Biomedical subjects

H Akedo

Publications and source records attributed to H Akedo.

At least 55 records · Page 3Linked to original sources

PubMed1995

Cell-cycle-dependent invasion in vitro by rat ascites hepatoma cells.

The relationship between cell cycle and experimental metastasis of tumor cells in vivo has been investigated, but it remains to be elucidate which step of metastasis, or whether tumor-cell invasion in particular, depends on cell cycle. We previously reported an in vitro cell-monolayer invasion (transcellular migration) assay system, in which the invasive capacity of tumor cells is measured by counting tumor cells penetrating beneath a cultured mesothelial cell monolayer after tumor-cell seeding. Using our invasion assay system, the relationship between invasive capacity and cell-cycle distribution of MMI cells, a highly invasive clone of rat ascites hepatoma AHI30, was investigated. Invasive capacity of aphidicolin- or hydroxyurea-synchronized tumor cells enriched in G1/S-early S-phase cells was about 2 to 6 times higher than that of asynchronous cells. According to time-course experiments to examine the relationship between invasive capacity and the size of fraction of cells in each phase after release from an aphidicolin or a nocodazole block, it was suggested that MMI cells are most invasive in G1/S-S phase. Phagokinetic assay using colloidal gold particles showed that one possible reason for the enhanced invasiveness might be the increased cell motility in such phases, as suggested by the in vitro invasion assay.

Animals↗

PubMed1995

Enhancement of in vitro tumor-cell transcellular migration by tumor-cell-secreted endothelial-cell-retraction factor.

To investigate the factors affecting endothelial-cell retraction, we have studied the interaction of tumor cells with endothelial cells in 2 human pancreatic cancer cell lines, PSN-1 and MiaPaca-2. The extent of endothelial-cell retraction measured by the amount of intercellular junctional transport of FITC-dextran through an endothelial monolayer was increased by the addition of a conditioned medium (CM) from both cell lines, while CM from PSN-1 cells was 2 to 3 times more potent than that from MiaPaca-2 cells. After the treatment of endothelial monolayer with CM of PSN-1 cells, the ability of both PSN-1 cells and MiaPaca cells to adhere to or invade the monolayer increased. The addition of CM from PSN-1 cells did not affect the growth rate of either the endothelial or the tumor cells. The activity in the CM was heat-stable and bound to heparin-Sepharose, but was inactivated when treated by 0.5% trypsin. Protease inhibitors did not influence the activity. Pre-treatment of PSN-1 cells by an inhibitor of protein synthesis, cycloheximide, or of protein processing, benzyl-N-acetyl-alpha-D-galactosaminide, reduced endothelial-cell-retraction activity in the CM. The active substance in the CM fractionated in the molecular-weight range of 10,000 to 50,000. These results suggest that PSN-1 cells produce and secrete (a) soluble factor(s) that can induce endothelial-cell retraction, thus facilitating tumor-cell invasion.

Animals↗

PubMed1995

Participation of rhop21 in serum-dependent invasion by rat ascites hepatoma cells.

Rat ascites hepatoma cells (MM1 cells) penetrate through a cultured mesothelial cell monolayer (MCL) in the presence of fetal calf serum (FCS), but scarcely do so in its absence. Inactivation of rhop21 of MM1 cells by ADP-ribosyltransferase C3 resulted in the suppression of this serum effect on the penetration, suggesting that the serum effect was mediated by rhop21. To ascertain this assumption MM1 cells were transfected with an activated (Val14) human rhoA cDNA (Neo/RhoA 1-7). The transfectants penetrated MCL extensively even in the absence of FCS and became largely independent of serum for the penetration. These results suggest that serum-induced invasion by MM1 cells is mainly mediated by rhop21.

ADP Ribose Transferases↗

PubMed1995

Increased experimental pulmonary metastasis in pregnant mice.

