Showing papers on "Angiogenesis published in 1983"

Oxygen tension regulates the expression of angiogenesis factor by macrophages

Abstract: When cultured in a hypoxic environment similar to that found in the center of a wound, macrophages secreted active angiogenesis factor into the medium. Under conditions similar to those of well-oxygenated tissue, macrophages did not secrete active angiogenesis factor. Macrophages that secreted the factor at hypoxic conditions stopped secreting it when returned to room air. Thus the control of angiogenesis in wound healing may be the result of macrophages responding to tissue oxygen tension without the necessity of interacting with other cell types or biochemical signals.

In vitro rapid organization of endothelial cells into capillary-like networks is promoted by collagen matrices.

Abstract: We have studied the behavior of cloned capillary endothelial cells grown inside a three dimensional collagen matrix. Cell monolayers established on the surface of collagen gels were covered with a second layer of collagen. This induced the monolayers of endothelial cells to reorganize into a network of branching and anastomosing capillary-like tubes. As seen by electron microscopy, the tubes were formed by at least two cells (in transverse sections) delimiting a narrow lumen. In addition, distinct basal lamina material was present between the abluminal face of the endothelial cells and the collagen matrix. These results showed that capillary endothelial cells have the capacity to form vessel-like structures with well-oriented cell polarity in vitro. They also suggest that an appropriate topological relationship of endothelial cells with collagen matrices, similar to that occurring in vivo, has an inducive role on the expression of this potential. This culture system provides a simple in vitro model for studying the factors involved in the formation of new blood vessels (angiogenesis).

Capillary endothelial cell cultures: phenotypic modulation by matrix components.

Abstract: Capillary endothelial cells of rat epididymal fat pad were isolated and cultured in media conditioned by bovine aortic endothelial cells and substrata consisting of interstitial or basement membrane collagens. When these cells were grown on interstitial collagens they underwent proliferation, formed a continuous cell layer and, if cultured for long periods of time, formed occasional tubelike structures. In contrast, when these cells were grown on basement membrane collagens, they did not proliferate but did aggregate and form tubelike structures at early culture times. In addition, cells grown on basement membrane substrata expressed more basement membrane constituents as compared with cells grown on interstitial matrices when assayed by immunoperoxidase methods and quantitated by enzyme-linked immunosorbent inhibition assays. Furthermore, when cells were grown on either side of washed, acellular amnionic membranes their phenotypes were markedly different. On the basement membrane surface they adhered, spread, and formed tubelike structures but did not migrate through the basement membrane. In contrast, when seeded on the stromal surface, these cells were observed to proliferate and migrate into the stromal aspect of the amnion and ultimately formed tubelike structures at high cell densities at longer culture periods (21 d). Thus, connective tissue components play important roles in regulating the phenotypic expression of capillary endothelial cells in vitro, and similar roles of the collagenous components of the extracellular matrix may exist in vivo following injury and during angiogenesis. Furthermore, the culture systems outlined here may be of use in the further study of differentiated, organized capillary endothelial cells in culture.

Mobilization of capillary endothelium in vitro induced by effectors of angiogenesis in vivo.

Abstract: An assay to measure endothelial cell mobilization on a gelatin substratum has been developed. Utilization of the gelatin-agarose and Boyden chamber assays established that: (a) fragments or extracts of corneas treated with several effectors of angiogenesis in vivo acquired the capacity to mobilize the capillary endothelium in vitro; (b) this mobilization was selective for the capillary endothelium; endothelium from aorta and fibroblasts from human skin or rabbit cornea were unresponsive; and (c) among the effectors of angiogenesis utilized alone; i.e., without the intermediary action of the cornea, none were able to mobilize the capillary endothelium in vitro, except for the heparin-copper complex. The data are interpreted to indicate that new formation of capillaries in vivo is the end result of a cascade of events of which heparin and copper are important components.

