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J A Nagy

Publications and source records attributed to J A Nagy.

At least 37 records · Page 2Linked to original sources
PubMed1997

Vascular permeability factor/vascular endothelial growth factor: a multifunctional angiogenic cytokine.

VPF/VEGF is a multifunctional cytokine that contributes to angiogenesis by both direct and indirect mechanisms. On the one hand, VPF/VEGF stimulates the endothelial cells lining nearby microvessels to proliferate, to migrate and to alter their pattern of gene expression. On the other hand, VPF/VEGF renders these same microvascular endothelial cells hyperpermeable so that they spill plasma proteins into the extravascular space, leading to profound alterations in the extracellular matrix that favor angiogenesis. These same principles apply in tumors, in several examples of non-neoplastic pathology, and in physiological processes that involve angiogenesis and new stroma generation. In all of these examples, microvascular hyperpermeability and the introduction of a provisional, plasma-derived matrix precede and accompany the onset of endothelial cell division and new blood vessel formation. It would seem, therefore, that tumors have made use of fundamental pathways that developed in multicellular organisms for purposes of tissue defense, renewal and repair. VPF/VEGF, therefore, has taught us something new about angiogenesis; namely, that vascular hyperpermeability and consequent plasma protein extravasation are important--perhaps essential--elements in its generation. However, this finding raises a paradox. While VPF/VEGF induces vascular hyperpermeability, other potent angiogenic factors apparently do not, at least in sub-toxic concentrations that are more than sufficient to induce angiogenesis (Connolly et al., 1989a). Nonetheless, wherever angiogenesis has been studied, the newly generated vessels have been found to be hyperpermeable. How, therefore, do angiogenic factors other than VPF/VEGF lead to the formation of new and leaky blood vessels? We do not as yet have a complete answer to this question. One possibility is that at least some angiogenic factors mediate their effect by inducing or stimulating VPF/VEGF expression. In fact, there are already clear example of this. A number of putative angiogenic factors including small molecules (e.g. prostaglandins, adenosine) as well as many cytokines (e.g. TGF-alpha, bFGF, TGF-beta, TNF-alpha, KGF, PDGF) have all been shown to upregulate VPF/VEGF expression. Further studies that elucidate the crosstalk among various angiogenic factors are likely to contribute significantly to a better understanding of the mechanisms by which new blood vessels are formed in health and in disease.

Adult↗
PubMed1996

Vesiculo-vacuolar organelles and the regulation of venule permeability to macromolecules by vascular permeability factor, histamine, and serotonin.

In contrast to normal microvessels, those that supply tumors are strikingly hyperpermeable to circulating macromolecules such as plasma proteins. This leakiness is largely attributable to a tumor-secreted cytokine, vascular permeability factor (VPF). Tracer studies have shown that macromolecules cross tumor vascular endothelium by way of a recently described cytoplasmic organelle, the vesiculo-vacuolar organelle or VVO (VVOs are grapelike clusters of interconnected, uncoated vesicles and vacuoles). However, equivalent VVOs are also present in the cytoplasm of normal venules that do not leak substantial amounts of plasma protein. To explain these findings, we hypothesized that VPF increased the permeability of tumor blood vessels by increasing VVO function and that the VVOs of normal venules were relatively impermeable in the absence of VPF stimulation. To test this hypothesis, VPF was injected intradermally in normal animals after intravenous injection of a soluble macromolecular tracer, ferritin, whose extravasation could be followed by electron microscopy. VPF caused normal venules to leak ferritin, and, as predicted by our hypothesis, ferritin extravasated by way of VVOs, just as in hyperpermeable tumor microvessels. Ultrathin (14-nm) serial electron microscopic sections and computer-aided three-dimensional reconstructions better defined VVO structure. VVOs occupied 16-18% of endothelial cytoplasm in normal venules. Individual VVOs were clusters of numerous (median, 124) interconnected vesicles and vacuoles that formed complex pathways across venular endothelium with multiple openings to both luminal and abluminal surfaces. Like VPF, histamine and serotonin also stimulated ferritin extravasation across venules by way of VVOs. Together, these data establish VVOs as the major pathway by which soluble plasma proteins exit venules in response to several mediators that increase venular hyperpermeability. These same mediators also increased the extravasation of colloidal carbon, but this large particulate nonphysiological tracer exited venules primarily through endothelial gaps.