The effect of pregnancy on experimental pulmonary metastasis was studied. Compared to the incidence of pulmonary metastasis induced by G6 cells in non-pregnant mice, the incidence of such metastasis was found to be greatly enhanced when the cells were injected i.v. in the latter half of pregnancy. The maximum enhancement was seen on the 15th day of pregnancy. The incidence of pulmonary metastasis returned to the level observed in non-pregnant mice when the cells were injected 4 days after parturition. Pregnancy also significantly increased the incidence of pulmonary metastasis of 2 other cell lines (3LL and Colon 26). Injection of G6 cells after hysterectomy performed on the 15th day of pregnancy resulted in decreased lung colonization, similar to that seen after parturition. Quantificative analysis of the arrest of G6 cells labeled with [125I]-5-iodo-2'-deoxyuridine in the lungs showed that the tumor-cell clearance from the lungs during the 24-72 hr after tumor-cell injection was much slower in pregnant than in non-pregnant mice. The continuous administration of beta-estradiol and/or progesterone, which maintained serum levels of the hormones equivalent to those prevailing on the 15th day of pregnancy, did not affect the lung colonization of G6 cells. Tumor-cell-platelet aggregation was more extensive with platelets obtained from mice at the 15th day of pregnancy than with those from non-pregnant mice. When platelets isolated from pregnant mice were injected into normal mice 5 min before G6 injection, lung metastasis was also enhanced. These findings suggest that a pregnant host is handicapped with regard to pulmonary metastasis, this being partly due to increased platelet-aggregating activity in response to tumor cells.

Adenocarcinoma↗

PubMed1995

Synergistic effects of basic fibroblast growth factor and hematopoietic growth factors on colony formation of K562 human leukemic cells.

The effect of basic fibroblast growth factor (bFGF) alone and in combination with other hematopoietic growth factors on the colony formation of K562 human leukemic cells was studied using soft agar colony assay. bFGF was found to have a weak colony-stimulating activity on K562 cells derived from the blastic crisis cells of human chronic myelogenous leukemia and to potentiate the K562 cell colony-stimulating activity of granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3) and stem cell factor (SCF) at a low concentration of below 1 ng/ml. These findings suggested that bFGF stimulates the growth of human leukemic cells directly in vivo alone and in synergy with other hematopoietic growth factors.

Blast Crisis↗

PubMed1995

Antiangiogenic substance(s) in a tumor cell line with low metastatic potential originating from the BALB/c mouse liver.

Two cell lines were established from liver cells (BALB/c mouse) exposed to benzo(a)pyrene: one was highly metastatic (G-5) and the other poorly metastatic (G-1) to the lung when subcutaneously implanted. However, there was no difference in lung colonization between G-1 and G-5 cells when they were intravenously injected. When G-1 cells were subcutaneously inoculated on one side of the back of mice followed by a challenge on the other side with G-5 cells 10 days later, the growth of the latter tumor was inhibited and the number of metastatic nodules in the lung was reduced. The functional vascular volume of G-1 tumor was less than the G-5 one. In mice bearing G-1 tumors, the neovascularization of intradermally inoculated G-5 cells was reduced. The conditioned medium from G-1 culture contained an inhibitory activity on the growth of endothelial cells from calf pulmonary artery. The inhibitory substance(s) was heat-stable, trichloroacetic acid-soluble, nondialyzable and resistant to various proteinases. The present results imply that G-1 cells produce an antiangiogenic substance(s), probably a polysaccharide(s), which inhibits the angiogenesis required for growth and metastasis of the G-5 tumor.

Animals↗

PubMed1994

Effects of NG-nitro-L-arginine and/or L-arginine on experimental pulmonary metastasis in mice.

Effects of NG-nitro-L-arginine methyl ester (L-NAME; an inhibitor of nitric oxide (NO) synthase) and/or L-arginine (substrate of NO synthase) on pulmonary metastasis of murine melanoma and Lewis lung carcinoma cells were investigated. L-NAME, L-arginine or both L-NAME and L-arginine was injected i.p. into mice 5, 3, and 1 h before and 1, 3, 5, and 7 h after the injection of tumor cells into mice via a tail vein. The administration of L-NAME (9.3 mumol/mouse) alone or L-arginine alone (46.5 or 186 mumol/mouse) potentiated pulmonary metastasis of highly and poorly metastatic B16 melanoma cells. L-NAME alone also increased the number of pulmonary metastasis of Lewis lung carcinoma cells, but L-arginine (185 mumol/mouse) did not. However, the combination of L-NAME and L-arginine increased the number of pulmonary metastasis of both the melanoma and Lewis lung carcinoma cells synergistically. L-NAME or L-arginine administration enhanced the retention of B16 melanoma cells in the lungs examined 24 h after the tumor cell injection. Synergistic effect of L-NAME and L-arginine was also seen in the tumor cell retention. The present results suggest that the metastatic potentials of the tumor cells do not simply correlate to NO production in vivo.