Angiogenesis-dependent Tumor Spread in Reinforced Fibrin Clot Culture

Abstract: To investigate the role played by developing microvessels in the spread of tumors, segments of rat aorta were cultured with aggregates of NBT-II-81, a cell line derived from squamous cell carcinoma of rat bladder. Aortic rings cultured in plasma clot gave rise to microvascular networks composed of branching endothelial channels. Aggregates of carcinoma in contact with fibrin clot alone grew slowly and by expansion. When the proliferating branching endothelial sprouts and channels contacted the tumor aggregates, the pattern of neoplastic growth changed abruptly, as carcinoma cells infiltrated the fibrin clot, migrating and proliferating in periendothelial location. Some vascular channels were disrupted and permeated by cords of invading tumor cells. Ultrastructural studies revealed intimate association between invading epithelial cells and endothelial cells. Focal fusion of the endothelial basal lamina with the basal lamina of the tumor cells was observed. Our results demonstrate that angiogenesis in vitro, i.e. , in absence of active circulation, markedly enhanced the spread of a carcinoma in plasma clot and modified its pattern of growth. This indicates that other vascular-related factors beside nutritional gradients from the circulation attract tumor cells along endothelial paths.

Neovascular responses induced by cultured aortic endothelial cells.

Abstract: Neovascularization was studied in the chorioallantoic membrane of the chick embryo after implantation of bovine aortic endothelial and smooth muscle cells, Swiss and BALB/c 3T3 cells and human diploid fibroblasts cultured separately on microcarrier beads. Quantitative analysis of neovascularization indicated a 3 1/2-fold increase in the number of blood vessels responding to endothelial cells while smooth muscle cells induced a twofold increase when compared to the response of beads without cells. Skin fibroblasts and Swiss 3T3 cells did not elicit a comparable response. The marked angiogenic response induced by endothelial cells was characterized by a 137% increase in total vessel length and a 35% increase in average vessel area when compared to controls. Two of the properties required for an angiogenesis factor--stimulation of cellular migration and proliferation--can also be demonstrated using endothelial cell-conditioned medium in cell culture systems. Medium from cultured bovine aortic endothelium stimulates DNA synthesis, proliferation, and migration of smooth muscle cells. In addition, conditioned media from both endothelial cells and smooth muscle cells produced an angiogenic response in the chorioallantoic membrane assay, which was comparable to that produced by intact cells growing on microcarrier beads. Similar responses were not evident with medium conditioned by other cell types. These results indicate the potential importance of endothelial cells and endothelial cell products in regulating blood vessel growth.

Angiogenic lymphokines of activated T-cell origin.

Abstract: Inflammation often is accompanied by neovascularization. This is especially evident in the normally avascular cornea, where an angiogenic response occurs with keratitis or during graft rejection. In the current experiments, supernatants from cultured lymph node cell populations were incorporated in Elvax 40 slow-release polymers and implanted in the corneal stroma. Separation of whole lymph node cells into highly T-enriched and macrophage populations permitted investigation of soluble cell products responsible for angiogenesis. It was found that sepharose-linked concanavalin A was an adequate stimulus for the generation of angiogenic activity, as assessed in the rabbit corneal micropocket assay. Cytoadherence, nylon wool column cell separation, and mitomycin C treatment of individual populations were used to demonstrate that stimulated T cells are a source of angiogenic material. The time-course and magnitude of the angiogenic response were equivalent in normal and x-irradiated leukopenic animals. These observations demonstrate that a potent angiogenic lymphokine secreted by stimulated T cells is active in the corneal micropocket assay system. Invest Ophthalmol Vis Sci 24:1595-1601, 1983 The growth of corneal vessels in response to inflammation has been studied extensively since Schoefl's 1 work on the fine structure of the new vasculature in corneal wound healing. An angiogenic response has been documented in a wide range of inflammatory events including arthritis, 2 delayed hypersensitivity, 34 and corneal graft rejection. 5 The class of leukocytes that can elicit an angiogenic response has been the subject of considerable investigation. Fromer and Klintworth 6 have implicated polymorphonuclear leukocytes (PMN) as the primary source of angiogenic activity in experiments using a rat corneal model. Activated macrophages and supernatants from activated macrophages have been found to elicit angiogenesis in guinea pig cornea, 4 and also have been found to be mitogenic for fibroblasts, 7 an important cell in wound repair. Using lymphocyte transfer reaction, Auerbach and his colleagues have evaluated the angiogenic response to stimulated allogeneic lymphocytes in mouse 8 and rabbit 9 corneas, on the chorioallantoic membrane (CAM), 9 and in mouse skin. 10 Recent work from Auerbach's laboratory, using the Lyt and Thy series of monoclonal antibodies, has implicated the effector T cell as a source of the angiogenic stimulus in mice," and has shown that the major antigenic stimulus in the event is associated with the I-region alloantigens. 12

