Showing papers on "Angiogenesis published in 1985"

Cutaneous tissue repair: Basic biologic considerations. I

Abstract: Wound repair of the integument is reviewed in the context of new developments in cell biology and biochemistry. Injury of the skin and concomitant blood vessel disruption lead to extravasation of blood constituents, followed by platelet aggregation and blood clotting. These events initiate inflammation and set the stage for repair processes. The macrophage plays a pivotal role in the transition between wound inflammation and repair (granulation tissue formation), since this cell both scavenges tissue debris and releases a plethora of biologically active substances that include growth factors. Although concrete evidence is lacking, growth factors are probably at least partially responsible for the angiogenesis and fibroplasia (granulation tissue) that gradually fill the wound void. If the epidermal barrier is disrupted during injury, reepithelialization begins within 24 hours and proceeds first over the margin of residual dermis and subsequently over granulation tissue. The signals for angiogenesis, fibroplasia, neomatrix formation, and reepithelialization in wound repair are not known, but a number of possibilities are discussed. Matrix remodeling is the last stage of wound repair and gradually increases the scar tensile strength to 70% to 80% of normal skin.

Angiogenesis is stimulated by a tumor-derived endothelial cell growth factor.

Abstract: A growth factor mitogenic for BALB/C 3T3 cells and capillary endothelial cells was isolated from a rat chondrosarcoma and purified to homogeneity. Purification was accomplished by a combination of BioRex 70 cation exchange chromatography and heparin affinity chromatography. The pure chondrosarcoma-derived growth factor (ChDGF) had a molecular weight of about 18.000. The angiogenesis activity of pure ChDGF was tested by measuring its ability to vascularize the chorioallantoic membrane (CAM) and yolk sac membrane of the developing chick. The ability of ChDGF to induce the growth of limbal vessels in the rat cornea was also measured. To quantitate the angiogenesis response, a unit system based on the growth factor activity of ChDGF for 3T3 cells was adopted. ChDGF was found to have a specific activity of about 5 units/ng when applied to 3T3 cells. About 300–600 units of ChDGF in the two types of developing chick membrane and 30–50 units of ChDGF in the rat cornea were found to stimulate noninflammatory angiogenesis.

Toward an understanding of angiogenesis: search and discovery.

Abstract: I was in the navy, assigned not to a ship but to the Naval Medical Research Institute in Bethesda. Frederick Becker and David Long were there also. The three of us had been invited to interrupt our respective residencies to fulfill 2 years of military obligation. It was 1960, and the navy was curious about the feasibility of blood substitutes for use in its hospitals aboard aircraft carriers. We experimented with hemoglobin solutions. Dog thyroid glands were perfused with hemoglobin solutions in isolated glass chambers for periods of more than a week. The perfused organs appeared to be viable, but could they support growth? We injected tumor cells from a mouse melanoma into the thyroid glands. Tiny tumors developed and grew rapidly at first but then stopped enlarging at diameters of 1-2 mm [I]. What was the explanation for the arrested growth of these tumors? Immune rejection seemed unlikely because these organs were perfused with acellular fluids. Furthermore, when tumors were transplanted to a freshly isolated thyroid gland, the size limit held. However, when reimplanted into the donor mice, these tumors grew rapidly, beyond 1 cm3, and killed their hosts. In the mice the tumors were vascularized; in the isolated perfused organs, they were not. (fig. 1). What we had witnessed,

Induction of angiogenesis by bovine brain derived class 1 heparin-binding growth factor.

Abstract: The angiogenic capacity of the class 1 heparin-binding growth factor from bovine brain, an anionic endothelial cell mitogen of Mr 16 000, has been evaluated. Its ability to induce the growth of new blood vessels has been assessed by means of two established assay systems. On the embryonic chick chorioallantoic membrane dose-response studies demonstrate that 160 ng (10 pmol) of mitogen is required to induce angiogenesis in greater than 50% of the eggs within 72 h. In the presence of 1 unit of exogenous heparin only 40 ng of mitogen (2.5 pmol) is needed to induce a similar response. Moreover, this occurs within 48 h, indicating that heparin also augments the angiogenic response by enhancing the rate of induction of angiogenesis. Eighty nanograms (5 pmol) of mitogen also induces the ingrowth of new blood vessels into the rabbit cornea, both in the presence and in the absence of heparin. These results establish that the class 1 heparin-binding growth factor from bovine brain is an angiogenesis factor. Importantly, the neovascularization induced by this angiogenesis factor is enhanced by heparin. The mechanistic implications for neovascularization under certain normal and pathological conditions are discussed.

