Sarcoma-180 Cells Research Articles

Immunological studies on sporamycin-treated animals.

Sporamycin showed a remarkable tumor regressive activity against sarcoma-180 with a single 5 mg/kg dose of intravenous administration. This antitumor effect on tumor and host animals was examined immunologically. As the results: (1) When sarcoma-180 tumor cells were used as an antigen macrophage migration inhibition reaction by spleen cells derived from the tumor-bearing mice treated with sporamycin was positive at day 7 approximately 14 after the medication and was negative thereafter. (2) The delayed hypersensitivity tested by the foot-pad reaction was positive in tumor-bearing mice treated with sporamycin, and no decrease of foot pad reaction was observed, whereas this reaction decreased remarkably in non-treated tumor-bearing mice. (3) Sarcoma-180 tumor cells were mixed with spleen cells derived from sporamycin-treated mice, and were inoculated into normal dd mice. The growth of tumor cells was inhibited markedly, but no inhibition of tumor growth was observed in case of spleen cells derived from non-treated tumor bearing mice. (4) Combined treatment of sporamycin with PS-K, an immunopotentiator, showed a remarkable synergistic effect.

Studies on antitumor activity of wood hemicelluloses. I. Antitumor effect of 4-O-methylglucuronoxylan on solid tumor in mice.

The present studies were undertaken to evaluate antitumor activities of 4-o-methylglucuronoxylan isolated from beech wood (Fagus clenata Blume), based on the inhibition of tumor growth of Sarcoma-180, Ehrlich carcinoma and L1210 leukemic cells in mice. The growth rate of Sarcoma–180 subcutaneously implanted into ICR mice was significantly inhibited by intraperitoneal administrations of 4-o-methylglucuronoxylan and the most significant effect was found at doses ranging 200 mg/kg. On the other hand, such a treatment failed to suppress the growth of ascites form of L1210 leukemic cells arising BDF1 mice. Attempts to induce prolongation of life span by intralesional injection into L1210 solid tumor itself were successful. The xylan studied here has a main chain of (1–4)β-d-xylopyranosyl unit with every nine or ten units carrying (1–2) α-4-O-methylglucuronic acid.For comparison, several common wood hemicelluloses were tested. These involved O-acetylgalactoglucomannan, arabino-4-O-methylglucuronoxylan, arabinog...

Antitumor activity of hot-water extract from delipidated BCG.

The antitumor activity of hot-water extract of delipidated BCG was investigated in mice inoculated with Sarcoma-180 cells and Ehrlich carcinoma cells, respectively. The hot-water extract was found to be effective when administered after and ineffective when administered before the inoculation of tumor cells. When this extract was given with anticancer drugs, such as Mitomycin C and cyclophosphamide, a combined effect was obtained in the treatment of Sarcoma-180 and of Ehrlich carcinoma.

Further evidence in support of cell-surface-associated deoxyribonucleic acid in tumor cells: an autoradiographic study.

Ascites sarcoma-180 cells, when stained with platinum-pyrimidine complexes as the sole electron dense stain, show distinct dense patches to granular appearance on the surface of the plasma membrane which has been suggested to be attributable to deoxyribonucleic acid. Swiss Webster mice, 4-5 weeks of age, weighing 24-26 g with 4 X 10(6) ascites sarcoma-180 cells when injected with 3 X 7.0 micronC of tritiated thymidine on day 5 of the tumor implant, show specific labeling on the plasma membrane surface. The photopositive silver grain distribution in both the light and electron microscope autoradiograms when followed from the nucleus outwards show a distinct peak over the nucleus and the plasma membrane. The quantity and origin and role of this surface-associated deoxyribonucleic acid is not clear.

Antitumor activity of a Brucella abortus preparation

Mice injected intraperitoneally with sarcoma-180 cells develop ascites and eventually die. Intraperitoneal injection of a nonviable, aqueous... ether-extracted Brucella abortus preparation (BRU-PEL) as early as 7 days before or as late as 7 days after injection of tumor cells significantly inhibited development of ascites and protected against death. BRU-PEL was not effective if injected after ascites was grossly apparent. BRU-PEL was significantly more active than a Corynebacterium parvum preparation administered in the same way.