Animals↗
PubMed1996

The vesiculo-vacuolar organelle (VVO): a distinct endothelial cell structure that provides a transcellular pathway for macromolecular extravasation.

The vesiculo-vacuolar organelle (VVO) is a recently described organelle found in the cytoplasm of endothelial cells that line tumor microvessels and normal venules. VVOs are grape-like clusters of interconnecting uncoated vesicles and vacuoles, bounded by trilaminar unit membranes, that span the entire thickness of vascular endothelium, thereby providing a potential trans-endothelial connection between the vascular lumen and the extravascular space. Macromolecular tracers preferentially cross hyperpermeable tumor microvessels through VVOs. The present investigation was undertaken to elucidate further the ultrastructure and function of VVOs in a murine ovarian carcinoma (MOT) and in normal venules. Morphometry revealed that VVOs were enormous cytoplasmic structures (median area, 0.12-0.14 microns2 in single electron micrographs). Moreover, the individual vesicles and vacuoles that comprised VVOs were on average substantially larger than capillary caveolae and followed a non-normal distribution that was skewed to the right. Specimen tilting provided conclusive evidence that individual VVO vesicles and vacuoles communicated with each other and with the endothelial cells' plasma membranes by stomata, some of which were closed by diaphragms composed of a single membrane. Studies with two tracers, ferritin (FE, diameter approximately 11 nm) and horseradish peroxidase (HRP, diameter approximately 5 nm), revealed that passage of macromolecules through VVOs was regulated at the level of stomatal diaphragms, thereby demonstrating a mechanism for controlling the passage of macromolecules across endothelial cells. Thus, compared with tumor microvessels, little circulating FE and HRP entered the VVOs of normal venular endothelium because stomata joining vesicles and vacuoles to each other and to the lumen and ablumen were closed. VVOs and their component vesicles/vacuoles were readily distinguished from endosomal organelles such as coated vesicles and multivesicular bodies, which also accumulated FE and HRP. Our findings indicate that VVOs provide a major pathway for the extravasation of circulating macromolecules across endothelia taller than capillary endothelium and suggest that upregulated VVO function accounts for the well-known hyperpermeability of tumor blood vessels.

Animals↗
PubMed1996

Increased hyaluronan at sites of attachment to mesentery by CD44-positive mouse ovarian and breast tumor cells.

The mouse ovarian ascites tumor, MOT, and mammary ascites tumor, TA3/St, served as models to follow changes in hyaluronan levels during tumor growth, attachment, and invasion. Subsequent to introduction of tumor cells into the peritoneal cavity, hyaluronan accumulated intraperitoneally and at the initial sites of attachment of tumor cells and cell clumps to the mesenteric surface; the latter co-localized with sites of fibrin deposition as reported earlier. Subsequently, high levels of hyaluronan accumulated throughout the interior of the mesentery. Because neither tumor cell line synthesized substantial amounts of hyaluronan in culture, the large accumulations observed in the mesenteries and ascites fluid of tumor-bearing animals most likely resulted from increased synthesis and secretion by peritoneal-lining mesothelial cells and/or fibroblasts in response to stimulation by the tumor cells or their products. TA3/St tumor cells were universally positive for the hyaluronan receptor, CD44, whereas approximately 90% of MOT tumor cells were CD44-negative. However, the great majority of MOT or TA3/St cells that initially attached to the mesentery were strongly CD44 positive. We propose that hyaluronan-rich matrix is involved in tumor cell attachment to the mesentery possibly via interaction with tumor cell surface CD44.

