Activation of a procollagenase by low-molecular-weight angiogenesis factor

Avascular tumours have the ability to establish a blood supply for themselves by secreting a humoral factor which stimulates their host's endothelial cells to proliferate and to migrate towards the tumour source. The mechanism of action of such a humoral anglo-genesis factor is more than that of an endothelial-cell growth factor since it requires an oriented migration of cells towards the tumour. We report here the activation of pure skin-fibroblast procollagenase by a low-molecular-weight angiogenesis factor capable of stimulating endothelial-cell growth in vitro. The activation was observed when either Type I or III collagen was used as substrate. It is suggested that at least one function of angiogenesis factor is to promote limited degradation of the connective tissue through which it passes causing channeling in the matrix along which stimulated endothelial cells may     

In vivo models of angiogenesis

The process of building new blood vessels (angiogenesis) and controlling the propagation of blood vessels (anti-angiogenesis) are fundamental to human health, as they play key roles in wound healing and tissue growth. More than 500 million people may stand to benefit from anti- or pro-angiogenic treatments in the coming decades [National Cancer Institute (USA), Cancer Bulletin, volume 3, no. 9, 2006]. The use of animal models to assay angiogenesis is crucial to the search for therapeutic agents that inhibit angiogenesis in the clinical setting. Examples of persons that would benefit from these therapies are cancer patients, as cancer growth and spread is angiogenesis-dependent, and patients with aberrant angiogenesis in the eye, which may lead to blindness or defective sight. Recently, anti-angiogenesis therapies have been introduced successfully in the clinic, representing a turning point in tumor therapy and the treatment of macular degeneration and heralding a new era for the treatment of several commonly occurring angiogenesis-related diseases. On the other hand, pro-angiogenic therapies that promote compensatory angiogenesis in hypoxic tissues, such as those subjected to ischemia in myocardial or cerebral hypoxia due to occluding lesions in the coronary or cerebral arteries, respectively, and in cases of poor wound healing, are also being developed. In this review, the current major and newly introduced preclinical angiogenesis assays are described and discussed in terms of their specific advantages and disadvantages from the biological, technical, economical and ethical perspectives. These assays include the corneal micropocket, chick chorioallantoic membrane, rodent mesentery, subcutaneous (s.c.) sponge/matrix/alginate microbead, s.c. Matrigel plug, s.c. disc, and s.c. directed in vivo angiogenesis assays, as well as, the zebrafish system and several additional assays. A note on quantitative techniques for assessing angiogenesis in patients is also included. The currently utilized preclinical assays are not equivalent in terms of efficacy or relevance to human disease. Some of these assays have significance for screening, while others are used primarily in studies of dosage-effects, molecular structure activities, and the combined effects of two or more agents on angiogenesis. When invited to write this review, I was asked to describe in some detail the rodent mesenteric-window angiogenesis assay, which has not received extensive coverage in previous reviews.     

Regulation of angiogenesis: wound healing as a model

Normal tissue function requires adequate supply of oxygen through blood vessels. Understanding how blood vessels form is a challenging objective because angiogenesis is vital to many physiological and pathological processes. Unraveling mechanisms of angiogenesis would offer therapeutic options to ameliorate disorders that are currently leading causes of mortality and morbidity, including cardiovascular diseases, cancer, chronic inflammatory disorders, diabetic retinopathy, excessive tissue defects, and chronic non-healing wounds. Restoring blood flow to the site of injured tissue is a prerequisite for mounting a successful repair response, and wound angiogenesis represents a paradigmatic model to study molecular mechanisms involved in the formation and remodeling of vascular structures. In particular, repair of skin defects offers an ideal model to analyze angiogenesis due to its easy accessibility to control and manipulate this process. Most of those growth factors, extracellular matrix molecules, and cell types, recently discovered and considered as crucial factors in blood vessel formation, have been identified and analyzed during skin repair and the process of wound angiogenesis. This article will review cellular and molecular mechanisms controlling angiogenesis in cutaneous tissue repair in light of recent reports and data from our laboratories. In this article we will discuss the contribution of growth factors, basement membrane molecules, and mural cells in wound angiogenesis. The article provides a rationale for targeting the angiogenic response in order to modulate the outcome of the healing response.     

