Morris Hepatoma Cells Research Articles

Synergistic induction of acyl-CoA oxidase activity, an indicator of peroxisome proliferation, by arachidonic acid and retinoic acid in Morris hepatoma 7800C1 cells

Morris hepatoma 7800C1 cells (a Wistar rat cell line) were exposed to 100 μM arachidonic acid in the medium for seven days. This treatment resulted in 150% and 60% increases (above control activities) in acyl-CoA oxidase (which catalyzes the first step in peroxisomal β-oxidation) and catalase activities, respectively. Arachidonic acid (C20:4) can be metabolized to 20- and 19-hydroxyarachidonic acid by cytochrome P-450IVA and it was shown that our cells are capable of forming 20-hydroxyarachidonic acid. However, 20-hydroxyarachidonic acid (0.l–0.8 μM, 4 days) had no effects on lauroyl-CoA oxidase and catalase activities in Morris hepatoma cells. Treatment of 7800C1 cells with 100 μM all- trans-retinoic acid resulted in inductions of catalase (160% above the control activity) and carnitine acetyltransferase (140% above the control activity) activities. The activity of lauroyl-CoA oxidase was often, but not always, slightly induced by treatment with all- trans-retinoic acid. When all- trans-retinoic acid was administered together with arachidonic acid, these two compounds had a synergistic effect on the induction of acyl-CoA oxidase activity (almost 700% above the control activity). However, treatment of Morris hepatoma cells with the man-made peroxisome proliferator, perfluorooctanoic acid, together with all- trans-retinoic acid did not result in any synergistic effect on this same enzyme activity. In summary, this study (1) corroborates findings from transfection experiments indicating that the heterodimer PPAR-RXR α activates transcription of the acyl-CoA oxidase gene using the Morris hepatoma cell line; (2) shows that arachidonic acid induces the activity of lauroyl-CoA oxidase; (3) suggests that transcription of the catalase gene is not regulated by a PPAR-RXR α heterodimer in this system; and (4) demonstrates that peroxisome proliferation in Morris hepatoma cells by perfluorooctanoic acid is not as dependent on the level of retinoic caused by arachidonic acid.

Polyamine acetylations in normal and neoplastic growth processes

The expression patterns of cytosolic and nuclear polyamine acetyltransferases were studied in normal and neoplastic growth processesin vivo andin vitro to evidentiate the roles played by these enzymes in cell proliferation. In regenerating liver, cytosolic spermidine/spermine N(1)-acetyltransferase showed similar augments of mRNA level and enzymatic activity during the prereplicative period (4-8 h), whereas spermidine N(8)-acetyltransferase activity increased later (24 h) when DNA synthesis was maximally enhanced. In fibroblasts continuously dividing, the messenger for spermidine/spermine N(1)-acetyltransferase rapidly accumulated after serum-stimulation. In cultured Morris hepatoma cells stimulated to logarithmic growth, spermidine N(8)-acetyltransferase activity remained at plateau for 1 day declining thereafter, while spermidine/spermine N(1)-acetyltransferase activity immediately decreased. In Yoshida AH-130 hepatoma cells transplanted in rat peritoneum, spermidine N(8)-acetyltransferase and spermidine/spermine N(1)-acetyltransferase activities rose, respectively, in concomitance with elevated proliferation-rate and quasi-stationary phase of growth. Since the expression of cytosolic and nuclear acetyltransferases underwent different temporal activation, an involvement of these enzymes in separate metabolic processes controlling normal and neoplastic growth may be suggested.

Induction of apoptosis by transforming growth factor-beta 1 in the rat hepatoma cell line McA-RH7777: a possible association with tissue transglutaminase expression.

We report here that transforming growth factor-beta 1 induces cell death in the Morris hepatoma cell line McA-RH7777. We assessed the type of cell death induced by transforming growth factor-beta 1 in this hepatoma cell line on the basis of morphological and biochemical characteristics. Dying cells, which detached from the cell monolayer, showed morphological characteristics of apoptosis (programmed cell death) such as chromatin condensation, nuclear disintegration and cellular fragmentation into clusters of eosinophilic globules. DNA isolated from these cells showed a ladder pattern consisting of multimers of 180 to 190 bp, indicating extensive DNA cleavage into oligonucleosomal units by an endogenous endonuclease. Treatment of the dead cells with detergents and chaotropic agents resulted in formation of insoluble shells, so-called apoptotic bodies, suggesting extensive cross-linking of cell proteins by tissue transglutaminase. Furthermore, increased amounts of cytosolic tissue transglutaminase, which has been recognized as a possible marker of apoptosis, and extensive cross-linking of cytokeratin polypeptides was demonstrated in TGF-beta 1-treated hepatoma cells on immunoblot analysis. These results provide strong evidence that the cell death induced by TGF-beta 1 in McA-RH7777 hepatoma cells is mainly apoptotic. It also suggests that a specific induction of the cytosolic tissue transglutaminase may be involved in the TGF-beta 1-induced pathways of apoptotic cell death in McA-RH7777 hepatoma cells.