Animals↗
PubMed1995

Pathogenesis of ascites tumor growth: angiogenesis, vascular remodeling, and stroma formation in the peritoneal lining.

In the accompanying papers, we demonstrated that two murine ascites tumors (MOT and TA3/St) induced peritoneal lining blood vessels to become hyperpermeable to plasma proteins, leading to extravasation of fibrinogen and its clotting to cross-linked fibrin in peritoneal lining tissues (peritoneal wall, mesentery, and diaphragm). In solid tumors, vascular hyperpermeability and fibrin deposition lead to the generation of vascularized connective tissue. In order to determine whether fibrin had similar consequences in ascites tumors, the vasculature and stroma of peritoneal lining tissues were analyzed at successive intervals after i.p. tumor cell injection. In both MOT and TA3/St ascites tumors, the size and number of peritoneal lining microvessels increased significantly by 5-8 days. Subsequently, peritoneal lining vessels increased in cross-sectional area by as much as 15-fold and peritoneal vascular frequency increased by up to 11-fold. Incorporation of [3H]thymidine by mesenteric blood vessels was negligible in control animals but came to involve 20 and 40% of endothelial cells lining mesenteric vessels in MOT and TA3/St ascites tumor-bearing mice, respectively. After an early dramatic increase in cross-sectional area, peritoneal lining microvessels subsequently underwent a novel form of remodeling to smaller average size as the result of transvascular bridging by endothelial cell cytoplasmic processes. Thus, both of the ascites tumors studied here induced angiogenesis and stroma similar to that elicited when these same tumors were grown in solid form. However, stroma developed more slowly in ascites than in solid tumors and was entirely confined to a compartment (peritoneal lining tissues) that was distinct from that (peritoneal cavity) containing the majority of tumor cells and ascites fluid. These findings are consistent with the hypothesis that vascular hyperpermeability, induced in both solid and ascites tumors by tumor cell-secreted vascular permeability factor, is a common early step in tumor angiogenesis, resulting in fibrinogen extravasation, fibrin deposition, and likely other alterations of the extracellular matrix that together stimulate new vessel and fibroblast ingrowth.

Animals↗
PubMed1995

Pathogenesis of ascites tumor growth: vascular permeability factor, vascular hyperpermeability, and ascites fluid accumulation.

Previous studies have shown that accumulation of tumor ascites fluid results in large part from increased permeability of peritoneal lining vessels (Nagy et al., Cancer Res., 49: 5449-5458, 1989; Nagy et al., Cancer Res., 53: 2631-2643, 1993). However, the specific microvessels rendered hyperpermeable have not been identified nor has the basis of peritoneal vascular hyperpermeability been established. To address these questions, TA3/St and MOT carcinomas, well-characterized transplantable murine tumors that grow in both solid and ascites form, were studied as model systems. Ascites tumor cells of either type were injected i.p. into syngeneic A/Jax and C3Heb/FeJ mice, and ascites fluid and plasma were collected at intervals thereafter up to 8 and 28 days, respectively. Beginning several days after tumor cell injection, small blood vessels located in tissues lining the peritoneal cavity (mesentery, peritoneal wall, and diaphragm) became hyperpermeable to several macromolecular tracers (125I-human serum albumin, FITC-dextran, colloidal carbon, and Monastral Blue B). Increased microvascular permeability correlated with the appearance in ascites fluid of vascular permeability factor (VPF), a tumor cell-secreted mediator that potently enhances vascular permeability to circulating macromolecules. VPF was measured in peritoneal fluid by both a functional bioassay and a sensitive immunofluorometric assay. The VPF concentration, total peritoneal VPF, ascites fluid volume, tumor cell number, and hyperpermeability of peritoneal lining microvessels were found to increase in parallel over time. The close correlation of peritoneal fluid VPF concentration with the development of hyperpermeable peritoneal microvessels in these two well-defined ascites tumors suggests that VPF secretion by tumor cells is responsible, in whole or in part, for initiating and maintaining the ascites pattern of tumor growth.

