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Biomedical subjects

I R Hart

Publications and source records attributed to I R Hart.

At least 91 records · Page 5Linked to original sources
PubMed1991

Influence of tumor-derived interleukin 1 on melanoma-endothelial cell interactions in vitro.

Human melanoma cell lines that express high constitutive levels of the metastasis-associated marker intercellular adhesion molecule 1 (ICAM-1) were found to secrete interleukin 1 (IL-1) in vitro. Experiments with neutralizing antibodies showed that this cytokine was responsible for their expression of ICAM-1 but not that of two other progression/metastasis markers, Muc-18 and Gp IIb/IIIa. The IL-1 present in melanoma-conditioned medium induced the expression of vascular cell adhesion molecule 1, endothelial-leukocyte adhesion molecule 1, and ICAM-1 on human umbilical vein endothelial cells (ECs) in culture and increased the rate at which melanoma cells and ECs adhered to each other. IL-1-producing melanoma lines adhered significantly more rapidly to ECs than did non-IL-1-producing lines, and this enhancement was reduced by prior incubation of the melanoma cells with neutralizing anti-IL-1 antibodies. Similarly, endothelial cells treated with conditioned medium from IL-1-producing melanoma lines adhered significantly more rapidly to melanoma cells than did ECs treated with medium from non-IL-1-producing melanoma lines, and this enhancement was abolished by addition of anti-IL-1 antibodies to EC cultures in conditioned medium. Blocking antibodies to endothelial vascular cell adhesion molecule 1, endothelial-leukocyte adhesion molecule 1, and ICAM-1 failed to inhibit melanoma-EC adhesion, but an antibody to tumor cell GpIIb/IIIa did block adhesion by up to 44%. The ability to secrete IL-1 could increase the metastatic potential of melanoma cells by stimulating tumor cell-EC adhesion.

Antigens, CD↗
PubMed1991

Protein kinase C down-regulation, and not transient activation, correlates with melanocyte growth.

The nontumorigenic, immortal line of murine melanocytes, Mel-ab, requires the continual presence of biologically active phorbol esters for growth (R.E. Wilson et al., Cancer Res., 49:711-716, 1989). Comparable treatments of B16 murine melanoma cells result in partial inhibition of cell proliferation. The role of protein kinase C (PKC) in the modulation of growth of cells from these two melanocytic cell lines has been investigated. Significant levels of PKC were present in quiescent Mel-ab cells as determined by Western blotting, whereas no immunoreactive protein was detected in cell extracts from either proliferating Mel-ab or B16.F1 cells. Phosphorylation of a Mr 80,000 protein, which by one- and two-dimensional gel analysis comigrated with the known Mr 80,000 protein substrate of PKC in fibroblasts, was induced in 12-O-tetradecanoylphorbol-13-acetate-stimulated quiescent Mel-ab cells but not in proliferating Mel-ab cells or B16.F1 melanoma cells. Direct measurement of PKC activity in these cells demonstrated a 10-fold greater level of activity in quiescent Mel-ab cells (262 +/- 50 pmol/min/mg SD) compared with growing cells (22.8 +/- 11.8 pmol/min/mg SD). An intermediate level of activity was detected in proliferating B16.F1 melanoma cells (148.5 +/- 20.4 pmol/min/mg SD). The subcellular distribution of PKC was dependent upon the growth state of the cells such that quiescent Mel-ab cells displayed a higher level of activity in the cytosol, whereas growing Mel-ab cells displayed greater activity in the particulate fraction. Like many other transformed lines, B16.F1 melanoma cells constitutively expressed the majority of enzyme activity in the particulate fraction. Measurement of [3H]phorbol ester binding in intact cells paralleled the PKC activation data such that quiescent Mel-ab cells displayed binding of 1612 +/- 147 cpm/10(6) cells, whereas proliferating Mel-ab and B16.F1 melanoma cells displayed binding of 652 +/- 28 and 947 +/- 81 cpm/10(6) cells, respectively. Membrane-permeant diacylglycerol analogues, which activated but did not down-regulate PKC, were devoid of growth-stimulating effects on melanocytes, even in the presence of the specific diacylglycerol kinase inhibitor, R59022. Together, these data show that PKC down-regulation, and not activation, correlates with the growth of melanocytes in culture.

Animals↗
PubMed1991

Cell adhesion receptor expression during melanoma progression and metastasis.