Angiogenesis and its inhibitors.

Abstract: The hypothesis that solid tumors are angiogenesis-dependent has, in the past decade, generated much new work aimed at understanding the mechanism of angiogenesis itself. Many laboratories in this country and abroad are now studying some aspect of this intriguing problem. Some investigations are focused mainly on tumor angiogenesis, whereas others are centered on angiogenesis that occurs in physiologic situations or that dominates certain non-neoplastic pathologic states. These efforts have brought about [a] the development of bioassays for angiogenesis; [b] the partial purification (and in one case the complete purification) of angiogenic factors from neoplastic and non-neoplastic cells; [c] the development of new polymer technology for the sustained release of these factors and of other macromolecules in vivo; [d] the cloning and long-term culture of capillary endothelial cells; [e] the demonstration of the role of nonendothelial cells, such as mast cells, in modulating angiogenesis; [f] the discovery of angiogenesis inhibitors; and [g] the recent demonstration that certain animal tumors will undergo complete regression when treated by antiangiogenesis alone. The effects of angiogenesis inhibitors provide the most compelling evidence for the role of angiogenesis in tumor growth. That it is now possible not only to inhibit tumor growth but also to eradicate some experimental tumors speaks strongly for a therapeutic approach that may some day be useful in clinical oncology. Conceivably, the original goal to understand the role of angiogenesis in tumor growth will lead to the use of angiogenesis inhibitors in other non-neoplastic diseases.

Fibrin degradation and angiogenesis: Quantitative analysis of the angiogenic response in the chick chorioallantoic membrane

Abstract: Fibrin deposition and removal is a feature common to major pathological processes such as wound healing, chronic inflammation and tumour invasion: processes involving the ingrowth of new blood vessels. Low molecular weight fibrin degradation products (MW less than 50,000) are now shown to induce angiogenesis in the chick chorioallantoic membrane (CAM). This effect has also been shown by new quantitative assays to be associated with stimulation of both DNA and protein synthesis. Autoradiography indicates that all cell types in the CAM are stimulated to divide, and it is proposed that fibrin degradation products are a pathological growth factor.

Mast cells and tumors. The specific enhancement of tumor proliferation in vitro.

Abstract: Mast cells were found to be unique among the peritoneal leukocytes by virtue of their capacity to enhance profoundly the proliferation of a variety of tumors in vitro. This phenomenon occurs at mast cell/tumor ratios which reflect the stoichiometry of host cell/tumor relationships in vivo. The growth factor was found to reside in mast cell granules and was identified as heparin by sequential purification and enzymatic degradation. This cellular interaction was tumor-specific, although isolated granules could enhance fibroblast proliferation. The findings are discussed in relation to previous morphologic studies, reports of in vitro mast-cell-mediated tumor cytotoxicity, and the role of mast cells in angiogenesis and connective tissue proliferation.

The fine structure of tumor blood vessels. I. Participation of non-endothelial cells in tumor angiogenesis.

Abstract: This report provides fine structural evidence that, dependent upon the malignancy, tumor as well as mesenchymal cells may participate actively in the neovascularization of experimental tumors grown in transparent tissue chambers implanted into skinfolds of syrian hamsters. Such non-endothelial cells may help to promote angiogenesis in two different ways: (1) They are incorporated into capillary sprouts thereby accelerating the growth rate of the latter independent of endothelial cell proliferation. (2) Extravascular cells (tumor and mesenchymal elements) become integrated in varying numbers into the linings of comparatively large blood-perfused vessels. This facilitates the rapid establishment and functional remodelling of the microvascular bed to adapt the microcirculation to the varying local demands of the growing tumor. If these results can be confirmed for other tumors, and if they are independent of the tumor's environment and the experimental protocol, then we will have to reconsider the significance of tumor angiogenesis as a realistic biological model from which general conclusions with regard to neovascularization in non-tumorous tissues may be drawn.