Animals↗

PubMed1994

Effect of dibutyryl cyclic AMP on morphologic features and marker production of human cervical argyrophil small-cell carcinoma cell line.

The effect of dibutyryl cyclic AMP (dB-cAMP) on the morphologic features and marker production of a human cervical argyrophil small-cell carcinoma (ASCC) cell line was examined. Following 1-5 days' exposure to 5 mM dB-cAMP, morphologic differentiation as defined by the expression of cytoplasmic processes (stellate cells) was observed. The number of stellate cells depended on the dose of dB-cAMP and incubation time. Shortly after removal of dB-cAMP from the culture medium, the treated cells returned to their original spherical shape. dB-cAMP caused a reduction in the growth rate of cells which recovered after removal of the agent. The morphological changes appeared not to be the result of growth inhibition by dB-cAMP, because the cells maintained in a serum-free medium did not show any change in shape. Electron microscopic study revealed the development of intracytoplasmic microtubules, microfilaments, and an increase in the number of neurosecretory granules in the treated cells. The levels of neuron-specific enolase, serotonin and gastrin in treated cells were significantly higher than those in untreated controls. These findings indicate that a reversible differentiation of cultured ASCC cells into neuroendocrine cells occurs in a growth medium containing dB-cAMP.

Biomarkers, Tumor↗

PubMed1994

Inhibitory effects of tumor invasion-inhibiting factor 2 and its conjugate on disseminating tumor cells.

Tumor invasion-inhibiting factor 2 (IIF-2) is a polypeptide of 21 amino acids which binds to the surface of tumor cells and inhibits experimental invasion in vitro. An albumin conjugate of IIF-2 was used to examine its potential as an antimetastatic compound. The conjugate inhibits in vivo lung metastasis of various highly metastatic tumor cells, including murine melanoma, colon adenocarcinoma, squamous cell carcinoma, forestomach carcinoma, and human fibrosarcoma. In addition to the anti-lung metastasis activity of this compound, it also showed the inhibitory effects on liver and spleen metastasis of murine T-lymphoma cells. A single administration of the conjugate with melanoma cells resulted in prolonged survival times, and their lung colonization was also inhibited when the conjugate was administrated i.v. at times ranging from 6 h before to 1 h after tumor cell inoculation. Similarly, i.p. administration 1 h prior to melanoma cell injection suppressed lung colonization. Pharmacokinetic analysis revealed that the conjugate was more stable than IIF-2 peptide alone. Approximately 10% of the conjugate remained circulating 2 h postinjection and persisted 20 h without degradation, compared with rapid clearing of the unconjugated IIF-2 peptide within 5 min. Furthermore, spontaneous lung metastasis of murine melanoma and colon adenocarcinoma cell was inhibited by successive i.p. administration of the conjugate before the removal of the primary site, with no effect on primary tumor growth. The conjugate significantly reduced tumor cell arrest in the lung and both the IIF-2 peptide and its conjugate demonstrated potent inhibition of basal as well as cytokine-induced-stimulated tumor cell motility. These results suggest that one mode of IIF-2 action may be inhibition of the extravasation of metastasizing cells which have arrested in a target organ, and that the IIF-2-albumin conjugate may be a potent antimetastatic substance with utility in the prevention of artificial seeding of tumor cells during surgical removal of the primary lesions as well as inhibiting metastasis from established metastases.

Albumins↗

PubMed1994

Distribution of C3-step regulatory proteins of the complement system, CD35 (CR1), CD46 (MCP), and CD55 (DAF), in hematological malignancies.