Cell-surface-associated nucleic acid in tumorigenic cells made visible with platinum-pyrimidine complexes by electron microscopy.

Platinum-pyrimidine complexes have been used as the sole electron-dense stains for electron microscopy on 18 types of mammalian cells. These recently discovered complexes react avidly with nucleic acids in vitro, are antitumor agents, and are highly soluble in water. In cells, they are selective for regions rich in nucleic acid and make visible the cellular chromatin, nucleolus, and ribosomes. In addition, cells that are tumorigenic exhibit electron-dense patches at the plasma membrane; normal cells do not. The results of treatment of ascites sarcoma-180 cells with concanavalin A, nucleases, glycosidases, or a protease suggest that the electron-dense patches are DNA, with neuroaminic acid involved in the surface binding.

Effect of antitumor agents on sarcoma-180 tumor cells transplanted to liver, kidney, and lung.

The survival time of animals, inhibition of the incorporation of thymidine-[6-3H] (3H-TdR) into DNA, and histopathological observation were made after the injection of Mitomycin-C, Bleomycin, cyclophosphamide, Daunomycin, Actinomycin-D, or 5-fluorouracil into mice transplanted with sarcoma-180 to their liver, kidney, and lung. The most prolonged survival time was obtained by the injection with cyclophosphamide and a moderate prolonged survival by Bleomycin and Actinomycin-D. In the case of 5-fluorouracil and Daunomycin, there were extreme variations in the survival time depending on the site of tumor growth. Cyclophosphamide and 5-fluorouracil showed greater and longer lasting inhibition of the incorporation of 3H-TdR into DNA of the tumor tissue, whereas the remaining agents caused transient inhibition on the tumor tissue. Inhibitory ratio and duration of the incorporation of 3H-TdR into DNA of normal site of the tissue of tumor-bearing organ were found to be more increased or almost the same compared with those of the tumor tissue. The most rapid recovery of the incorporation of 3H-TdR into DNA was observed in the small intestine among various organs and tumor in any treatment groups. From the histopathological observation, the degree of tumor cell damage by the agent was almost in agreement with inhibition of the incorporation of 3H-TdR up to 72 hr after the treatment.

Platinum Compounds as Stains for Electron Microscopy

Uranyl acetate has been used as an electron stain for en bloc staining as well as for staining ultrathin sections in conjunction with various lead stains (Fig. 1). Present studies reveal that various platinum compounds also show promise as electron stains. Certain platinum compounds have been shown to be effective anti-tumor agents. Of particular interest are the compounds with either uracil or thymine as one of the ligands (cis-Pt(II)-uracil; cis-Pt(II)-thymine). These compounds are amorphous, highly soluble in water and often exhibit an intense blue coloration. These compounds show enough electron density to be used as stains for electron microscopy. Most of the studies are based on various cell lines (human AV, cells, human lymphoma cells, KB cells, Sarcoma-180 ascites cells, chick fibroblasts and HeLa cells) while studies on tissue blocks are in progress.

Asparagine metabolism in mouse sarcoma cells I. Transport, metabolism and pools of dicarboxylic amino acids and their amides

In order to grow in vitro, mouse Sarcoma-180 (S-180) cells require glutamine (2 mM) or glutamate (10 mM). A subline (S-180/ASN) was developed that can also grow in the absence of glutamine and glutamate if supplied either with asparagine (2 mM), aspartate (30 mM) or oxaloacetate (10 mM). The purpose of this study was to elucidate the cellular and metabolic differences between S-180 and S-180/ASN cells. In both cells the transport velocity of amino acids decreased in the order: glutamine≈asparagine ⪢ glutamate≈aspartate. With the exception of aspartate, the transport proceses were saturable and led to intracellular accumulation of the free amino acids. The energy of activation for the transport of asparagine by S-180 cells, which did not metabolize it, was 9.6 kcals/mole, that for aspartate 2.6 kcals/mole. The parent cells accumulated large amounts of unaltered asparagine but incorporated very little into acid-insoluble material. In the subline the conversion of asparagine to aspartate and glutamate was rapid and was directly responsible for the ability of these cells to grow on asparagine (in the absence of glutamine). The subline also differed from S-180 cells by transporting glutamate slower but metabolizing it more rapidly. The intracellular pools of amides never exceeded 3.2 mM while the corresponding acids reached up to 26 mM in cell water.