Mast cells and tumour angiogenesis: The tumour‐mediated release of an endothelial growth factor from mast cells

Abstract: Exposure to tumour cells has previously been shown to induce mast cells to degranulate and release heparin. Isolated mast-cell granules were found to be mitogenic for endothelial cells in vitro. This effect was a property of mast-cell heparin, whose potency as a mitogen exceeded that of commercial heparins. The basis of this difference lay in the proteoglycan structure of the molecule. The release of heparin in mast-cell-tumour co-cultures was examined by both endothelial cell proliferation and isotopic techniques. The kinetics and mode of release are described. The results are discussed in relation to the role of the mast cell in angiogenesis assays and tumour neovascularization.

Fibrin-enhanced endothelial cell organization.

Abstract: The formation of cloned bovine endothelial cells into capillary-like tubes is accelerated from 3–7 days to 2–18 h in the presence of fibrin. Indirect immu-nofluorescence showed the presence of both fibrin and fibronectin in the strands along which the cells organized. Electronmicroscopy revealed the same type of cell structures as form in the absence of fibrin; it also revealed a gradual decrease with time of the fibrin within the putative lumen. Fibrin and fibronectin are commonly present during angiogenesis in vivo, thus these in vitro observations may well have relevance to the in vivo process.

Tumor-derived angiogenesis factors from rat Walker 256 carcinoma: an experimental investigation and review.

Abstract: Angiogenesis, the process of developing a hemovascular network, is an essential feature of the growth of solid tumors, and is induced by factors secreted by tumor cells Assay procedures suitable for the investigation of angiogenesis, and for the screening of angiogenesis factors during purification are reviewed; and a number of reports describing the purification of angiogenesis factors, primarily from the rat Walker 256 carcinoma as starting material, are discussed Work from the authors' laboratory is also presented Walker 256 cells grown in large-scale culture were the source of a reproducible and homogeneous source of angiogenic material Factors secreted by these cells were isolated by a series of chromatographic steps Ion exchange chromatography on carboxymethyl-Sephadex produced two active fractions, one of which was fractionated into several macromolecular species by lectin affinity and hydrophobic adsorption chromatography The other gave a high molwt, active fraction that was resolved into a low molwt, active component and a non-angiogenic but possibly carrier molecule with a molwt of 140,000 While none of the angiogenic factors were identified chemically, the results demonstrate the existence of both high and low molwt tumor-secreted angiogenic substances, confirming the hypothesis for tumor-induced angiogenesis and predicting potential means to interfere with the process of tumor growth

Regulation of cell growth by vitreous humour.

Abstract: Extracts of normal vitreous have been found to inhibit angiogenesis in two animal models: tumour-induced neovascularization in the rabbit corneal micropocket and retinal extract-induced angiogenesis in the chick chorioallantoic membrane assay. Using in vitro assays, we have found recently that an extract of bovine vitreous, free of hyaluronic acid, inhibits proliferation of cells in the aortic wall, i.e. endothelium and smooth muscle cells, as well as capillary and corneal endothelium. The inhibition is dose-dependent, as determined by either cell count or [3H]thymidine incorporation, and not due to cytotoxicity, as demonstrated with a double-label thymidine assay. The inhibitor is trypsin-sensitive and heat-stable (95 degrees C for 10 min). Conversely, proliferation of pericytes, lens epithelium and fibroblasts (dermal and corneal) was stimulated by the vitreous extract. This mitogenic activity was heat-labile. Growth of pigment epithelium and several tumour cell lines was unaffected. The data demonstrate that normal vitreous contains a heat-stable growth inhibitor specific for endothelium and smooth muscle cells, and a non-specific heat-labile mitogen. The paradoxical effect of this antiangiogenic factor on arterial and capillary contractile cells, smooth muscle and pericytes, suggests a basic difference in the regulation of the two vasculatures. The results suggest that a substance in normal vitreous may be important in controlling neovascularization that results from diabetic and other retinopathies, and could be useful for inhibiting tumour-induced angiogenesis.

