calcium-permeable cation channels and gated via receptor- or store-operated mechanisms. Recent studies have revealed the requirement of TRPC channels in mediating guidance cue-induced calcium influx and their essential roles in regulating axon navigation and angiogenesis. Dissecting TRPC functions in these physiological processes may provide therapeutic implications for suppressing pathological angiogenesis and improving nerve regeneration.     

The regulatory function of SPARC in vascular biology

SPARC is a matricellular protein, able to modulate cell/ECM interactions and influence cell responses to growth factors, and therefore is particularly attuned to contribute to physiological processes involving changes in ECM and cell mobilization. Indeed, the list of biological processes affected by SPARC includes wound healing, tumor progression, bone formation, fibrosis, and angiogenesis. The process of angiogenesis is complex and involves a number of cellular processes such as endothelial cell proliferation, migration, ECM degradation, and synthesis, as well as pericyte recruitment to stabilize nascent vessels. In this review, we will summarize current results that explore the function of SPARC in the regulation of angiogenic events with a particular emphasis on the modulation of growth factor activity by SPARC in the context of blood vessel formation. The primary function of SPARC in angiogenesis remains unclear, as SPARC activity in some circumstances promotes angiogenesis and in others is more consistent with an anti-angiogenic activity. Undoubtedly, the mercurial nature of SPARC belies a redundancy of functional proteins in angiogenesis as well as cell-type-specific activities that alter signal transduction events in response to unique cellular milieus. Nonetheless, the investigation of cellular mechanisms that define functional activities of SPARC continue to contribute novel and exciting paradigms to vascular biology.     

Vascular integrins: pleiotropic adhesion and signaling molecules in vascular homeostasis and angiogenesis

New blood vessel formation, a process referred to as angiogenesis, is essential for embryonic development and for many physiological and pathological processes during postnatal life, including cancer progression. Endothelial cell adhesion molecules of the integrin family have emerged as critical mediators and regulators of angiogenesis and vascular homeostasis. Integrins provide the physical interaction with the extracellular matrix necessary for cell adhesion, migration and positioning, and induction of signaling events essential for cell survival, proliferation and differentiation. Antagonists of integrin alpha V beta 3 suppress angiogenesis in many experimental models and are currently tested in clinical trials for their therapeutic efficacy against angiogenesis-dependent diseases, including cancer. Furthermore, interfering with signaling pathways downstream of integrins results in suppression of angiogenesis and may have relevant therapeutic implications. In this article we review the role of integrins in endothelial cell function and angiogenesis. In the light of recent advances in the field, we will discuss their relevance as a therapeutic target to suppress tumor angiogenesis.     

Transfection of primary brain capillary endothelial cells for protein synthesis and secretion of recombinant erythropoietin: a strategy to enable protein delivery to the brain

Treatment of chronic disorders affecting the central nervous system (CNS) is complicated by the inability of drugs to cross the blood–brain barrier (BBB). Non-viral gene therapy applied to brain capillary endothelial cells (BCECs) denotes a novel approach to overcome the restraints in this passage, as turning BCECs into recombinant protein factories by transfection could result in protein secretion further into the brain. The present study aims to investigate the possibility of transfecting primary rat brain endothelial cells (RBECs) for recombinant protein synthesis and secretion of the neuroprotective protein erythropoietin (EPO). We previously showed that 4% of RBECs with BBB properties can be transfected without disrupting the BBB integrity in vitro, but it can be questioned whether this is sufficient to enable protein secretion at therapeutic levels. The present study examined various transfection vectors, with regard to increasing the transfection efficiency without disrupting the BBB integrity. Lipofectamine 3000? was the most potent vector compared to polyethylenimine (PEI) and Turbofect. When co-cultured with astrocytes, the genetically modified RBECs secreted recombinant EPO into the cell culture medium both luminally and abluminally, and despite lower levels of EPO reaching the abluminal chamber, the amount of recombinant EPO was sufficient to evolve a biological effect on astrocytes cultured at the abluminal side in terms of upregulated gene expression of brain-derived neurotropic factor (BDNF). In conclusion, non-viral gene therapy to RBECs leads to protein secretion and signifies a method for therapeutic proteins to target cells inside the CNS otherwise omitted due to the BBB.     

Cardiomyokines from the heart

The heart is regarded as an endocrine organ as well as a pump for circulation, since atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were discovered in cardiomyocytes to be secreted as hormones. Both ANP and BNP bind to their receptors expressed on remote organs, such as kidneys and blood vessels; therefore, the heart controls the circulation by pumping blood and by secreting endocrine peptides. Cardiomyocytes secrete other peptides besides natriuretic peptides. Although most of such cardiomyocyte-derived peptides act on the heart in autocrine/paracrine fashions, several peptides target remote organs. In this review, to overview current knowledge of endocrine properties of the heart, we focus on cardiomyocyte-derived peptides (cardiomyokines) that act on the remote organs as well as the heart. Cardiomyokines act on remote organs to regulate cardiovascular homeostasis, systemic metabolism, and inflammation. Therefore, through its endocrine function, the heart can maintain physiological conditions and prevent organ damage under pathological conditions.     