The distribution and levels of three membrane proteins, CD35, CD46, and CD55, which serve as complement regulators, were examined in normal peripheral blood and hematologically malignant cells. CD35 was negative in most leukemia cells regardless of the type of leukemia, although granulocytes, monocytes, and some populations of lymphocytes were CD35+. CD46 was present in all blood cells except erythrocytes, and levels were 2-8 times higher in most leukemia cells than in their mature counterparts, particularly in CML and CLL cells, except for those of B cell lineage. CD55, a widely-distributed phosphatidyl inositol-anchored protein, was more frequently lost in NHL cells than in other types of hematological malignancies. In this review, we discuss the roles, mechanisms, and clinical applications of cell-associated complement regulatory proteins in hematological malignancies.

Antigens, CD↗

PubMed1994

An experimental model for ovarian tumor invasion of cultured mesothelial cell monolayer.

BACKGROUND: Peritoneal spread of tumor cells is one of the characteristic features of biologic behavior of ovarian cancers. To understand the mechanism by which human tumor cell invasion takes place, we have tried to establish an in vitro experimental model for ovarian tumor cell invasion of the mesothelial cell monolayer. EXPERIMENTAL DESIGN: Mesothelial cells were isolated from normal rat mesentery by trypsin digestion and the cells (1 x 10(5)/dish) were cultured in Eagle's minimum essential medium supplemented with 10% fetal calf serum. Cultured mesothelial cells (M cells) grew forming a pavement-like monolayer. When M cells grew to a confluent state, tumor cells (1 x 10(5)/dish) were seeded on M cell monolayers and cultured. Four tumor cell lines derived from human ovarian cancers were tested for their invasive behaviors. The penetration of M cell monolayers by the tumor cells was confirmed by a perpendicular section of the cell layers. The number of penetrated single tumor cells and colonies/cm2 was counted under a phase contrast microscope after the tumor cell seeding. RESULTS: Several hours after the tumor cell seeding, the cells adhered to M cell monolayers and started to penetrate by extending pseudopodia-like cytoplasmic processes through junctional margins of neighboring M cells, resulting in the formation of penetrated single tumor cells that then proliferated to form colonies under the monolayer. The number of penetrated single tumor cells and colonies/cm2 increased up to 24 hours after the tumor cell seeding, and thereafter stayed almost constant. The number increased with the number of tumor cells seeded, when counted at 48 hours, and therefore was taken to be the number of tumor cells invaded. The in vitro invasiveness of tumor cells varied with the tumor cell lines examined. CONCLUSIONS: Application of this system appears to provide rapid determinations of the invasive potential of ovarian tumor cells and to make it easy to screen substances that modify the invasion of mesothelial cells.

Adenocarcinoma↗

PubMed1993

Induction of in vitro tumor cell invasion of cellular monolayers by lysophosphatidic acid or phospholipase D.

Serum is known to be required for invasion or phagokinesis of certain tumor cells, although the mechanism of its action is not well understood yet. In the in vitro invasion assay system we have developed, MM1 cells exhibiting extensive invasiveness against cultured mesothelial cell monolayers in the presence of 10% fetal calf serum did not invade them without serum. Human small cell lung cancer (OC10) cells also required serum in invasion. Serum could be completely substituted by oleoyl-lysophosphatidic acid (LPA) or bacterial phospholipase D (PLD). Phosphatidic acid also had an invasion-inducing activity, though to a lesser degree. Since human ovarian cancer (RMUG-S) cells require neither serum, LPA nor the PLD for invasion of mesothelial cell monolayers, the production of phospholipid like LPA, a candidate for intracellular second messenger, in tumor cells seems critical for the invasion by MM1 cells or OC10 cells. This result suggests a possible participation of particular signaling cascade, PLD-LPA(PA) system, in the invasion of certain tumor cells.

Animals↗

PubMed1993

A potent anti-metastatic activity of tumor invasion-inhibiting factor-2 and albumin conjugate.