Many steps in melanoma metastasis involve cell-cell or cell-matrix adhesive interactions. The surface molecules which mediate these processes therefore play an important role in regulating melanoma dissemination and their level of expression may alter during the course of tumor progression. Human melanocyte strains and melanoma cell lines have been characterised with regard to levels of cell surface receptors of the integrin family. Increased amounts of at least two integrins, VLA-4 (alpha 4 beta 1) and VnR (alpha v beta 3), appeared to correlate with progression in this tumor, type. A novel VnR composed of an alpha v beta 1 association has been observed in one melanoma cell line and there is the possibility that heterogeneity of integrin composition could affect biological behavior of these tumors. CD44, a cell surface glycoprotein which functions as the major receptor for hyaluronate, is another molecule whose expression increases in transformed cells of the melanocytic lineage. Iterative sorting on the FACS for stable variants, of both human and murine melanomas, expressing low and high levels of CD44 established that lack of expression of this molecule correlated with impaired ability to form pulmonary tumor nodules subsequent to i.v. injection into appropriate recipient mice. These findings illustrate that an understanding of the regulation of melanoma adhesion receptors could provide insights into the process of tumor spread.

Humans↗
PubMed1991

Divergent effects of flavone acetic acid on established versus developing tumour blood flow.

Flavone Acetic Acid (FAA) exerts much of its effect by reducing tumour blood flow. Previous studies on FAA-induced changes in blood flow have used established tumours with a functional microvasculature. Using radioactive Xenon(133Xe) clearance to monitor local blood flow we show that the effects of FAA are dependent on the presence of this functional microvasculature with no evidence that FAA inhibits the actual development of tumour microcirculation. Thus, administration of multiple doses of FAA around the time of tumour cell injection failed to diminish t1/2 values of 133Xe (e.g. t1/2 16 min for FAA vs 14 min for saline controls at 10 days) or to affect tumour volumes (5.55 +/- 0.06 cm3 in FAA-treated animals vs 5.7 +/- 1.3 cm3 in controls at 25 days). In marked contrast a single dose of FAA (200 mg kg-1 body weight) 2 weeks after tumour cell injection dramatically extended t1/2 times (47 min for FAA vs 7 min for controls; P less than 0.001) and significantly reduced tumour burden. This effect is specific for tumour microvasculature and is not directed simply at new vessels since a similar treatment of animals with implanted-sponge-induced granulation tissue had no effect on t1/2 times (6.8 +/- 1.1 min for FAA at 200 mg kg-1 vs 7.2 +/- 1.0 min for saline-treated controls.

Animals↗
PubMed1991

Photocytotoxic efficacy of sulphonated species of aluminium phthalocyanine against cell monolayers, multicellular spheroids and in vivo tumours.

The problem of relying solely on in vitro data to predict photosensitiser efficacy was demonstrated by examining the uptake and the ability to mediate photocytotoxicity of mono-, di-, tri- and tetra-sulphonated species of chloroaluminium phthalocyanine (AlS1-4Pc) in monolayer cultures of murine Colo 26 cells and in both monolayer and spheroid cultures of human WiDr cells. Cells treated in vitro, whether in monolayer or as spheroids, with the less sulphonated derivatives, AlS1Pc and AlS2Pc, were more susceptible to photocytotoxicity than those treated with AlS3Pc, cells treated with AlS4Pc were even less susceptibile to the cytotoxic effects of light irradiation. Generally these results mirrored the cellular uptake in vitro. When WiDr spheroids were increased in size from 250 microns to 500 microns there was a reduction in uptake of AlS1Pc and AlS2Pc which was reflected by the decreased sensitivity of the larger spheroids to the effects of light irradiation. AlS1Pc had no effect against Colo 26 cells growing as s.c. tumours in syngeneic BALB/c mice; whereas AlS3Pc, AlS2Pc and AlS4Pc produced significant reductions in tumour weights 5 days post laser light irradiation. Of these, AlS2Pc had the most dramatic effect on the colony forming efficiency of tumour cells recovered 24 h after PDT. While, despite their effects on tumour size, AlS3Pc and AlS4Pc scarcely affected the subsequent viability of cells from dissociated tumours. Thus the in vitro efficacy of the sulphonated species of phthalocyanines is not necessarily predictive of their in vivo effectiveness.

Animals↗
PubMed1991

Tumor cell progression and differentiation in metastasis.

The development and evolution of tumors is regulated by both genetic and epigenetic events. It is thought that these processes tend to drive neoplastic development in opposing directions so that tumor progression, predominantly as a consequence of mutational events, leads to increasing tumor aggression. Conversely the induction of differentiation, largely through epigenetic mechanisms, tends to cause tumors to evolve to a more benign phenotype. However, these generalizations are a simplistic view of a complex dynamic event where both processes can be overlaid within a single neoplasm. Using malignant melanoma as a model system the alterations in gene expression and their effects upon metastatic dissemination, that accompany some of these changes, both natural and induced, are described.

Animals↗
PubMed1990

Role of tumor necrosis factor in flavone acetic acid-induced tumor vasculature shutdown.