Nitric oxide-dependent blood-brain barrier permeability alteration in the rat brain

The role of nitric oxide (NO), a well known vasodilator, in the regulation of blood-brain barrier (BBB) permeability is not clear. Therefore, the present study was planned to assess the role of NO-releasing compounds like sodium nitroprusside (SNP) and the active metabolite of molsidomine, SIN-1, as well as a precursor of NO, L-arginine, on this physiological barrier. The permeability was assessed by using several tracers. All three agents increased the permeability of BBB to the tracer. The increase in permeability caused by L-arginine was not blocked by N-nitro-L-arginine methyl ester (L-NAME). L-Arginine-treated brains did not show an elevation of nitrite content, thus ruling out the possibility of NO generation and its involvement in BBB permeability alteration. It is concluded that NO itself causes an increase in the permeability of BBB. However arginine-induced opening is not NO mediated.CDRI Communication Number: 5178.     

Secretogranin III: a diabetic retinopathy-selective angiogenic factor

Secretogranin III (Scg3) is a member of the granin protein family that regulates the biogenesis of secretory granules. Scg3 was recently discovered as an angiogenic factor, expanding its functional role to extrinsic regulation. Unlike many other known angiogenic factors, the pro-angiogenic actions of Scg3 are restricted to pathological conditions. Among thousands of quantified endothelial ligands, Scg3 has the highest binding activity ratio to diabetic vs. healthy mouse retinas and lowest background binding to normal vessels. In contrast, vascular endothelial growth factor binds to and stimulates angiogenesis of both diabetic and control vasculature. Consistent with its role in pathological angiogenesis, Scg3-neutralizing antibodies alleviate retinal vascular leakage in mouse models of diabetic retinopathy and retinal neovascularization in oxygen-induced retinopathy mice. This review summarizes our current knowledge of Scg3 as a regulatory protein of secretory granules, highlights its new role as a highly disease-selective angiogenic factor, and envisions Scg3 inhibitors as “selective angiogenesis blockers” for targeted therapy.     

Iron transport across the blood–brain barrier: development,neurovascular regulation and cerebral amyloid angiopathy

There are two barriers for iron entry into the brain: (1) the brain–cerebrospinal fluid (CSF) barrier and (2) the blood–brain barrier (BBB). Here, we review the literature on developmental iron accumulation by the brain, focusing on the transport of iron through the brain microvascular endothelial cells (BMVEC) of the BBB. We review the iron trafficking proteins which may be involved in the iron flux across BMVEC and discuss the plausible mechanisms of BMVEC iron uptake and efflux. We suggest a model for how BMVEC iron uptake and efflux are regulated and a mechanism by which the majority of iron is trafficked across the developing BBB under the direct guidance of neighboring astrocytes. Thus, we place brain iron uptake in the context of the neurovascular unit of the adult brain. Last, we propose that BMVEC iron is involved in the aggregation of amyloid-β peptides leading to the progression of cerebral amyloid angiopathy which often occurs prior to dementia and the onset of Alzheimer’s disease.     

Cytokine-mediated proteolysis in tissue remodelling

Summary Proteolytic enzymes play a key role in a variety of physiological processes in which the degradation of macromolecules is essential: angiogenesis, embryogenesis, bone and tissue remodelling, blood hemostasis and cell migration. The action of these enzymes is also crucial in the development of many pathological conditions such as wound healing, neoplasia, inflammation and arthritic disorders.the activity of proteases is negatively affected by specific protease-inhibitors. Various growth factors and other cytokines modulate the synthesis and secretion of both proteases and protease-inhibitors. The study of this regulation results in a better insight into (patho)physiology at the molecular level and promises to result in alternative treatment strategies.     

Cytokine-mediated proteolysis in tissue remodelling

Proteolytic enzymes play a key role in a variety of physiological processes in which the degradation of macromolecules is essential: angiogenesis, embryogenesis, bone and tissue remodelling, blood hemostasis and cell migration. The action of these enzymes is also crucial in the development of many pathological conditions such as wound healing, neoplasia, inflammation and arthritic disorders. The activity of proteases is negatively affected by specific protease-inhibitors. Various growth factors and other cytokines modulate the synthesis and secretion of both proteases and protease-inhibitors. The study of this regulation results in a better insight into (patho)physiology at the molecular level and promises to result in alternative treatment strategies.     

Pre- and post-natal ontogeny of neutral endopeptidase 24-11 ('enkephalinase') studied by in vitro autoradiography in the rat.