Tumor invasion-inhibiting factor-2 (IIF-2) peptide was chemically conjugated to albumin with 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide hydrochloride. The molar ratio of the covalently linked IIF-2 peptide and albumin in the purified conjugate was 1.4 +/- 0.4, as determined with a radioactive peptide. The conjugate inhibited the invasion of HT1080 and B16FE7 cells in vitro at 40- to 60-fold lower concentrations than IIF-2 peptide. Scatchard analysis of binding data demonstrated that the IIF-2-albumin conjugate bound to HT1080 and B16FE7 cells with Kd values of 240 and 340 nM, respectively. The conjugate suppressed the lung colonization of B16FE7 cells more effectively than the IIF-2 peptide in an experimental metastasis. These results indicate that the covalent linkage of the IIF-2 peptide to a carrier macromolecule provides a structure that enables this peptide to exhibit increased inhibitory action in cancer cell invasion and metastasis, and that IIF-2 exerts its inhibitory action on invasion by binding to a specific binding site on the tumor cell surface.

Albumins↗

PubMed1993

Membrane cofactor protein (MCP, CD46) in seminal plasma and on spermatozoa in normal and "sterile" subjects.

A sperm protein of molecular mass 43 kDa (the spermatozoa membrane cofactor protein, smMCP) and a seminal plasma protein of 60 kDa (ssMCP) were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblotting with four monoclonal antibodies (mAb) against membrane cofactor protein (MCP, CD46). These proteins served as factor I cofactors for the cleavage of methylamine-treated C3 (C3ma), the activity of which was blocked by M75, an MCP cofactor-activity-blocking mAb. Thus, these semen proteins are antigenic and functional homologous of MCP. On SDS-PAGE analysis these MCP migrated as single-band proteins which differed from the two-band forms of MCP expressed on other cells. smMCP was N-glycosylated but not O-glycosylated, while ssMCP was O-glycosylated: after deglycosylation of these proteins bands were detected at 38-40 kDa and 43 kDa on SDS-PAGE, respectively. These semen MCP are therefore, structurally different from the conventional MCP. ssMCP in both normal and "sterile" subject groups was determined by sandwich enzyme-linked immunosorbent assay. Seminal plasma in the two groups contained 250-700 ng/ml ssMCP. The difference between the two groups was marginal, although samples from normal subjects tended to show higher concentrations of ssMCP than samples from "sterile" subjects. No molecular difference was observed with ssMCP and smMCP in the two groups by SDS-PAGE/immunoblotting analysis. Immunohistochemical analysis suggested that MCP was positive in glandular epithelial cells and the lumen of the prostate, and in most intra-lumen cells of the testis. Using antibody M177, solubilized prostate and testis were analyzed by immunoblotting and compared with other cell MCP. The major band of MCP in the testis, but not in the prostate, was of 60 kDa, which aligned with ssMCP. No band of testis or prostate MCP, however, aligned with smMCP. ssMCP may be produced in the testis, while the origin of smMCP remains unknown. We hypothesize that ssMCP is important in the survival of spermatozoa, protecting them against local secretion of immunoglobulin and complement in the female genital tract, and that smMCP, which is expressed on acrosome-reacted spermatozoa, plays an essential role in the interaction of spermatozoa with oocytes.

Antigens, CD↗

PubMed1993

Retraction of cultured endothelial cell monolayer by human breast cancer cells, MCF-7.

A human breast cancer cell line (MCF-7), when sealed on confluent bovine pulmonary aortic endothelial cell (CPAE) monolayers, induced morphological changes (retraction) in CPAE cells. The area of retraction depended on the incubation time and the number of MCF-7 cells, suggesting that MCF-7 cells had the capacity to retract CPAE cells. This capacity was reduced by 60% by pretreatment of MCF-7 cells with 17 beta-estradiol (E) and progesterone (Pg). The extent of retraction was not affected by the addition of various protease inhibitors. CPAE retraction was induced also by adding conditioned medium (CM) from the culture of MCF-7 cells. Considerably less activity was detected in the CM obtained from MCF-7 cells cultured in the presence of E and Pg. The retraction was reversed in 24 h by culturing the monolayer in fresh medium without CM and was not induced by trypsin treatment of the CM.