Flavone acetic acid (FAA), a novel investigational antitumor agent, has been shown to cause early vascular shutdown in several experimental murine tumors, and this phenomenon is believed to be crucial to FAA's antitumor effects. However, the basis of this FAA-induced tumor vascular shutdown is unknown. In this study a radioactive tracer-clearance technique has been used as an objective indication of tumor blood flow to show that i.p. administered FAA induces a progressive and sustained reduction in blood flow in a colon 26 tumor growing s.c. in syngeneic mice. As early as 1 h after administration, there was a significant increase in the t1/2 clearance value for intratumorally injected 133Xe, reaching a peak at 3 h (117.3 +/- 36.4 versus 7.8 +/- 0.85 min for controls). Significant inhibition of blood flow was still apparent 48 h after a single injection of drug. This FAA-induced vascular shutdown was virtually abolished in tumor-bearing mice pretreated with an antiserum against tumor necrosis factor, while no such effect was observed in controls pretreated with nonimmune serum (t1/2 of 10.8 +/- 1.2 versus 65.6 +/- 8.0 min for controls). Furthermore, in vitro FAA was seen to induce tumor necrosis factor secretion from murine peritoneal cells and splenocytes. These studies suggest that FAA-induced tumor vascular shutdown in the colon 26 tumor is mediated by tumor necrosis factor.

Animals↗
PubMed1990

Effect of sulfonation on the cell and tissue distribution of the photosensitizer aluminum phthalocyanine.

Aluminum sulfonated phthalocyanine has potential as a suitable photosensitizer for use in the photodynamic therapy of cancer. In the present study, cellular uptake and retention of the individual mono-, di-, tri-, and tetrasulfonated derivatives (AlS1-4Pc) were examined in tissue culture and in normal and neoplastic tissue of tumor-bearing mice. Uptake and retention of the various derivatives by cells in tissue culture correlated inversely with the degree of sulfonation. Accordingly, Colo 26 cells in monolayer culture, 24 h after addition of 10 microM of appropriate photosensitizer, had accumulated approximately 25-fold more AlS1Pc than AlS3Pc and retained this species longer than more sulfonated derivatives. In contrast to these in vitro results, it was found that Colo 26 growing s.c. in BALB/c mice accumulated photosensitizer to a greater extent when the degree of sulfonation increased, such that A1S4Pc greater than AlS3Pc greater than AlS2Pc greater than AlS1Pc. By 24-48 h after the i.v. injection of 0.1 ml 2.27 mM solution of individual photosensitizer, the relative ratios of tumor:adjacent tissue varied from greater than 10:1 to greater than 2:1, showing that selective tumor uptake may be affected profoundly by the composition of the phthalocyanine compound. The livers and spleens of both normal and tumor-bearing mice, unlike other normal tissue, took up the sulfonated derivatives in an order that provided a mirror image of that observed in neoplastic tissue. These complex in vivo distribution and retention characteristics appear to be a consequence of relative hydrophilicity/hydrophobicity properties of the sulfonated species and indicate the extent to which these characteristics may influence photosensitizer distribution and accumulation.

Animals↗
PubMed1990

Protein kinase C-induced stimulation or inhibition of cellular proliferation in a murine macrophage tumor cell line.

M5076, a tumorigenic murine macrophage cell line, demonstrates diverse proliferative responses to a panel of protein kinase C activators. Thus, the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate and mezerein potently inhibit cellular proliferation (by greater than 90%), whereas the diacylglycerol analogue 1-oleoyl-2-acetyl-sn-glycerol markedly stimulates proliferation of serum-starved, quiescent M5076 cells. Another DG analogue, 1,2-dioctanoyl-sn-glycerol, has no effect on growth. However, all of these agents induce expression of c-fos oncogene mRNA levels to a similar magnitude and activate protein kinase C as determined by Mr 80,000 phosphorylation. Levels of c-fos protein induced by these treatments were markedly different with the antiproliferative agents producing greater c-fos protein levels.

Animals↗
PubMed1990

Selection and preliminary characterization of variant lines of a murine macrophage tumor resistant to the antiproliferative effects of phorbol esters.