Long-term serial cultivation of arterial and capillary endothelium from adult bovine brain.

Abstract: Cerebrovascular endothelial cells from adult bovine brain were carried successfully in long-term, serial culture. Endothelial cells were obtained from the middle and anterior cerebral arteries and from capillaries isolated from grey matter of the cerebral cortex or caudate nucleus. Capillary cells were found to grow best in RPMI 1640 with 20% fetal bovine serum. They did not require tumor-conditioned medium or matrix-coated surfaces, although fibronectin was used to enhance the initial plating efficiency of the primary cultures. The same conditions were used to support satisfactory growth of arterial endothelial cells; however they did not grow as rapidly as the capillary cells. Retention of endothelial-specific characteristics were shown for capillary-derived cells carried up to Passage 28, arterial-derived cells up to Passage 11, and after frozen storage of both types of cultured cells. Cultures of both arterial and capillary cells stained positively for Factor VIII antigen, exhibited a nonthrombogenic surface, and produced prostacyclin in response to arachidonic acid. Arterial endothelial cells produced more prostacyclin than capillary endothelium. The capillary cells had a unique tendency to assume a ringlike morphology after subculture and sometimes formed capillarylike networks of cell cords in dense cultures. When cultured in a three-dimensional plasma clot, capillary and arterial endothelial cells, but none of the other cell types studied, organized into tubelike structures reminiscent of capillary formation in vivo. The availability of long-term cultures of cerebrovascular endothelial cells provides an opportunity to compare properties of arterial and capillary endothelium from the same tissue and to investigate such processes as angiogenesis and blood-brain barrier induction.

Adenosine nucleotides, adenosine and adenine as angiogenesis factors.

Abstract: Investigations of migratory behaviour of porcine endothelial cells using Boyden technique showed strong chemotactic activity of ADP, adenosine and adenine. These substances were postulated to be angiogenesis factors in vivo.

Ultrastructure of cerebellar capillary hemangioblastoma

Abstract: Electron microscopy and computerized morphometric techniques were employed to examine pericyte ultrastructure and to assess quantitatively their relationship to endothelial cells in five cases of cerebellar capillary hemangioblastoma A total of 97 cross-sectioned capillary profiles were studied Pericyte coverage of capillary ranged from 302% to 973% with a mean value of 687%, which is higher as compared with the available data from the cerebral cortex, skeletal and cardiac muscle, and pulmonary capillaries The higher pericyte coverage of capillary suggests that pericyte is an active component of cerebellar capillary hemangioblastoma and may have a close functional relationship to endothelial cells Pericytes contained bundles of parallel microfilaments along the adluminal side and in the terminal processes, and exhibited an intimate “peg-and-socket” relationship with endothelial cells, suggesting a contractile function of pericytes and their possible role in regulating capillary lumina and focal blood flow The finding of abundant micropinocytic vesicles along the abluminal side of the cytoplasmic membrane indicates an active metabolic exchange between pericytes and the interstitium It is possible that in cerebellar hemangioblastoma pericytes may act as a mechanical and metabolic monitor barrier for endothelial cells

Cooperation between fibronectin and heparin in the mobilization of capillary endothelium.

Abstract: Angiogenesis is indispensable to sustain promotion and growth of metastases. As a contribution to the understanding of the angiogenesis process, the experiments reported showed that: (a) fibronectin is involved in the mobilization of capillary endothelium which is the first event in angiogenesis; (b) antifibronectin serum can block the mobilization, and neutralization of the antiserum can restore it; (c) the combination of fibronectin + heparin is a powerful mobilizer of capillary endothelium, and (d) fragments of the fibronectin and heparin molecules in combination can mobilize capillary endothelium as effectively as the intact molecules. The results are interpreted to indicate that molecules normally present in the extracellular matrix like heparin and fibronectin, may act as angiogenesis effectors when the physiological structure of the tissue is altered, for instance by lytic enzymes released by metastatic neoplastic cells.