Animals↗
PubMed1995

Pathogenesis of ascites tumor growth: fibrinogen influx and fibrin accumulation in tissues lining the peritoneal cavity.

In the immediately preceding paper, we demonstrated that the microvasculature supplying peritoneal lining tissues of mice bearing either of two transplantable ascites carcinomas was hyperpermeable to circulating macromolecules. Solid tumors have been shown to exhibit similar levels of microvascular hyperpermeability, leading to extravasation of plasma proteins, including fibrinogen which clots on extravasation to form an extravascular fibrin gel. To determine whether similar extravasation and clotting of plasma fibrinogen occurred in ascites tumors, we used 125I-labeled fibrinogen (125I-F) as a tracer to measure inflow of fibrinogen into the peritoneal cavities, and influx and accumulation of fibrinogen/fibrin in the peritoneal lining tissues (peritoneal wall, mesentery, and diaphragm) of mice bearing syngeneic TA3/St or MOT ascites tumors. The percentage of circulating 125I-F that extravasated into the peritoneal cavity was increased from 10- to 50-fold in mice bearing either ascites tumor. Influx into the peritoneal walls of ascites tumor-bearing mice was 3-7 times that of control mice and became maximal on day 8 (TA3/St) and day 15 (MOT). Accumulation of 125I-F in ascites fluid and peritoneal lining tissues was also increased substantially in mice bearing these ascites tumors, reaching maximal values on days 7-8 (TA3/St) and 19-29 (MOT) at levels 2- to 3-fold (peritoneal wall) and 33- to 148-fold (ascites fluid) above control levels. Significant amounts of the 125I-F that accumulated in the peritoneal lining tissues of ascites tumor-bearing animals were insoluble in 3 M urea, consistent with clotting of 125I-F to cross-linked fibrin. Autoradiographs of SDS-PAGE gels performed on extracts of peritoneal lining tissues of both ascites tumors revealed the characteristic signature of cross-linked fibrin, i.e., gamma-gamma dimers and alpha-polymers. Fibrin was also identified in peritoneal lining tissues of both ascites tumors by immunohistochemistry. Taken together, these data indicate that fibrinogen, like other circulating macromolecules, extravasates into the peritoneal cavity and peritoneal lining tissues of ascites tumor-bearing mice and does so with kinetics similar to those of other macromolecular tracers we have studied. Moreover, a portion of the fibrinogen that extravasated into peritoneal lining tissues clotted to form a cross-linked fibrin meshwork which trapped tumor cells and favored their attachment to the peritoneal surface. By analogy with solid tumors, such fibrin deposits may also be expected to have a role in initiating angiogenesis and the generation of mature tumor stroma.

Animals↗
PubMed1995

Keratinocyte-derived vascular permeability factor (vascular endothelial growth factor) is a potent mitogen for dermal microvascular endothelial cells.

Expression of vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is markedly increased in the epidermis of lesional psoriatic skin and in healing skin wounds. In this study, we characterized the effects of several cytokines and growth factors on the expression and secretion of VPF/VEGF mRNA and protein by cultured human epidermal keratinocytes, as well as the effect of VPF/VEGF on the growth of cultured human dermal microvascular endothelial cells. Transforming growth factor-alpha, epidermal growth factor, and phorbol myristate acetate markedly stimulated VPF/VEGF mRNA expression by cultured keratinocytes; as in psoriatic skin, the three most common VPF/VEGF isoforms (encoding proteins of 121, 165, and 189 amino acids) were upregulated to an equal extent. Transforming growth factor (TGF)-alpha, epidermal growth factor, and phorbol myristate acetate also enhanced the secretion of VPF/VEGF by keratinocytes; in contrast, a number of other cytokines including interleukin (IL)-1, IL-6, IL-8, tumor necrosis factor-alpha, interferon-gamma, and transforming growth factor-beta did not induce VPF/VEGF secretion. The VPF/VEGF secreted by keratinocytes was biologically active in that, like recombinant human VPF/VEGF, it potently stimulated dermal endothelial cell proliferation. Scatchard analysis revealed two high-affinity VPF/VEGF binding sites on dermal endothelial cells with dissociation constants of 51 pM and 2.9 pM. These results suggest that the avascular epidermis has the capacity to regulate dermal angiogenesis and microvascular permeability by a paracrine mechanism involving the secretion of VPF/VEGF. Similar mechanisms may be anticipated in a variety of inflammatory and neoplastic skin diseases characterized by microvascular hyperpermeability, edema, and angiogenesis.