Treatment of M5076 wild-type cells with 50 ng/ml of 12-O-tetradecanoylphorbol-13-acetate (TPA) almost completely inhibited cellular proliferation. Continuous culture in the presence of TPA was used to derive four lines, one polyclonal (TPAR) and three clonally derived (TPAR-1, -2, and -3), which exhibited variable resistance to the antiproliferative effects of phorbol esters. Protein kinase C (PKC) activation and c-fos expression in wild-type cells and the stably resistant line (TPAR-3) were examined after phorbol ester treatment. Both lines exhibited a comparable rapid and transient induction of c-fos mRNA expression, but induction of c-fos protein was reduced markedly in the TPAR-3 cells. Similarly in both cell lines, prolonged culture in phorbol ester produced down-regulation of PKC, as measured by inducible Mr 80,000 phosphorylation and an in vitro PKC assay. This decrease in PKC levels was paralleled by a decrease in c-fos mRNA and protein induction. Thus, c-fos expression in both wild-type and TPAR-3 cells is a consequence of PKC activation, and the development of resistance to TPA-antiproliferative effects in the TPAR-3 cell line was not linked causally to alterations in PKC levels or the c-fos mRNA induction response. The malignant capacity of the TPAR line was not reduced relative to wild-type cells. PKC activation and c-fos mRNA expression do not appear to determine changes in the in vivo or in vitro growth behavior of M5076 cells, whereas variations in c-fos protein expression may determine the anti-proliferative response to tumor-promoting phorbol esters.

Animals↗
PubMed1990

Protein kinase C levels and protein phosphorylation associated with inhibition of proliferation in a murine macrophage tumor.

Treatment of M5076 tumor cells with the phorbol estes 12-O-tetradecanoylphorbol 13-acetate (TPA) and phorbol 12,13 dibutyrate (PdBu) inhibited cellular proliferation, whereas 1,2-dioctanoyl-glycerol (DiC8) and 1-oleoyl2-acetyl-glycerol (OAG) did not affect cell growth. Inhibition of cellular proliferation in this cell line appears to be a consequence of protein kinase C (PKC) down-regulation since phorbol esters, but not a single application of diacylglycerols (DGs) down-regulated cellular PKC levels. By repeated application of DGs, PKC down-regulation was achieved and correlated with inhibition of proliferation. Phorbol ester-induced PKC down-regulation was reversible, upon removal of the phorbol ester, and the reappearance of PKC was associated with resumption of proliferation. The mitogenic responsiveness of these cells to added serum depended upon cellular PKC levels. Phorbol esters also caused the phosphorylation of two proteins which were not phosphorylated in response to DG treatment. Inhibition of growth of M5076 cells appears to be associated with phosphorylation of two novel proteins and/or PKC down-regulation.

Animals↗
PubMed1990

High frequency of double drug resistance in the B16 melanoma cell line.

Methotrexate (MTX) and N-(phosphonacetyl)-L-aspartate (PALA) are two agents to which cellular resistance can be conferred by gene amplification, but they do not generally show cross resistance. However, combined treatment with these two agents produced drug resistant cells in the B16 melanoma cell line at a much higher frequency than would be expected if resistance to the two agents was totally independent. An isolated doubly resistant clone, B16-F1 MP, showed a high frequency of resistance to pyrazofurin and ouabain, which are also agents to which resistance can be conferred by gene amplification. Thus MTX combined with PALA selected cells with an 'amplificator' phenotype (an increased ability to amplify parts of the genome). These B16-F1 MP cells had a decreased ability to form experimental lung metastases compared with the parent line but this difference was not found in baby hamster kidney cells with the amplificator phenotype. The mechanism underlying drug resistance may need to be considered when designing combination chemotherapy.

Amides↗
PubMed1990

Inhibition of [125I]epidermal growth factor binding to murine fibroblasts and keratinocytes by irradiation with UV-B light. Evidence for a protein kinase C-independent mechanism.

Treatment of murine Swiss 3T3 fibroblasts and XB/2 keratinocytes with UV-B light (302 nm) resulted in a dose-dependent inhibition of [125I]epidermal growth factor (EGF) binding. The light dose required to achieve 50% inhibition of binding in both cell types was 80-85 J/m2. Decreased [125I]platelet-derived growth factor binding was not evoked even by light doses of up to 280 J/m2. UV-B irradiation did not stimulate phosphorylation of the 80 kd protein substrate for protein kinase C. Furthermore, its effect on [125I]EGF binding was not altered as a consequence of protein kinase C down-regulation following prolonged exposure of cells to phorbol esters. These results indicate that UV-B-induced transmodulation of the epidermal growth factor receptor is a specific event mediated through a protein kinase C-independent pathway. Transfer of culture medium from irradiated cells to untreated control cells showed this effect was not induced as a result of transforming growth factor alpha release and subsequent binding to the EGF receptor in these cells.

Animals↗
PubMed1990

Metastasis and angiogenesis.

The relationship of angiogenesis, the growth of new blood vessels, to the process of tumour metastasis is examined. While the occurrence of angiogenesis generally is a necessary prerequisite for the formation of secondary tumours its presence is not a guarantee that cancer dissemination will occur. Novel approaches to antimetastatic therapy, using the newly formed vascular bed of tumours as the target, have produced successful results in experimental animals though such approaches have yet to be undertaken in the clinic.

Angiogenesis Inducing Agents↗