The Effect of Inhibition of Angiogenesis in Granulation Tissue on Wound Healing and the Fibroblast

Abstract: Protamine sulfate given in high doses can inhibit angiogenesis in the granulation tissue generated in an open wound. This is reflected by changes consistent with delayed vascular maturation in the morphology of the regenerating vessels seen at the gross, microscopic, and ultrastructural levels. A coincidental delay in wound healing as evidenced by impaired wound contraction occurs, although fibroblast duplication and myofibroblast differentiation appear intact. However, the fibroblasts contain little endoplasmic reticulum, the site of synthetic activity, and the surrounding collagen appears loose and disorganized. To unite these observations into a coherent pattern, we review the proposal that the endothelial cell, the fibroblast, and the extracellular stroma are interdependent and that maturation of each is necessary to maintain the momentum of wound healing. Our findings fit this mechanistic hypothesis but do not prove it. The abnormal vasoformation that may be initiated by protamine's anticoagulant properties could set the stage for impaired fibroblast synthetic activity. If collagenous stroma is deficient, both endothelial maturation and wound contraction wound fail. Although we saw these final events, to prove a series of cause and effect changes would require further study of the oxygen tension and the fibrin and collagen levels in granulation tissue.

Weibel-Palade bodies as a marker for neovascularization induced by tumor and rheumatoid angiogenesis factors.

Abstract: An analysis was made of ultrastructural changes in capillary endothelial cells in experimentally induced angiogenesis and in a human pathological situation known to involve increased angiogenesis. Chick chorioalloantoic membrane (CAM) showing a positive angiogenic response to low-molecular weight angiogenesis factor isolated from rat Walker sarcoma or from human rheumatoid joint was compared with untreated CAM. Serotonintreated CAM provided an additional control in that serotonin has the capacity to stimulate endothelial cell growth in vitro but did not induce angiogenesis on the CAM. Human rheumatoid joints were studied using normal healthy human joints as controls. The number of Weibel-Palade (W-P) bodies per unit of cytoplasmic area were higher in tumor angiogenesis factor-treated CAMs (not significant) and rheumatoid angiogenesis factor-treated CAMs ( P < 0.008) than in untreated controls. These differences were more pronounced if W-P body volumetric density was analyzed ( P in both cases <0.008). Serotonin-treated control CAMs did not show higher numbers of W-P body or greater WPV than untreated controls. Numbers of W-P body and W-P body volumetric density were higher ( P < 0.008) in rheumatoid joints than normal joints. Median values for W-P body number were 16-fold higher and, for W-P body volumetric density, they were up to 30-fold higher in rheumatoid joints.

Effects of cortisone with and without heparin on angiogenesis induced by prostaglandin E1 and by S180 cells, and on growth of murine transplantable tumours.

Abstract: Cortisone acetate, locally applied in sustained-release pellets, is effective in inhibiting angiogenesis induced by prostaglandin E1 in the rabbit cornea. The inhibitory effect of cortisone is not increased by addition of heparin. Similar results were obtained with angiogenesis induced by S180 cells. The effects of cortisone with and without heparin were also studied on 5 transplantable murine tumours: 3 variants of B16 melanoma, Lewis lung carcinoma and fibrosarcoma M4. The effect of cortisone in slowing the growth rate of tumours was modestly potentiated by heparin, but no regression of the tumour mass occurred.

Rheumatoid arthritis and osteoarthritis synovial fluid effects on primary human endothelial cell cultures.