Binding Sites↗
PubMed1995

Ultrastructural localization of vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) to the abluminal plasma membrane and vesiculovacuolar organelles of tumor microvascular endothelium.

Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a cytokine secreted by many animal and human tumors, activated macrophages, keratinocytes, rheumatoid synovial cells, embryonic tissues, and by cultured epithelial and mesenchymal cell lines. It acts selectively on vascular endothelial cells to increase their permeability to circulating macromolecules and to stimulate their replication. Although not detectably expressed by vascular cells in the human and animal tumors we have studied, VPF/VEGF accumulates in the microvessels supplying tumors and certain inflammatory reactions in which VPF/VEGF is also overexpressed. Light microscopic immunohistochemistry lacked the resolution necessary to localize VPF/VEGF precisely in such vessels. Therefore, we used a pre-embedding immunocytochemical method to localize VPF/VEGF at the ultrastructural level in the new blood vessels that are elicited in the peritoneal walls of mice bearing a transplantable mouse ascites tumor of ovarian origin. Intense immunostaining for VPF/VEGF was observed on the abluminal plasma membrane of tumor-associated microvascular endothelial cells and in vesiculovacuolar organelles (VVOs) present in these same endothelial cells. (VVOs are recently described cytoplasmic organelles present in tumor vascular endothelium that provide an important pathway for extravasation of circulating macromolecules.) In contrast to labeling of the abluminal plasma membrane and VVO vesicles and vacuoles, endothelial cytoplasmic organelles, such as multivesicular bodies and Weibel-Palade bodies, and the underlying basal lamina, did not stain with antibodies to VPF/VEGF. The distribution of VPF/VEGF here described corresponds to that anticipated for high-affinity VFP/VEGF receptors, although binding of VPF/VEGF to other endothelial cell surface structures, such as plasma membrane proteoglycans, is also a possibility.

Animals↗
PubMed1995

Mesothelial cell-mediated gene therapy: feasibility of an ex vivo strategy.

We have developed a model system in the rat to test the feasibility of recombinant protein expression by genetically modified peritoneal mesothelial cells following autologous peritoneal implantation. Rat primary peritoneal mesothelial cells, isolated from parietal peritoneum by enzymatic digestion, were stably transduced (using a Moloney murine leukemia virus (MoMLV)-derived retroviral vector, BAG, expressing the Escherichia coli lacZ gene) to mark the cells with a reporter protein (beta-galactosidase, beta-gal). Such transduced mesothelial cells, tagged with DiO, a fluorescent lipophilic dye used for long-term tracing of transplanted cells, were then reseeded on the denuded peritoneal surface of syngeneic recipients. DiO-labeled, BAG-transduced mesothelial cells were observed to repopulate the denuded areas and remain attached there for > 90 days. Moreover, these genetically modified mesothelial cells continued to express the reporter gene product in vivo (ie beta-gal activity was present for at least 1 month). Our results demonstrate the feasibility of ex vivo gene therapy using peritoneal mesothelial cells.

Animals↗
PubMed1995

Systemic delivery of a recombinant protein by genetically modified mesothelial cells reseeded on the parietal peritoneal surface.