The enzymic phosphorylation of N6-(delta 2-isopentenyl) adenosine in chick liver homogenates.

Enzyme preparations from chick kidney and liver rapidly phosphorylate the modified nucleoside, N6-(Δ2-isopentenyl) adenosine. The resulting metabolite behaves electrophoretically as a mononucleotide and is quantitatively hydrolyzed to the nucleoside by 5′-nucleotidase. The isopentenyl side chain is not altered in the phosphorylated derivative. It is concluded that the metabolite is N6-(Δ2-isopentenyl) adenosine-5′-monophosphate. The properties of the enzyme activity responsible for the phosphorylation resemble the properties of adenosine kinase isolated from sarcoma-180 cells, and from rabbit liver, with respect to pH optimum, Km value, and sensitivity to magnesium ion and ATP. Thus, it seems probable that the phosphorylation observed in the chick tissue preparations is due to adenosine kinase. The mononucleotide could not be further phosphorylated in vitro by adenylate kinase. This observation, coupled with the fact that isopentenyladenosine is biosynthesized at the macromolecular level, suggests that the 5′-monophosphate ester may be the form through which some of the biological effects of this modified nucleoside are manifested.A major end product of N6-(Δ2-isopentenyl) adenosine catabolism in chick liver and kidney homogenates is uric acid.

Regression of Sarcoma-180 after cis-Dichlorodiammineplatinum (II) : A Fine Structural Study

Sarcoma-180, a transplantable tumor in Swiss White mice regresses completely after a single intraperitoneal injection of cis-Dichlorodiammineplatinum (II). Fine structural studies before and after cis (Pt (NH3)2C12) treatment show certain morphological changes indicative of possible mechanisms involved in the tumor regression. The mitotic activity of tumor cells is inhibited soon after platinum injection possibly due to its interaction with DNA. Appearance of plasma cells (Fig. 1) and lymphocytes within the tumor mass after a 4-6 days period of platinum treatment indicates a possible enhancement of the immune system. The plasma cells display a well developed RER and a typical nuclear morphology. Further studies on spleen of both treated and control animals show an increase in the number of plasma cells after about 5 days of platinum treatment (Fig. 2).The increase in the lymphocytes and the plasma cells indicates an immune system at the cellular level as these antibody producing cells directly come in contact with the Sarcoma-180 cells. Also the double diffusion method on agar plates indicates the enhanced humoral antibody production against Sarcoma-180 as opposed to the controls.

Uptake and distribution of 4,4'-diacetyl-diphenyl-urea-bis-guanylhydrazone in sensitive and resistant sarcoma 180 cells in vitro

Sarcoma 180cells sensitive to 4,4'-diacetyl-diphenyl-urea-bis-guanylhydrazone (DDUG) were compared with a resistant subline (S-180/DDUG) with respect to crossresistance to methylglyoxal-bis-guanylhydrazone (CH 3G), 2-chloro-4',4″-bis(2-imida-zolin-2-yl)-terephthalanilide (NSC 38280) and vincristine (VCR); they were also compared with respect to the nature and rate of cellular uptake of DDUG and its intracellular distribution and binding. Resistance to DDUG was found to be associated with a reduced rate of uptake of DDUG while the intracellular distribution (65–70 per cent in nuclei and 30 per cent in mitochondria) and the capacity for lipid-associated drug binding was unchanged. The uptake was a rapid passive process leading to extensive intracellular accumulation; it was not affected by CH 3G or VCR, but was inhibited by NSC 38280 or Janus Green. S-180/DDUG cells were cross-resistant to NSC 38280 but sensitive to CH 3G and VCR, while S-180/VCR cells were cross-resistant to NSC 38280 but sensitive to DDUG and CH3G. In uptake, intracellular distribution and binding DDUG resembles NSC 38280. Combination and cross-resistance studies suggest a lack of similarity between DDUG, NSC 38280, VCR and CH 3G with respect to the site of growth inhibitory action.

