Growth Of Mouse Melanoma Cells Research Articles

Uncarboxylated Osteocalcin Induces Antitumor Immunity against Mouse Melanoma Cell Growth.

Because of the poor response to chemotherapy and radiation therapy, new treatment approaches by immune-based therapy involving activated T cells are required for melanoma. We previously reported that the uncarboxylated form of osteocalcin (GluOC), derived from osteoblasts, potentially suppresses human prostate cancer cell proliferation by direct suppression of cell growth. However, the mechanisms in vivo have not been elucidated. In this study, we found that GluOC suppressed tumor growth of B16 mouse melanoma transplants in C57Bl/6N wild-type mice. Our data demonstrated that GluOC suppressed cell growth by downregulating phosphorylation levels of receptor tyrosine kinases and inducing apoptosis in vitro. Additionally, stimulation of primary mouse splenocytes with concanavalin A, a polyclonal T-cell mitogen, in the presence of GluOC increased T cell proliferation and their interferon-γ production. Taken together, we demonstrate that GluOC exerts multiple antitumor effects not only in vitro, but also in vivo through cellular immunostimulatory effects against B16 mouse melanoma cells.

Low Molecular Weight Components in Aqueous Echinacea Purpurea Leaf Extract Inhibit Melanoma Cell Growth

Melanoma is a skin cancer associated with high mortality. The three year survival rate from advanced melanoma is between 10-15%. One reason for this high mortality rate is that melanoma cells are resistant to traditional chemotherapeutics. Echinacea is a plant genus native to North America with putative anticancer properties. Here, we examined effects of aqueous Echinacea purpurea leaf extract on the growth of melanoma cells and nontransformed fibroblasts. This aqueous extract reduced B16 mouse melanoma cell growth at concentrations that did not inhibit the growth of nontransformed NIH3T3 fibroblasts, suggesting that the extract had biological specificity against transformed cells. We also examined the effect of different fractions of the extract on melanoma cell growth. These data indicate that components less than 3 kD in size exhibited the greatest inhibitory action on melanoma cell growth. In addition, these data indicated that larger components in the extract ameliorate the ability of these low molecular weight compounds to inhibit melanoma cell growth. Furthermore, Echinacea extract inhibited the growth of v-Src transformed LA25 cells without reducing Src kinase activity. Taken together, these results suggest that aqueous Echinacea purpurea extract contains low molecular weight compounds that preferentially inhibit tumor cell growth in the face of oncogenic tyrosine kinase activity. These data suggest future studies to better define bioactive compounds in Echinacea purpurea and evaluate their therapeutic efficacy in vivo.

<i>In-Vitro</i> Inhibition of Human Melanoma (BLM) Cell Growth by Progesterone Receptor Antagonist RU-486 (Mifepristone)

RU-486 is an abortifacient which is used to terminate early pregnancy. It acts by blocking progesterone receptor. In our earlier study with progesterone, RU-486 was used as a progesterone receptor antagonist to find out the mechanism of progesterone action on melanoma cells. Results indicated that the effect of progesterone was not mediated through progesterone receptor. In the course of experiments, it was observed that RU-486 by itself inhibited mouse melanoma cell growth. Further research work with RU-486 showed a dose dependent inhibition of human melanoma cell growth. The mechanism of inhibition of cell growth was due to apoptosis and this effect of RU-486 was neither mediated through progesterone receptor nor glucocorticoid receptor. This in-vitro study suggested that melanoma also could be a target for RU-486 action, apart from breast, ovary and prostate cancers.

Challenge to the suppression of tumor growth by the β4-galactosyltransferase genes.

It has been well established that structural changes in glycans attached to proteins and lipids are associated with malignant transformation of cells. We focused on galactose residues among the sugars since they are involved in the galectin-mediated biology, and many carbohydrate antigens are frequently expressed on this sugar. We found changes in the expression of the β4-galactosyltransferase (β4GalT) 2 and 5 genes in cancer cells: decreased expression of the β4GalT2 gene and increased expression of the β4GalT5 gene. The growth of mouse melanoma cells showing enhanced expression of the β4GalT2 gene or reduced expression of the β4GalT5 gene is inhibited remarkably in syngeneic mice. Tumor growth inhibition is probably caused by the induction of apoptosis, inhibition of angiogenesis, and/or reduced MAPK signals. Direct transduction of human β4GalT2 cDNA together with the adenovirus vector into human hepatocellular carcinoma cells grown in SCID mice results in marked growth retardation of the tumors. β4GalT gene-transfer appears to be a potential tool for cancer therapy.