Relationship of angiogenesis factor in synovial fluid to various joint diseases.

Abstract: A low-molecular-weight freely dialysable angiogenesis factor has been isolated from 49 synovial fluids obtained from patients with various joint diseases. An analysis of disease type and incidence of freely dialysable angiogenesis activity showed that the osteoarthrotic group had a significantly higher incidence than all the other groups (p = 0.0332). Angiogenesis factor has also been detected in a bound form in the retentates of fluids which gave positive results for dialysable factor. The possibility that an imbalance between carrier-bound and free factor may have a causative role in disease is discussed.

Extracellular nonmitogenic angiogenesis factor and method of isolation thereof from wound fluid

Abstract: A nonmitogenic angiogenesis factor is isolated from wound fluid by dialysis to include materials in the molecular size range of 2,000 to 14,000, lyophilization, and chromatography. The nonmitogenic angiogenesis factor is identified by activity by corneal implant assay and by cell migration assay. The angiogenesis factor is also characterized by inactivity by mitogenesis assay.

Role of mast cells in experimental tumour angiogenesis.

Abstract: Although the morphological features of angiogenesis are well documented and many promoting factors are known, the pharmacological mechanisms for the development of new vessels are not understood. Compounds found in platelets and/or mast cells--adenosine diphosphate, 5-hydroxytryptamine, histamine and heparin--caused endothelial cell growth stimulation in vitro: tumour angiogenesis factor did not. These same vasoactive compounds, as well as tumour angiogenesis factor, induced neovascularization on the chick chorioallantoic membrane. The increased vascularity produced by tumour angiogenesis factor was associated with considerable numbers of mast cells. These findings, together with an appreciation of the biochemical armoury of the mast cell and how its products could relate to the morphological steps of angiogenesis, and a realization that known anti-angiogenesis factors could all act through inhibition of mast cell products, strongly implicate the mast cell in the inductive mechanisms of neovascularization.

Protease-mediated enhancement of lymphocyte-induced angiogenesis in X-ray irradiated mice.

Abstract: SummaryAngiogenesis was induced in mice by intradermal injection of semi-syngeneic splenocytes, and after three days the number of newly formed blood vessels at the injection site was counted. When recipients were total-body irradiated with 700 R 2 hours before the lymphocyte injection, the angiogenesis was significantly higher than in non-irradiated mice. The angiogenesis enhancement was of a systemic (not local) character as revealed in experiments with shielding of irradiated animals. This enhancement was not due to X-ray dependent immunosuppression, as shown in experiments with non-irradiated, pharmacologically immunosuppressed mice. Decreased angiogenesis was observed in irradiated mice after treatment with cortisone acetate, aprotinin, and EACA. The results suggest that porteases might be involved in mediating the angiogenesis enhancement after X-irradiation.

Angiogenic responses elicited from chorioallantoic membrane vessels by neoplastic, preneoplastic, and normal mammary tissues from GR mice.