Biological effects of anthramycin methyl ether. II. Inhibition of DNA synthesis in the intestinal epithelium and sarcoma-180 tumor cells.

The temporal course of DNA synthesis after subcutaneous injection on Anthramycin Methyl Ether (AME) was investigated in Sarcoma-180 ascites tumor cells and in the proliferative cells of the intestinal epithelium. Both cell types demonstrated dramatic inhibition of DNA synthesis: however, there was no appreciable difference in response of the 2 cell systems to the drug. After sub-lethal doses, recovery was seen to occur in an oscillatory fashion which eventually reached control values. This pattern of recovery suggested partial <i>in vivo </i>synchronization of the proliferative cell population, which could be due to preferential block in one stage of cell cycle or selective killing of cells in a sensitive stage.

Substances in sera influencing growth of sarcoma 180 in mice.

IN 1953 John Kidd 1 reported a series of experiments showing that subcutaneous mouse lymphomata of two types regressed promptly following intraperitoneal injection of normal guinea pig serum into the mice. The tumor grew normally in untreated controls and in mice injected with rabbit or horse serum. The guinea pig serum had no discernible effect on the tumor cells in vitro. This was the first report of a naturally occurring substance which would cause tumor regression in a single type of cancer without harm to the injected animal. The experiments reported in this paper were designed to assess the effects of direct contact between sarcoma-180 cells and various animal and human sera on the growth of and immunologic reactions to this tumor in mice. Kidd's experiments suggested that incubation of cancer cells with foreign sera, especially guinea pig serum, might cause the cells to become inactive without discernible effects on

On the nature of permeability of sarcoma-180 cells to amethopterin in vitro.

The rate of net influx of amethopterin into sublines of Sarcoma-180 cells in vitro (4.6 · 10 −8 cm/sec at 36°) was shown to be dependent on at least four processes: (a) slow influx, apparently by diffusion; (b) rapid, energy-dependent efflux; (c) very rapid binding of inhibitor to intracellular folic acid reductase (EC 1.5.1.3); and (d) slow dissociation of the latter complex. The influx of amethopterin requires a large activation energy of 30.5 kcal which is explained by poor lipid solubility and the net negative charge of the molecule. Influx is increased by increasing the H + -ion concentration, and also by high concentrations of Ca 2+ . Both of these are likely to decrease the ionization of the two carboxyl groups at the glutamyl-end of the molecule and thus facilitate their entry through the negatively charged cell surface. Ca 2+ probably acts by forming an [amethopterin-Ca 2 ] 2+ chelate. Combination of Ca 2+ and 2,4-dinotrophenol, an inhibitor of oxidative phosphorylation, increased the apparent rate of influx of amethopterin by 5–6-fold. Riboflavin, a natural ingredient of the growth medium, and folinic acid were found to be able to partially slow down the apparent rate of influx of amethopterin.

On the role of drug penetration in amethopterin resistance of Sarcoma-180 cells in vitro.

Abstract 1. 1. The measurement of the net rate of influx of amethopterin was based on the rate of formation of the intracellular folic acid reductase-amethopterin complex. This rate was found to be identical for the sensitive and resistant sublines of Sarcoma-180 cells cultured in vitro . It was slow (4.6·10 −8 cm/sec at 36°) and proportional to the concentration of amethopterin over a 1000-fold range. The net rates of influx of amethopterin and aminopterin were the same. 2. 2. It was mathematically proven that the degree of resistance to amethopterin would be directly proportional to the content of folic acid reductase in these cells if amethopterin were irreversibly bound by the enzyme. The lack of direct proportionality is caused by the intracellular dissociation of the enzyme-amethopterin complex. The final degree of resistance to amethopterin is determined by (a) the cellular folic acid reductase (EC 1.5.1.3) content, (b) the rate of growth, which determines the rate of synthesis of new enzyme, (c) the slow influx of amethopterin and its great affinity to folic acid reductase. These cause the concentration of the intracellular free amethopterin to be only a fraction of the extracellular amethopterin. Consequently, even a small degree of dissociation of the enzyme-amethopterin complex contributes to the degree of resistance to amethopterin. This dissociation becomes especially significant in the 3000-fold resistant subline in which folic acid reductase constitutes about 4% of the total cellular protein. In this subline less than 1% of the total enzyme needs to be functional for normal growth.