To evaluate the ability of genetically modified peritoneal mesothelial cells to deliver recombinant proteins to the systemic circulation, we used our previously described mesothelial cell-based ex vivo gene therapy strategy. Rat primary peritoneal mesothelial cells, isolated from parietal peritoneum by enzymatic digestion, were stably transfected (using strontium phosphate DNA co-precipitation) with the plasmid pSVTKgh to express a secreted reporter gene product, human growth hormone (hgh). Such hgh-secreting mesothelial cells were reseeded on the denuded peritoneal surface of syngeneic recipients and delivery of the reporter gene product to the systemic circulation was monitored by analysis of serum samples for the presence of hgh at various times after mesothelial cell implantation. Polymerase chain reaction (PCR) analysis demonstrated that the hgh-transfected mesothelial cells repopulated the denuded areas and remained attached there for at least 12 weeks. Moreover, these genetically modified mesothelial cells continued to express the reporter gene product in vivo and secreted hgh in sufficient quantity to be detected in the systemic circulation (ie statistically significant amounts of hgh could be measured in the serum of cyclosporine A-treated rats for at least 2 months; Mann-Whitney test, P < 0.05). Our results demonstrate the successful, sustained, systemic delivery of a recombinant protein by genetically modified peritoneal mesothelial cells following their reattachment to the peritoneal surface, and suggest the potential of ex vivo mesothelial cell-mediated gene therapy for the treatment of inherited or acquired disorders requiring delivery of therapeutic proteins to the circulation.

Animals↗
PubMed1994

Compartmental distribution of tumor-specific monoclonal antibodies in human melanoma xenografts.

Monoclonal antibodies (MAb) are attractive for tumor therapy because of their exquisite specificity. Although a majority of tumor cells in small (< or = 20 mg) solid tumors can be labeled following systemic administration of antitumor cell MAbs, little quantitative information is available as to the distribution of these MAbs within the several compartments that comprise solid tumors. Our goal was to provide such data in a well-characterized melanoma xenograft system. In accord with earlier work, i.v.-injected, melanoma-specific MAbs 436 and IND1, directed, respectively, against the 125 kD and HMW-melanoma-associated antigens, accumulated in M21 and SK-MEL-2 tumor xenografts in amounts of approximately 20% of injected dose/g. However, only 20-24% of the MAbs present in tumor xenografts was bound to tumor cells; the great majority (76-80%) was in the tumor extracellular fluid (ECF) and collagenous residue fractions. These results could not be accounted for by MAb degradation or release of MAbs from tumor cells during xenograft dissociation. Rather, they reflected in large part interactions of MAbs with antigens which tumors had shed into the ECF. Thus, 48 h after i.v. injection of 20 micrograms of melanoma-specific, biotin-tagged MAb, 46-66% of that present in the tumor ECF was complexed with melanoma-associated antigens. Overall, 61-73% of the MAbs recovered from tumor xenografts were bound to tumor antigens (either to tumor cells themselves or to tumor-shed antigens). In contrast, only approximately 4% of a melanoma-nonspecific MAb (B72.3) accumulated per g tumor after i.v. injection and nearly all of this was free in the ECF. Consistent with these data, fluorescence microscopy revealed that i.v.-injected, fluorescein-tagged MAbs achieved highest concentrations in tumor stroma, particularly at the tumor-host interface. Flow cytometry of dissociated solid tumors revealed that both the fraction of MAb-labeled tumor cells and the amount of MAb/tumor cell could be increased by increasing the administered i.v. dose of melanoma-specific MAb. Nonetheless, even at the highest i.v. injected dose (300 micrograms), 15-37% of tumor cells lacked detectable MAb labeling. Taken together, the data indicate that delivery of tumor cell-specific MAbs to solid tumors cannot be equated with their delivery to tumor cells. This distinction is important for immunotherapeutic approaches that require MAb contact with tumor cells.

Animals↗
PubMed1994

Induction of vascular endothelial growth factor expression by prostaglandin E2 and E1 in osteoblasts.