Abstract: Neoplastic tumors are able to elicit the ingrowth of new capillaries, a process known as angiogenesis. The chorioallantoic membrane (CAM) of chicken embryos was used in an assay for this response, and normal mammary glands and various mammary growths from GR mice, including plaques, hyperplasic alveolar nodules, and hormone-dependent and hormone-independent tumors were tested. Fifteen percent of the male mammary glands tested were positive, as were 28% of the resting female mammary glands. Fifty percent of the plaques and 63% of the hyperplastic alveolar nodules tested induced neovascularization. Eighty percent of the hormone-dependent tumors and 97% of the hormone-independent tumors tested elicited angiogenesis. A fine-structural study revealed that capillaries invaded to within less than 0.5 microns of the tumor cells, but no penetration of tumor cells through the basal lamina was observed. Positive responses were directly correlated with the neoplastic potential of the tissues tested, indicating that angiogenesis can predict mammary gland growths most likely to become malignant.

Cultured tumor cells produce chemotactic factors specific for endothelial cells: a possible mechanism for tumor-induced angiogenesis.

Abstract: The production of stimulants of endothelial cell motility by cultured tumor cells was studied. Spontaneously transformed murine fibroblasts in culture produced activity that stimulated migration of endothelial cells, while this kind of activity was not detected in media from cultures of the normal counterparts of the transformed cells. Furthermore, a line of murine tumor cells (HB4), known to induce vasoformative sarcomas in vivo, was found to produce in culture a strong chemoattractant specific for endothelial cells. Since the tumor-derived material also caused vessel ingrowth when implanted in the rabbit eye, these results suggest that the angiogenesis observed during tumor growth may involve chemoattractants for endothelial cells produced by tumor cells.

Quantitation of angiogenesis factor in bovine retina and tumour extracts by means of radioimmunoassay.

Abstract: Using an antiserum raised against tumour angiogenesis factor (TAF) we have developed a radioimmunoassay for retina and tumour angiogenesis factor(s). This antiserum was previously shown to bind to both human and animal tumour extracts and to inhibit the angiogenesis induced by TAF in vivo. TAF from rat Walker tumour was used for iodination by the chloramine-T method. An excess of 125I-labelled TAF was incubated with TAF antibody in the absence (maximum binding) and presence (inhibition of maximum binding) of unlabelled tissue extract. A double antibody technique was used to separate free and bound TAF. Unlabelled human Wilms tumour TAF was used as a standard. The extent of inhibition of 125I-TAF-anti-TAF binding provided a measure of TAF in tissue extracts examined. Extracts of normal bovine retina, cornea, lung, aorta, lymph nodes, iris, vitreous humour, and human tumours and normal human pituitary and liver were assayed. Only bovine retina and human tumours were found to contain angiogenesis factor. These findings, together with our earlier results, suggest that angiogenesis factor from both bovine retina and human tumours induce angiogenesis in vivo and possess common antigenic determinants. The presence of angiogenesis factor in healthy retina and its relationship to neovascularisation in clinical conditions is discussed.

Signs of capillary formation in the early phase of wound healing in children

Abstract: Summary Small silicone rubber tubes containing a standard size viscose cellulose sponge (Cellstic) were implanted in wounds of 23 children at the end of routine surgery. The Cellstics were drawn out of the wounds 1–4 days after implantation. Single cells and cell aggregates, called cell aggregation centers (CAC), were washed out of the Cellstic sponges by the retrograde injection technique, centrifuged, studied histologically and histochemically for cytochrome oxidase activity. Small leukocytic CACs and solitary cells, also including some macrophages and a few endothelial cells, were observed on the first postoperative day. On the second postoperative day and thereafter 2–4 endothelial cells were attached to each other surrounding a lumen. These capillary segment-looking formations (CSF) resembled capillary segments and they were seen both inside and outside the (now larger) CACs. Outside the CACs solitary endothelial cells occasionally exhibited ring-like formations. High cytochrome oxidase activity was observed especially in many CSFs but the solitary and the ring-shaped endothelial cells, in- and outside the CACs, also showed intense oxidative metabolism.

Inhibition of tumor-induced angiogenesis and of tumor growth by activated lymphocytes.

Abstract: Inhibition in angiogenesis (neo-vascularization) and of growth of a tumor piece graft in the anterior chamber of the eye of mice have been observed in the presence of activated syngeneic lymphocytes.