Studies on the interaction of vitamin B 12: Intrinsic factor and receptors : III. Chemical and biological properties of vitamin B 12 combining substances from the stomach, serum, and ascites fluid of the mouse

The chemical and biological properties of vitamin B 12 combining substances in mouse stomach extracts (MIF), mouse serum (MS), and mouse ascites fluid (AS) were compared. MIF was found to enhance the absorption of vitamin B 12 by rings of mouse and guinea pig ileum and to inhibit its absorption by sarcoma-180 (S-180) cells in vitro. MS and AS, on the other hand, enhanced the absorption of vitamin B 12 by S-180 cells but inhibited its absorption by rings of mouse and guinea pig ileum. It was shown that MIF is capable of forming a stable complex with vitamin B 12 at pH values from 1.0 to 11.0, while MS and AS are not capable of forming such stable complexes at pH values below 5.0. The complex formed between vitamin B 12 and MIF (MIF-vitamin B 12) is not adsorbed by zirconyl phosphate gel (Z-gel) at pH values higher than 5.0; however, the complexes formed between vitamin B 12 and MS (MS-vitamin B 12) or AS (AS-vitamin B 12) are adsorbed by Z-gel at pH values as high as 7.0. By chromatography on CM-cellulose, it was possible to separate MIF-vitamin B 12 from MS-vitamin B 12 and AS-vitamin B 12. The latter two complexes behaved identically when chromatographed on CM-cellulose.

Capacity for synthesis of a pyrimidine biosynthetic enzyme in mammalian cells

1. 1. The control of formation of the first enzyme unique to the pyrimidine pathway was studied in Sarcoma-180 cells growing in culture. Pyrimidine starvation, induced by the addition of 6-azauridine, produced a three- to four-fold increase in the level of aspartate transcarbamylase relative to total cell protein. 2. 2. The level of this enzyme was not changed either by the addition of preformed pyrimidines to growing cultures, or by the simultaneous addition of uridine and 6-azauridine. 3. 3. The results show that the level of aspartate transcarbamylase is under the control of end-product pyrimidines, and that the cells have a potential forincreased enzyme synthesis ordinarly not expressed.

Capacity for synthesis of a pyrimidine biosynthetic enzyme in mammalian cells

1. 1. The control of formation of the first enzyme unique to the pyrimidine pathway was studied in Sarcoma-180 cells growing in culture. Pyrimidine starvation, induced by the addition of 6-azauridine, produced a three- to four-fold increase in the level of aspartate transcarbamylase relative to total cell protein. 2. 2. The level of this enzyme was not changed either by the addition of preformed pyrimidines to growing cultures, or by the simultaneous addition of uridine and 6-azauridine. 3. 3. The results show that the level of aspartate transcarbamylase is under the control of end-product pyrimidines, and that the cells have a potential forincreased enzyme synthesis ordinarly not expressed.

Effect of 5-bromodeoxyuridine incorporation on survival of cultured mammalian cells

The analog 5-bromodeoxyuridine replaced 93–100% of DNA thymine in human-uterine fibroblasts and in mouse firboblasts when amethopterin was used to inhibit the synthesis de novo of thymidylate. The incorporation of the analog was followed by unbalanced growth and finally cell death. Even when extensive incorporation had occurred, some cells were able to resume normal growth in analog-free medium. This involved a drastic selection revealed by the appearance of isolated clones after 2.5–4 weeks. It is suggested that the few surviving cells are those which have escaped a lethal mutation. The possible occurrence of resistance to the analog or to amethopterin is excluded. Prolonged growth of amethopterin-resistant Sarcoma-180 cells in the analog-containing medium failed to interfere with the capacity of these cells to produce large amounts of folic acid reductase.