PGE1 and PGE2 are potent stimulators of bone formation. Osteogenesis is strongly dependent on angiogenesis. Vascular endothelial growth factor (VEFG), a secreted endothelial cell-specific mitogen, has been implicated in physiological and pathological angiogenesis. The aim of this study was to examine the possible role of VEGF in PG stimulation of bone formation. We found that in rat calvaria-derived osteoblast-enriched cells and in the osteoblastic RCT-3 cell line PGE2 and E1 increased VEGF mRNA and protein levels. The increased expression of VEGF mRNA produced by PGE2 was rapid (maximal at 1 h), transient (declined by 3 h), potentiated by cycloheximide, and abolished by actinomycin D. PGE2 had no effect on VEGF mRNA stability, suggesting transcriptional regulation of VEGF expression by PGF2. Rp-cAMP, a cAMP antagonist, suppressed VEGF mRNA induced by PGE2, indicating cAMP mediation. The upregulation of VEGF expression by PGE2 in the preosteoblastic RCT-1 cells was potentiated by treatment with retinoic acid, which induces the differentiation of these cells. The upregulation of VEGF mRNA by PGE2 was inhibited by dexamethasone treatment. In addition, Northern blot analysis showed that VEGF mRNA is expressed in adult rat tibia. In summary, we documented, for the first time, the expression of VEGF in osteoblasts and in bone tissue. Stimulation of VEGF expression by PGs and its suppression by glucocorticoids, which, respectively, stimulate and suppress bone formation, strongly implicate the involvement of VEGF in bone metabolism.

Alprostadil↗
PubMed1993

Vascular permeability factor (vascular endothelial growth factor) in guinea pig and human tumor and inflammatory effusions.

Vascular permeability factor (VPF), also known as vascular endothelial growth factor, is a dimeric M(r) 34,000-42,000 glycoprotein that possesses potent vascular permeability-enhancing and endothelial cell-specific mitogenic activities. It is synthesized by many rodent and human tumor cells and also by some normal cells. Recently we developed a sensitive and specific time-resolved immunofluorometric assay for quantifying VPF in biological fluids. We here report findings with this assay in guinea pigs and patients with both malignant and nonmalignant effusions. Line 1 and line 10 tumor cells were injected into the peritoneal cavities of syngeneic strain 2 guinea pigs, and ascitic fluid, plasma, and urine were collected at various intervals. Within 2 to 4 days, we observed a time-dependent, parallel increase in VPF, ascitic fluid volume, and tumor cell numbers in animals bearing either tumor line; in contrast, VPF was not detected in plasma or urine, even in animals with extensive tumor burdens. However, low levels of VPF were detected in the inflammatory ascites induced by i.p. oil injection. In human studies, high levels of VPF (> 10 pM) were measured in 21 of 32 effusions with cytology-documented malignant cells and in only seven of 35 effusions without cytological evidence of malignancy. Thus, VPF levels in human effusions provided a diagnostic test for malignancy with a sensitivity of 66% and a specificity of 80% (perhaps as high as 97% in that six of the seven cytology-negative patients with VPF levels > 10 pM had cancer as determined by other criteria). As in the animal tumor models, VPF was not detected in serum or urine obtained from patients with or without malignant ascites. Many nonmalignant effusions contained measurable VPF but, on average, in significantly smaller amounts than were found in malignant effusions. VPF levels in such fluids correlated strongly (p = 0.59, P < 0.001) with monocyte and macrophage content. Taken together, these data relate ascitic fluid accumulation to VPF concentration in a well-defined animal tumor system and demonstrate, for the first time, the presence of VPF in human malignant effusions. It is likely that VPF expression by tumor and mononuclear cells contributes to the plasma exudation and fluid accumulation associated with malignant and certain inflammatory effusions. The VPF assay may prove useful for cancer diagnosis as a supplement to cytology, especially in tumors that grow in the pleural lining but not as a suspension in the effusions that they induce.