Breast Neoplasms↗

PubMed1993

Transforming growth factor(TGF)-beta 1 induces leukemic cell-growth-promoting activity in fibroblast cells.

The production of the leukemic cell-growth-promoting factor (LGF) in TGF-beta 1-treated fibroblast cells was studied. BALB/c3T3 mouse fibroblast (3T3) cells cultured in Eagle's medium containing a low concentration of TGF-beta 1 (0.04-1 ng/ml) secreted 3-5 times more LGF than the cells cultured in the absence of TGF-beta 1. The amount of LGF secretion was dose-dependent on the concentration of post-cultured medium and time-dependent after the addition of TGF-beta 1. Similar findings were obtained in human diploid fibroblasts, WI-38 cells. LGF is a 18KD glycoprotein that is acid-stable but heat-unstable.

3T3 Cells↗

PubMed1993

A monoclonal antibody against human decay-accelerating factor (DAF, CD55), D17, which lacks reactivity with semen-DAF.

Human decay-accelerating factor (DAF, CD55) is a phosphatidyl inositol-anchored glycoprotein consisting, from the N-terminus, of 4 short consensus repeats (SCR), a Ser/Thr (ST)-rich region providing O-glycosylation sites, and the membrane-anchoring unit. A mAb, named D17, was raised against purified erythrocyte-DAF. This mAb recognized DAF on blood cells and most cell lines as determined by flow cytometry and immunoblotting. Its reactivity was similar to but weaker than that of two other well-characterized mAbs to DAF, IA10 (seeing an epitope within SCR1) and 1C6 (seeing an epitope within SCR3). The reactivity of D17 with erythrocyte DAF became increased by treatment with sialidase/O-glycanase, suggesting that its epitope is located close to the O-glycosylation sites, probably within the ST-rich region or SCR4. D17 barely blocked the decay-accelerating activity of DAF. Using the three mAbs, tissue-associated and soluble forms of DAF were identified by SDS-PAGE/immunoblotting and immunohistochemical staining. IA10 and 1C6 recognized a 50 kDa protein in spermatozoa lysate and two proteins of Mr 70 and 55 kDa, respectively, in seminal fluid. These represented membrane-associated and soluble forms of DAF, which were neither recognized by mAb against membrane cofactor protein (MCP, CD46) and C3b/C4b receptor (CR1, CD35) nor by non-immune IgG. In contrast to IA10 and 1C6, D17 did not recognize either spermatozoa-DAF or seminal plasma-DAF, or the deglycosylated or untreated forms of them. Immunohistochemical analysis showed that testis was stained with IA10 but not with D17.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

[The evaluation of interval period to prepare the left ventricle for Jatene operation for simple transposition of the great arteries].

Laboratory studies indicate that it can be as short as one week after a first-stage operation to prepare the left ventricle for a subsequent second-stage arterial switch (Jatene operation) for simple transposition of the great arteries (TGA). In order to ascertain it, we evaluated the left ventricular (LV) function immediately after the first-stage procedure and late postoperatively by echocardiography and catheterization. There were 6 cases with simple TGA. The age at palliation was 6.6 +/- 4.5 (mean +/- SD) months, and the interval between the first-stage and Jatene operation was 16 days-16 months (6.7 +/- 6.4 months). All patients survived after arterial switch. In 2 early infant cases, arterial switch operation was performed with ventilatory support due to cardiac and respiratory failures 16 and 21 days after the first-stage operation respectively. LV end-diastolic dimension increased by 1 week and showed no change after 1 week (n = 4). LV ejection fraction was depressed from 0.84 +/- 0.03 preoperatively to 0.61 +/- 0.08 at 1-4 days after the first-stage operation but improved to 0.83 +/- 0.06 by 7-9 days after that (n = 4). LV end-diastolic posterior wall thickness increased to more than 4 mm in 3 infant cases, more than 3 mm in 2 early infant cases by 7 days. The results of catheterization after the first-stage operation were as follows (n = 5); left ventricular right ventricular peak systolic pressure ratio increased from 0.53 +/- 0.16 to 0.94 +/- 0.04 (p < 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation, Physiological↗