Neutral endopeptidase (NEP, enkephalinase, CALLA) which is present in various neural and non-neural tissues, is able to cleave a variety of regulatory peptides. The distribution of NEP has been studied during rat pre- and post-natal development by autoradiography after in vitro binding of the tritiated inhibitor [3H]HACBO-Gly to whole-body and organ sections. In the central nervous system (CNS), where the presence of NEP has been related to the termination of the action of enkephalins, the external layer of the olfactory bulbs is the only structure prominently labeled before birth. Other CNS structures rich in NEP in the adult, such as the nigrostriatal tract, are progressively labeled after birth. Outside the CNS, the progressive appearance of NEP in the kidney, the lungs and the salivary glands suggests its concomitant involvement in adult physiological functions, including fluid balance control, possibly by cleaving the atrial natriuretic peptide (ANP) and other peptides. On the other hand, transient or enhanced expression of NEP is observed during the development of several organs such as the sensory organs, the heart and the major blood vessels, the intestine, the bones and the genital tubercle. In addition to the still incompletely known physiological functions of the enzyme, the developmental pattern of its expression in several tissues strongly suggests a modulatory role for NEP in the ontogeny of a large number of organs.     

Janus head: the dual role of HLA-G in CNS immunity

The central nervous system (CNS) is considered an immune-privileged organ that maintains an adaptable immune surveillance system. Dysregulated immune function within the CNS contributes to the development of brain tumor growth, and robust immune activation results in excessive inflammation. Human lymphocyte antigen-G (HLA-G) proteins with tolerogenic immunoreactivity have been implicated in various pathophysiological processes including immune surveillance, governing homeostasis and immune regulation. In this review, we describe the wealth of evidence for the involvement of HLA-G in the CNS under physiological and pathological conditions. Further, we review regulatory functions that may be applicable as beneficial strategies in the therapeutic manipulation of immune-mediated CNS immune responses. Additionally, we try to understand how this molecule cooperates with other CNS-resident cells to maintain normal immune homeostasis, while still facilitating the development of the appropriate immune responses.     

Pre-and post-natal ontogeny of neutral endopeptidase 24-11 (‘enkephalinase’) studied by in vitro autoradiography in the rat

Neutral endopeptidase (NEP, enkephalinase, CALLA) which is present in various neural and non-neural tissues, is able to cleave a variety of regulatory peptides. The distribution of NEP has been studied during rat pre-and post-natal development by autoradiography after in vitro binding of the tritiated inhibitor [³H]HACBO-Gly to whole-body and organ sections. In the central nervous system (CNS), where the presence of NEP has been related to the termination of the action of enkephalins, the external layer of the olfactory bulbs is the only structure prominently labeled before birth. Other CNS structures rich in NEP in the adult, such as the nigrostriatal tract, are progressively labeled after birth. Outside the CNS, the progressive appearance of NEP in the kidney, the lungs and the salivary glands suggests its concomitant involvement in adult physiological functions, including fluid balance control, possibly by cleaving the atrial natriuretic peptide (ANP) and other peptides. On the other hand, transient or enhanced expression of NEP is observed during the development of several organs such as the sensory organs, the heart and the major blood vessels, the intestine, the bones and the genital tubercle. In addition to the still incompletely known physiological functions of the enzyme, the developmental pattern of its expression in several tissues strongly suggests a modulatory role for NEP in the ontogeny of a large number of organs.     

Transgenic mouse models of multiple sclerosis

Multiple sclerosis (MS) is an inflammatory demyelinating disease affecting the central nervous system (CNS) and a frequent cause of neurological disability in young adults. Multifocal inflammatory lesions in the CNS white matter, demyelination, oligodendrocyte loss, axonal damage, as well as astrogliosis represent the histological hallmarks of the disease. These pathological features of MS can be mimicked, at least in part, using animal models. This review discusses the current concepts of the immune effector mechanisms driving CNS demyelination in murine models. It highlights the fundamental contribution of transgenesis in identifying the mediators and mechanisms involved in the pathophysiology of MS models.     

Mesenchymal stem cells secretome: a new paradigm for central nervous system regeneration?

The low regeneration potential of the central nervous system (CNS) represents a challenge for the development of new therapeutic strategies. Mesenchymal stem cells (MSCs) have been proposed as a possible therapeutic tool for CNS disorders. In addition to their differentiation potential, it is well accepted nowadays that their beneficial actions can also be mediated by their secretome. Indeed, it was already demonstrated, both in vitro and in vivo, that MSCs are able to secrete a broad range of neuroregulatory factors that promote an increase in neurogenesis, inhibition of apoptosis and glial scar formation, immunomodulation, angiogenesis, neuronal and glial cell survival, as well as relevant neuroprotective actions on different pathophysiological contexts. Considering their protective action in lesioned sites, MSCs’ secretome might also improve the integration of local progenitor cells in neuroregeneration processes, opening a door for their future use as therapeutical strategies in human clinical trials. Thus, in this review we analyze the current understanding of MSCs secretome as a new paradigm for the treatment of CNS neurodegenerative diseases.     

Endothelial cell apoptosis in angiogenesis and vessel regression

Blood vessel regression is an essential process for ensuring blood vessel networks function at optimal efficiency and for matching blood supply to the metabolic needs of tissues as they change over time. Angiogenesis is the major mechanism by which new blood vessels are produced, but the vessel growth associated with angiogenesis must be complemented by remodeling and maturation events including the removal of redundant vessel segments and cells to fashion the newly forming vasculature into an efficient, hierarchical network. This review will summarize recent findings on the role that endothelial cell apoptosis plays in vascular remodeling during angiogenesis and in vessel regression more generally.