Adult↗
PubMed1993

Pathogenesis of malignant ascites formation: initiating events that lead to fluid accumulation.

Initiating events leading to the accumulation of malignant ascites in the peritoneal cavity were investigated in two syngeneic transplantable murine ascites-producing tumors, MOT mouse ovarian tumor and the TA3/St mammary carcinoma. The transport of two tracers, 125I-labeled human serum albumin (125I-HSA) and 51Cr-labeled red blood cells (51Cr-RBC), into and out of the peritoneal cavity was studied at early times after i.p. tumor cell injection, prior to abundant fluid accumulation, and at intervals of 5 to 360 min after i.v. or i.p. tracer injection. Tracer influx and efflux rates were estimated from the mass of tracer passing into or out of the peritoneal cavity following a bolus injection of tracer into either the blood or the peritoneal cavity. Efflux of 125I-HSA from the peritoneal cavity was markedly reduced (3- to 5-fold) within 1 day of i.p. injection of either type of tumor cell. Significantly reduced efflux preceded any increase in tumor cell number and by itself did not induce peritoneal fluid accumulation. 125I-HSA tracer influx from plasma to peritoneal fluid did not increase detectably until 5 to 7 days after tumor cell injection, when the tumor cell number had increased by 10- to 100-fold. Only at relatively late stages of ascites tumor growth, when the flow rate into the peritoneal cavity had increased relative to the flow rate out of the peritoneum, was there net peritoneal fluid accumulation. Thus, increased influx, in addition to impaired efflux, were required for malignant ascites accumulation. Following i.p. injection, the efflux rates of 125I-HSA always exceeded those of 51Cr-RBC, even in ascites tumor-bearing animals. Furthermore, 125I-HSA tracer disappeared from the peritoneal cavity more rapidly than it appeared in the plasma, suggesting that 125I-HSA moves more rapidly through the channels by which 51Cr-RBC egress from the peritoneum (primarily diaphragmatic lymphatics) and/or has access to additional pathways not open to 51Cr-RBC. Finally, flow rates into and out of the blood and peritoneum were used to obtain kinetic parameters that characterized tracer transport: k1, the rate constant for tracer transport from the blood to the peritoneum; k2, the rate constant for tracer transport from the peritoneal cavity to the blood; and k6, the rate constant for tracer transport from the peritoneal cavity to surrounding interstitial tissue.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Vascular permeability factor (VPF, VEGF) in tumor biology.

Vascular permeability factor (VPF), also known as vascular endothelial growth factor (VEGF), is a multifunctional cytokine expressed and secreted at high levels by many tumor cells of animal and human origin. As secreted by tumor cells, VPF/VEGF is a 34-42 kDa heparin-binding, dimeric, disulfide-bonded glycoprotein that acts directly on endothelial cells (EC) by way of specific receptors to activate phospholipase C and induce [Ca2+]i transients. Two high affinity VPF/VEGF receptors, both tyrosine kinases, have thus far been described. VPF/VEGF is likely to have a number of important roles in tumor biology related, but not limited to, the process of tumor angiogenesis. As a potent permeability factor, VPF/VEGF promotes extravasation of plasma fibrinogen, leading to fibrin deposition which alters the tumor extracellular matrix. This matrix promotes the ingrowth of macrophages, fibroblasts, and endothelial cells. Moreover, VPF/VEGF is a selective endothelial cell (EC) growth factor in vitro, and it presumably stimulates EC proliferation in vivo. Furthermore, VPF/VEGF has been found in animal and human tumor effusions by immunoassay and by functional assays and very likely accounts for the induction of malignant ascites. In addition to its role in tumors, VPF/VEGF has recently been found to have a role in wound healing and its expression by activated macrophages suggests that it probably also participates in certain types of chronic inflammation. VPF/VEGF is expressed in normal development and in certain normal adult organs, notably kidney, heart, adrenal gland and lung. Its functions in normal adult tissues are under investigation.

Animals↗