Abstract: Since vascular proliferation may be important in the pathogenesis of rheumatoid arthritis (RA) and/or osteoarthritis (OA), this study examined the induction of angiogenesis by these synovial fluids (SF). Four of 11 (36%) RA and 2 of 6 (33%) OA SF caused early morphological changes in human endothelial cell cultures. SF from 7 of 11 (63%) RA and 4 of 8 (50%) OA patients resulted in the late formation of tabular networks morphologically resembling capillaries observed in vivo. Early morphological changes in cultures were associated with a significantly (p less than 0.05) longer duration of disease in patients with RA. Factors present in the SF of RA and OA patients may play a role in the excessive vascularization which often occurs in these arthropathies.

A morphometric study of the microvasculature of a rat glioma.

Abstract: Luthert P. J. & Lantos P. L. (1985) Neuropathology and Applied Neurobiology, 11, 461–473 A morphometric study of the microvasculature of a rat glioma Intracerebral tumours were produced in BD IX rats by the inoculation of neoplastic astrocytes. Following perfusion-fixation, 10 firw sections were stained with toluidine-blue and photographed so as to construct photomontages embracing deep tumour, tumour edge and surrounding brain. Thirteen montages from 9 tumours were studied. Successive 200 mm samples were analysed with a digitizing tablet and, for each sample the following were measured (a) the number of vascular profiles per unit area, (b) the mean vessel circumference and diameter and (c) the approximate fractions of the sample occupied by tumour and necrosis. Towards the tumour centre, vascular density dropped to 20% of that for normal cortex but vessel size more than doubled. The surface area fraction of the vessels reached a maximum at the tumour edge. These results provide not only an anatomical framework for both a variety of physiological studies and for the investigation of angiogenesis in this system but also have pharmacokinetic implications for the treatment of brain tumours.

Long-term serial cultivation and growth requirements for human umbilical vein endothelial cells.

Abstract: Human umbilical vein endothelial cells (HUV-EC) grew rapidly in vitro in medium supplemented with epidermal growth factor, fetal bovine serum (FBS) and human diploid fibroblast-conditioned medium. The effect of FBS could be replaced partially by bovine serum albumin, cholesterol, and vitamin E, and completely by further addition of serum dialysate or refeeding every other day. Among these components, fibroblast-conditioned medium is essential for HUV-EC growth. The HUV-EC were cultured serially for over 50 population doublings in the 10% FBS containing fibroblast-conditioned medium and for over 40 population doublings in the serum-free medium. Mitogenic factor(s) present in the medium conditioned by fibroblasts may be related to endothelial cell growth factor and play an important role in angiogenesis and regeneration of vascular endothelium in vitro.

Wound fluid angiogenesis factor stimulates the directed migration of capillary endothelial cells

Abstract: During wound healing, new capillaries grow into the wound site. An angiogenesis factor isolated from wound fluid stimulates the movement of capillary endothelial cells in a filter migration assay. Experiments were carried out to determine whether the movement seen in the assay was chemokinetic or chemotactic. Capillary endothelial cells were plated onto a collagen-coated coverslip and inverted over a visualization apparatus. Cells exposed to a constant concentration of wound fluid angiogenesis factor (WAF) were more mobile than cells not exposed to WAF, and this movement was chemokinetic. When exposed to a gradient of WAF, the cells translocated toward the higher concentration; this directional movement was chemotactic. Cells in a gradient of WAF morphologically aligned with the gradient. These data support the idea that wound healing angiogenesis is regulated by the chemotaxis of capillary endothelial cells.

Interspecies chimeras: an experimental approach for studies on embryonic angiogenesis.

Abstract: Most studies on the growth of vessels have so far focused on the tumour vascularization or that occurring in the area of inflammation. The mechanisms of embryonic angiogenesis have not been characterized in such a detail and only relatively few experimental studies have been carried out to analyse the origin and development of the embryonic vasculature. It is not known if the vessels develop in situ in each organ rudiment or by invasion of earlier committed vascular cells. Because morphological analyses of in vivo tissues have proved to be unreliable for judging the origin of vascular cells, new methods have been presented. Nuclear differences between of some species can be used to trace the origin of cells in interspecies transplantation experiments. This review presents data on the biology of vasculogenesis and shows how interspecies chimeras can be used in studies on angiogenesis. For example, the transplantation experiments with embryonic kidneys are described in more detail.