An intracrine view of angiogenesis

Angiogenesis, the generation of new blood vessels from pre-existing vessels, is an integral component of wound healing, responses to inflammation and other physiologic processes. It is also an essential part of tumor growth; in the absence of new vessel formation, tumors cannot expand beyond a small volume. Although much is known about angiogenesis and its regulation, there is no overall theory that describes or explains this process. It is here suggested that the intracrine hypothesis, which ascribes to certain extracellular signaling peptides (whether hormones, growth factors, DNA-binding proteins or enzymes) a role in both intracellular biology and extracellular signaling, can contribute to a more general understanding of angiogenesis. Intracrine factors participate in angiogenesis in the following ways: (1) they can act within the cells that synthesized them (type I intracrine action), (2) they can be secreted and then taken up by their cell of synthesis to act intracellularly (type II intracrine action ), or (3) they can be secreted and internalized by a distant target cell (type III intracrine action). The parallels between the intracrine growth factor mechanisms cancer cells employ in stimulating their own growth and the mechanisms operative in endothelial cell proliferation during angiogenesis ("intracrine reciprocity") are discussed. Collectively, these explorations lead to testable hypotheses regarding the regulation of normal and pathological angiogenesis, and point to similarities between tumor-induced angiogenesis and tissue differentiation.     

Pathways in the emergence of developmental neuroethology: antecedents to current views of neurobehavioral ontogeny.

The historical forces that have contributed to our current views of neurobehavioral development (and thus to the fields of developmental psychobiology and neuroethology) are many and varied. Although similar statements might be made about almost any field of science, it is in particular true of this field, which represents a kind of mongrel discipline derived from at least three major sources (psychology, embryology, and neuroscience) and several more minor ones (including developmental psychology and psychiatry, psychoanalysis, education, zoology, ethology, and sociology). Although I attempt to demonstrate here how each of these sources may have influenced the emergence of a unified field of developmental psychobiology or developmental neuroethology, because the present article represents the first attempt of which I am aware to trace the history of these fields I am certain that there is considerable room for improvement, correction, and revision of the views expressed here. Accordingly, I consider this inaugural effort a kind of reconnaissance intended to trace a necessarily imperfect historic path for others to follow and improve upon. In the final analysis, I will be satisfied if this article only serves to underscore two related points: first is the value derived from historical studies of contemporary issues in development, and the second concerns the extent to which our current ideas and concepts about neurobehavioral development, ideas often considered new and contemporary, were already well known to those who came before us. The first point underscores the arguments expressed in the Introduction that the present must always be reconciled with the past, for the past is never entirely past. The second point returns full circle to an important thought expressed in the opening quotation to this article, namely, that even though our historic predecessors lacked much of the empirical facts available to us they were nonetheless able to attain a surprisingly deep understanding of neurobehavioral ontogeny.     

Expression and purification of human recombinant GST-FGF receptor-1

When tumors undergo the angiogenic switch, cell growth and tissue invasion is facilitated by the formation of new capillaries from preexisting blood vessels, a process known as angiogenesis. Growth factors such as vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) and fibroblast growth factor (FGF) trigger the process of angiogenesis. Here we describe a protocol for the expression and one-step purification of human recombinant GST-FGF receptor type 1 (FGFR-1) from Sf9 cells. This protocol allows generating an active kinase as indicated by its reactivity with a monoclonal antibody to phosphorylated tyrosine. The purified enzyme displays a specific activity of 1.2 x 10(4) pmol mg(-1) min(-1), which is in the range of activities reported for homogeneously purified recombinant kinases. We have employed a number of compounds to show that the GST-FGFR-1 preparation is suitable to the identification of tyrosine kinase inhibitors. Considering that inhibitors of angiogenesis may represent an attractive tool in therapeutic strategies targeting invasive metastatic tumors the results presented here, along with available data on the structure of the ATP-binding pocket of FGFR-1, should facilitate the rational design of specific FGFR-1 inhibitory compounds.     