Differentiation-inducing and anti-proliferative activities of lupeol on canine melanoma cells.

Canine melanoma is the most common oral malignant tumor reported in the field of veterinary medicine. We found that lupeol, a lupine triterpene, inhibited mouse melanoma cell growth in vitro and in vivo by inducing cell differentiation. In the present study, we examined the differentiation-inducing activities of lupeol on 4 canine melanoma cells in vitro and in vivo.The induction of canine melanoma cell differentiation by lupeol was confirmed by evaluating some differentiation markers such as tyrosinase with real-time RT-PCR. Furthermore, we transplanted canine melanoma cells into a severe combined immunodeficiency mouse, and studied the anti-progressive effects of lupeol on tumor tissue.The gene expression of microphthalmia-associated transcription factor, tyrosinase, and tyrosinase-related protein-2, which are markers of pigment cell differentiation, was induced in 4 canine oral malignant melanoma cells by lupeol, and the agent markedly inhibited tumor progression in canine melanoma-bearing mice.

A Novel and Effective Cancer Immunotherapy Mouse Model Using Antigen-Specific B Cells Selected In Vitro

Immunotherapies such as adoptive transfer of T cells or natural killer cells, or monoclonal antibody (MoAb) treatment have recently been recognized as effective means to treat cancer patients. However, adoptive transfer of B cells or plasma cells producing tumor-specific antibodies has not been applied as a therapy because long-term culture and selective expansion of antigen-specific B cells has been technically very difficult. Here, we describe a novel cancer immunotherapy that uses B-cell adoptive transfer. We demonstrate that germinal-center-like B cells (iGB cells) induced in vitro from mouse naïve B cells become plasma cells and produce IgG antibodies for more than a month in the bone marrow of non-irradiated recipient mice. When transferred into mice, iGB cells producing antibody against a surrogate tumor antigen suppressed lung metastasis and growth of mouse melanoma cells expressing the same antigen and prolonged survival of the recipients. In addition, we have developed a novel culture system called FAIS to selectively expand antigen-specific iGB cells utilizing the fact that iGB cells are sensitive to Fas-induced cell death unless their antigen receptors are ligated by membrane-bound antigens. The selected iGB cells efficiently suppressed lung metastasis of melanoma cells in the adoptive immunotherapy model. As human blood B cells can be propagated as iGB cells using culture conditions similar to the mouse iGB cell cultures, our data suggest that it will be possible to treat cancer-bearing patients by the adoptive transfer of cancer-antigen-specific iGB cells selected in vitro. This new adoptive immunotherapy should be an alternative to the laborious development of MoAb drugs against cancers for which no effective treatments currently exist.

<i>In-Vitro</i> Effect of Sex Steroids on Mouse Melanoma (B16F10) Cell Growth

A battery of sex steroids were used to check their effect on mouse melanoma (B16F10) cell growth in-vitro. Progesterone and its synthetic receptor antagonist RU-486 showed maximum inhibition on in-vitro melanoma cell growth [1]. Further research work showed that the inhibition by progesterone was not a toxic, spurious or non-specific effect on mouse melanoma cell growth and the inhibition by progesterone was not mediated through progesterone receptor.

1,3-Dichloro-2-propanol induces apoptosis via both calcium and ROS in mouse melanoma cells

We demonstrated that the apoptotic cell death of mouse melanoma cells exposed to 1,3- dichloro-2-propanol (DCP) was Ca(2+) dependent. 1,3-DCP inhibited the growth of mouse melanoma cells in a concentration-dependent manner. Furthermore, a terminal nick-end labeling of fragmented DNA (TUNEL) assay demonstrated that 1,3-DCP-induced apoptosis. Moreover, reactive oxygen species and Ca(2+), generated by mouse melanoma cells that were exposed to 1,3-DCP, resulted in the activation of NF-jB and MAPKs. Finally, treatment with BAPTA, an intracellular Ca(2+) chelator, completely inhibited the apoptosis that was induced by 1,3-DCP treatment of B16F10 cells.