Embryonic and adult vasculogenesis

Two mechanisms account for the formation of blood vessels, vasculogenesis and angiogenesis. Unfortunately, the terms vasculogenesis and angiogenesis literally have the same meaning, i.e., the genesis of blood vessels, and thus do little to distinguish between the two processes. Despite the nomenclature, the two processes are clearly distinct. Vasculogenesis, the de novo formation of blood vessels from mesoderm, is driven by the recruitment of undifferentiated mesodermal cells to the endothelial lineage and the de novo assembly of such cells into blood vessels. Angiogenesis is the generation of new blood vessels from endothelial cells of existing blood vessels, a process driven by endothelial cell proliferation. Recent years have seen dramatic changes in our understanding of the process of vasculogenesis, expanding the scope of its occurrence beyond the earliest stages of development to include involvement in neovascular processes throughout development as well as in the adult. In this review, emphasis is placed on discussion of emerging perspectives on the process of vasculogenesis in both the embryo and the adult.     

A pharmacologically based multiscale mathematical model of angiogenesis and its use in investigating the efficacy of a new cancer treatment strategy

Tumor angiogenesis is the process by which new blood vessels are formed and enhance the oxygenation and growth of tumors. As angiogenesis is recognized as being a critical event in cancer development, considerable efforts have been made to identify inhibitors of this process. Cytostatic treatments that target the molecular events of the angiogenesis process have been developed, and have met with some success. However, it is usually difficult to preclinically assess the effectiveness of targeted therapies, and apparently promising compounds sometimes fail in clinical trials.We have developed a multiscale mathematical model of angiogenesis and tumor growth. At the molecular level, the model focuses on molecular competition between pro- and anti-angiogenic substances modeled on the basis of pharmacological laws. At the tissue scale, the model uses partial differential equations to describe the spatio-temporal changes in cancer cells during three stages of the cell cycle, as well as those of the endothelial cells that constitute the blood vessel walls.This model is used to qualitatively assess how efficient endostatin gene therapy is. Endostatin is an anti-angiogenic endogenous substance. The gene therapy entails overexpressing endostatin in the tumor and in the surrounding tissue. Simulations show that there is a critical treatment dose below which increasing the duration of treatment leads to a loss of efficacy.This theoretical model may be useful to evaluate the efficacy of therapies targeting angiogenesis, and could therefore contribute to designing prospective clinical trials.     

Glioma angiogenesis: Towards novel RNA therapeutics

Brain tumors exhibit marked and aberrant blood vessel formation indicating angiogenic endothelial cells as a potential target for brain tumor treatment. The brain tumor blood vessels are used for nutrient delivery, and possibly for cancer cell migration. The process of angiogenesis is complex and involves multiple players. The current angiogenesis inhibitors used in clinical trials mostly target single angiogenic proteins and so far show limited effects on tumor growth. Besides the conventional angiogenesis inhibitors, RNA-based inhibitors such as small-interfering RNAs (siRNAs) are being analyzed for their capacity to silence the message of proteins involved in neovascularization. More recently, a new family of non-coding RNAs, named angiomirs [microRNAs (miRNAs) involved in angiogenesis] has emerged. These small RNAs have the advantage over siRNAs in that they have the potential of silencing multiple messages at the same time and therefore they might become therapeutically relevant in a “one-hit multiple-target” context against brain tumor angiogenesis. In this review we will discuss the emerging technologies in anti-angiogenesis emphasizing on RNA-based therapeutics.Key words: glioma, angiogenesis, anti-angiogenesis therapy, siRNA, miRNA, endothelial cells, blood vessels     

Glioma angiogenesis

Brain tumors exhibit marked and aberrant blood vessel formation indicating angiogenic endothelial cells as a potential target for brain tumor treatment. The brain tumor blood vessels are used for nutrient delivery, and possibly for cancer cell migration. The process of angiogenesis is complex and involves multiple players. The current angiogenesis inhibitors used in clinical trials mostly target single angiogenic proteins and so far show limited effects on tumor growth. Besides the conventional angiogenesis inhibitors, RNA-based inhibitors such as small-interfering RNAs (siRNAs) are being analyzed for their capacity to silence the message of proteins involved in neovascularization. More recently, a new family of non-coding RNAs, named angiomirs [microRNAs (miRNAs) involved in angiogenesis] has emerged. These small RNAs have the advantage over siRNAs in that they have the potential of silencing multiple messages at the same time and therefore they might become therapeutically relevant in a “one-hit multiple-target” context against brain tumor angiogenesis. In this review we will discuss the emerging technologies in anti-angiogenesis emphasizing on RNA-based therapeutics.     

Nitric oxide signaling during myocardial angiogenesis

Ischemic heart disease develops as a consequence of coronary atherosclerotic lesion formation. Coronary collateral vessels and microvascular angiogenesis develop as an adaptive response to myocardial ischemia, which ameliorates the function of the damaged heart. Angiogenesis, the formation of new blood vessels from pre-existing vascular bed, is of paramount importance in the maintenance of vascular integrity both in the repair process of damaged tissue and in the formation of collateral vessels in response to tissue ischemia. Angiogenesis is modulated by a multitude of cytokines/chemokines and growth factors. In this regard, angiogenesis cannot be viewed as a single process. It is likely that different mediators are involved in different phases of angiogenesis. Vascular endothelial cells (ECs) produce nitric oxide (NO), an endothelium-derived labile molecule, which maintains vascular homeostasis and thereby prevents vascular atherosclerotic changes. In patients with ischemic heart disease, the release of endothelium-derived NO is decreased, which plays an important role in the atherosclerotic disease progression. In recent years, endothelium-derived NO has been shown to modulate angiogenesis in vitro and in vivo. In this review, we summarize recent progress in the field of the NO-mediated regulation of postnatal angiogenesis, particularly in response to myocardial ischemia.     

Angiogenesis in the female reproductive system.

In adult tissues, capillary growth (angiogenesis) occurs normally during tissue repair, such as in healing of wounds and fractures. Rampant capillary growth is associated with various pathological conditions, including tumor growth, retinopathies, hemangiomas, fibroses and rheumatoid arthritis. The female reproductive organs (i.e., ovary, uterus, and placenta) exhibit dynamic, periodic growth and regression accompanied by equally dramatic changes in rates of blood flow. It is not surprising, therefore, that they are some of the few adult tissues in which angiogenesis occurs as a normal process. Thus, the female reproductive system provides a unique model for studying regulation of angiogenesis during growth and differentiation of normal adult tissues. Ovarian, uterine, and placental tissues recently have been shown to contain and produce angiogenic and anti-angiogenic factors. This review discusses the current state of knowledge regarding angiogenic processes and their regulation in female reproductive tissues. In addition, implications of this research for regulation of fertility as well as for control of angiogenesis in other normal and pathological processes are discussed.     

alphavbeta3 integrin and angiogenesis: a moody integrin in a changing environment

In the adult, angiogenesis, the formation of new blood vessels from pre-existing vasculature contributes to the pathogenesis of many disorders including cancer. The role of adhesion molecules, especially integrins, in pathological angiogenesis has long been the subject of investigation, mostly because of their potential as anti-angiogenic targets. Recent studies have highlighted the complexities connected with understanding the roles of one particular integrin, alphavbeta3, in neovascularization. This integrin is notoriously promiscuous and its precise functions in angiogenesis are unclear. Here, I have firstly summarized some of the salient features of the roles played by alphavbeta3 during angiogenesis; secondly attempted to address the apparently conflicting issues surrounding this topic; and finally raised some questions that appear to be unanswered.     

Strategic Endothelial Cell Tube Formation Assay: Comparing Extracellular Matrix and Growth Factor Reduced Extracellular Matrix

Malignant tumors require a blood supply in order to survive and spread. These tumors obtain their needed blood from the patient''s blood stream by hijacking the process of angiogenesis, in which new blood vessels are formed from existing blood vessels. The CXCR2 (chemokine (C-X-C motif) receptor 2) receptor is a transmembrane G-protein-linked molecule found in many cells that is closely associated with angiogenesis¹. Specific blockade of the CXCR2 receptor inhibits angiogenesis, as measured by several assays such as the endothelial tube formation assay. The tube formation assay is useful for studying angiogenesis because it is an excellent method of studying the effects that any given compound or environmental condition may have on angiogenesis. It is a simple and quick in vitro assay that generates quantifiable data and requires relatively few components. Unlike in vivo assays, it does not require animals and can be carried out in less than two days. This protocol describes a variation of the extracellular matrix supporting endothelial tube formation assay, which tests the CXCR2 receptor.     