ID: 212 Effect of plasminogen activator inhibitor-1 on growth of mouse melanoma cells

ID: 212 Effect of plasminogen activator inhibitor-1 on growth of mouse melanoma cells

Inhibition of B16 Mouse Melanoma Cell Growth and Induction of Apoptotic Cell Death with 8‐Chloroadenosine‐3′,5′‐monophosphate and Tiazofurin

Novel antineoplastic agents, 8-chloroadenosine 3',5'-monophosphate (8-Cl-cAMP) and tiazofurin (TR), have been shown to be effective against different malignant cells. Through specific mechanisms of action they modulate the cellular signal transduction pathway, thereby causing growth inhibition, cell differentiation, and apoptosis. The aim of this work was the in vitro study of either 8-Cl-cAMP or TR effects on B16/F10 and B16/C3 mouse melanoma cell growth and cell death. Significant cell growth inhibition was obtained after the application of 8-Cl-cAMP or TR. The presence and number of apoptotic cells was evaluated using agarose gel electrophoresis and flow cytometry. The number of apoptotic nuclei, after treatment with antineoplastic agents, did not significantly change in B16/F10 cells, although it did show a significant increase in B16/C3 cells. The expression of c-myc did not significantly change in B16/F10 cells after treatment with 8-Cl-cAMP or TR. The same results were obtained in B16/C3 cells after treatment with 8-Cl-cAMP. The level of c-myc expression showed a significant increase in B16/C3 cells after treatment with TR. Concerning the effects that the analyzed agents exhibited on melanoma cells and other cancer cells, further preclinical studies of these drugs will potentially lead to better understanding of the molecular mechanisms of their action and finally more efficient therapeutic approaches to malignant diseases.

6-deoxy-6-bromo-ascorbic acid inhibits growth of mouse melanoma cells

The influence of 6-deoxy-6-bromo-ascorbic acid (6-Br-AA) and ascorbic acid (AA) on the growth of mouse melanoma cell line B16, mouse fibroblasts (L929), and human cervical carcinoma HeLa cells was examined by in vitro experiments. 6-Br-AA was added in concentrations 10(-1) to 10(-8)M and for incubation periods of 2, 18, 24 and 72 h. The present results indicate that 6-Br-AA exhibits a highly pronounced inhibiting effect on growth and DNA synthesis of melanoma cells. Inhibitory effect of 6-Br-AA was confirmed by the in vivo experiments. The tumor-suppressing effect on solid melanoma B16 was attained with 9 mg per mouse given three times daily for 16 days. Our preliminary results indicate that 6-Br-AA could serve as a potential antitumor agent.

The growth of mouse melanoma cells in hormone-supplemented, serum-free medium

A mouse melanoma line, M2R, derived from the B 16 transplantable tumor can be grown in a hormone-supplemented serum-free medium. Cells grown in medium supplemented with insulin, transferrin, testosterone (or progesterone), follicle-stimulating hormone, nerve growth factor (NGF) and leutinizing releasing hormone show growth rates equivalent to that seen in medium supplemented with 5% fetal calf serum (FCS). This hormone-supplemented medium will support the growth of cells indefinitely. In cells grown in hormone-supplemented medium, insulin is shown to increase incorporation of glucose into glycogen and fatty acids. Transferrin is shown to act in part as an iron transport protein, although another role in growth stimulation cannot be eliminated. It is suggested that progesterone stimulates growth via an androgenic breakdown product and thus acts in a manner similar to testosterone.

The role of adenosine 3′,5′‐monophosphate in the transformation of cloudman mouse melanoma cells

The role of adenosine 3',5'-monophosphate (cyclic AMP) in the regulation of mouse melanoma cell growth and differentiation was investigated. A variant melanoma (Cloudman S91-F) which displays a greater degree of transformation than the parental cell (Cloudman S91) was isolated. A correlation between cyclic AMP metabolism and transformation was made. Dibutyryl cyclic AMP depressed cell growth and increased pigmentation in both parental and variant cell lines. The parental cell line, however, was more responsive to melanocyte-stimulating hormone (MSH) which was found to affect cell growth and pigmentation by increasing cyclic AMP levels. The more transformed S91-F cell line contained lower levels of cyclic AMP than the parental cell line, and this fact correlated well with the higher degree of growth and lesser degree of pigmentation in the variant. Enzymatic analysis revealed that the hydrolysis of cyclic AMP in both cell lines was similar, while the adenylate cyclase activity of the variant cell line was lower than that of the parental cell line. Lineweaver-Burk plots demonstrated that the Km's for the enzymes in the two cell lines were the same but that the Vmax of the S91-F cell line was significantly less that that of the S91 cell line. Thus, the lesion in the S91-F cell which is responsible for its more transformed characteristics seems to be one which affects adenylate cyclase at the level of the cell membrane.