Pathways in the emergence of developmental neuroethology: Antecedents to current views of neurobehavioral ontogeny

The historical forces that have contributed to our current views of neurobehavioral development (and thus to the fields of developmental psychobiology and neuroethology) are many and varied. Although similar statements might be made about almost any field of science, it is in particular true of this field, which represents a kind of mongrel discipline derived from at least three major sources (psychology, embryology, and neuroscience) and several more minor ones (including developmental psychology and psychiatry, psychoanalysis, education, zoology, ethology, and sociology). Although I attempt to demonstrate here how each of these sources may have influenced the emergence of a unified field of developmental psychobiology or developmental neuroethology, because the present article represents the first attempt of which I am aware to trace the history of these fields I am certain that there is considerable room for improvement, correction, and revision of the views expressed here. Accordingly, I consider this inaugural effort a kind of reconnaissance intended to trace a necessarily imperfect historic path for others to follow and improve upon. In the final analysis, I will be satisfied if this article only serves to underscore two related points: first is the value derived from historical studies of contemporary issues in development, and the second concerns the extent to which our current ideas and concepts about neurobehavioral development, ideas often considered new and contemporary, were already well known to those who came before us. The first point underscores the arguments expressed in the Introduction that the present must always be reconciled with the past, for the past is never entirely past. The second point returns full circle to an important thought expressed in the opening quotation to this article, namely, that even though our historic predecessors lacked much of the empirical facts available to us they were nonetheless able to attain a surprisingly deep understanding of neurobehavioral ontogeny. © 1992 John Wiley & Sons, Inc.     

De-extinction and the conception of species

Developments in genetic engineering may soon allow biologists to clone organisms from extinct species. The process, dubbed “de-extinction,” has been publicized as a means to bring extinct species back to life. For theorists and philosophers of biology, the process also suggests a thought experiment for the ongoing “species problem”: given a species concept, would a clone be classified in the extinct species? Previous analyses have answered this question in the context of specific de-extinction technologies or particular species concepts. The thought experiment is given more comprehensive treatment here. Given the products of three de-extinction technologies, twenty-two species concepts are “tested” to see which are consistent with the idea that species may be resurrected. The ensuing discussion considers whether or not de-extinction is a conceptually coherent research program and, if so, whether or not its development may contribute to a resolution of the species problem. Ultimately, theorists must face a choice: they may revise their commitments to species concepts (if those concepts are inconsistent with de-extinction) or they may recognize de-extinction as a means to make progress in the species problem.     

Pivotal role for decorin in angiogenesis

Angiogenesis, the formation of new blood vessels from preexisting vessels, is a highly complex process. It is regulated in a finely-tuned manner by numerous molecules including not only soluble growth factors such as vascular endothelial growth factor and several other growth factors, but also a diverse set of insoluble molecules, particularly collagenous and non-collagenous matrix constituents. In this review we have focused on the role and potential mechanisms of a multifunctional small leucine-rich proteoglycan decorin in angiogenesis. Depending on the cellular and molecular microenvironment where angiogenesis occurs, decorin can exhibit either a proangiogenic or an antiangiogenic activity. Nevertheless, in tumorigenesis-associated angiogenesis and in various inflammatory processes, particularly foreign body reactions and scarring, decorin exhibits an antiangiogenic activity, thus providing a potential basis for the development of decorin-based therapies in these pathological situations.     

Model of angiogenesis in mice with severe combined immunodeficiency (SCID) and xenoengrafted with Epstein-Barr virus-transformed B cells

Xenoengraftment of human cells in mice with severe combined immunodeficiency (SCID) has been used as a model system to study the mechanisms of B-cell lymphomagenesis. In the study reported here, we determined that SCID mice can also be used as a model to study angiogenesis in B-cell lymphomas. The C.B-17 scid/scid mice were xenotransplanted with Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCL), and we determined whether CD31, a marker found on endothelial cells, was detected in the human B-cell lymphomas that developed in these mice. Microvessel formation was identified by use of immunohistochemical staining for CD31. To assess possible mechanisms of angiogenic stimulus, we analyzed the expression of interleukin 8 (IL-8), a chemokine documented to promote angiogenesis, in non-small-cell lung cancer and bronchogenic carcinomas. We observed that a panel of LCL and LCL-lymphomas expressed IL-8 mRNA and protein. Neutralization of IL-8, however, did not inhibit lymphomagenesis, suggesting that IL-8 is not essential for angiogenesis in this model. To examine other parameters of angiogenesis, we identified expression of vascular endothelial growth factor in the lymphomas. These data suggest that angiogenesis accompanies EBV-associated B-cell lymphoma development, but IL-8 is not essential for this process. Thus, the SCID mouse model is amenable to testing of anti-angiogenic factors.