Substrate and hormone regulation of palmitoyl-CoA synthetase in 7800 C1 Morris hepatoma cells and cultured rat hepatocytes

The effects of tetradecylthioacetic acid (TTA), insulin and dexamethasone on palmitoyl-CoA synthetase activity and its mRNA both in 7800 Cl hepatoma cells and cultured rat hepatocytes were studied. (1) When the hepatoma cells were cultivated in the presence of fatty acids or alkyl thioacetic acids (3-thia fatty acids) palmitoyl-CoA synthetase activity was increased several fold. The stronger effect was obtained with TTA, which also increased long-chain acyl-CoA synthetase mRNA significantly. TTA has no inducing effect on butyryl-CoA synthetase and little effect on octanoyl-CoA synthetase in the same cells. Dexamethasone also had inducing effect on palmitoyl-CoA synthetase in the hepatoma cells. Insulin counteracted the induction given by TTA. All of these regulation actions take place at the pretranslational level. (2) In isolated hepatocytes the activity of palmitoyl-CoA synthetase was much higher than in hepatoma cells, but it was lost rapidly in culture. The loss of the enzyme activity was slowed down in the presence of TTA and insulin, either alone or combined. Dexamethasone combined with TTA reversed the loss of enzyme activity, while dexamethasone alone even increased the loss. Analysis of palmitoyl-CoA synthetase mRNA shows that TTA prevents the loss of the enzyme activity by inducing mRNA of the enzyme, dexamethasone enhances the effect of TTA, while insulin stabilizes the enzyme activity in the cultured cells without increasing the mRNA level.

Ornithine decarboxylase and malignant growth

There is indirect evidence that key enzymes which catalyse both synthesis and degradation of polyamines (PA) may be useful as biological markers of malignant growth. Ornithine decarboxylase (ODC) activity was assessed in fetal, regenerating and adult rat liver, in hepatoma with different growth rate, in Morris hepatoma cell lines and in rat liver in the course of nitrosopiperidine (NP) carcinogenesis. A much higher ODC activity was observed in all the investigated tumor tissues and in regenerating rat liver as well. A significant decrease of ODC inductive synthesis was demonstrated both in liver and hepatoma of rats maintained on vitamin B6-deficient diet. The experimental suggest that inductive synthesis of ODC may be considered not only early, but also the mandatory event in any hyperproliferating processes including carcinogenesis.

Transcription of the gene for the gap junctional protein connexin43 and expression of functional cell-to-cell channels are regulated by cAMP.

We investigated the mechanism by which cyclic AMP (cAMP) induces gap junctional communication via cell-to-cell channels in a communication-deficient rat Morris hepatoma cell line. We found that under basal conditions, the cells transcribe cx43 at a low level but do not transcribe cx26 or cx32. Elevation of intracellular cAMP, which induced communication, increased cx43 mRNA 15- to 40-fold and the rate of cx43 transcription 6-fold. Cx43 protein was detected by immunostaining in junctions of only those cells in which communication had been induced. We found the regulation by cAMP also in other cell lines; namely, in those with a low basal level of cx43 mRNA.

Distribution of fibroblast growth factors in cultured tumor cells and their transplants

The distributions of acidic fibroblast growth factor (aFGF) and basic FGF (bFGF) in extracts of various cultured mammalian cells were determined from their elution profiles on heparin-affinity chromatography, and assay of activity as ability to stimulate DNA synthesis in BALB/c3T3 cells. Only aFGF was found in extracts of mouse melanoma B 16 cell and rat Morris hepatoma cell (MH1C1) lines. Other tumor cell lines established from solid tumors and some normal cells contained bFGF as a main component, but blood tumor cell lines contained no aFGF or bFGF. The FGFs in extracts of solid tumor tissues derived by transplantations of these cultured tumor cells and various normal tissues of mice were also examined. Tumors formed by all cell lines, regardless of whether they produced aFGF, bFGF, or neither, contained bFGF that was probably derived from host cells including capillary endothelial cells, in addition to the tumor-derived aFGF or bFGF, if produced. The content of bFGF, possibly derived from the host, in these tumor tissues was comparable to those of various mouse organs other than thymus, lung, spleen, and testis, which have higher bFGF contents. Tumor tissues derived from cultured cells producing bFGF had relatively higher bFGF contents. Like bFGF, aFGF was distributed almost ubiquitously in normal mouse tissues.

Bidirectional morphological changes induced by dexamethasone in Morris hepatoma cell lines McA-RH7777 and McA-RH8994: Independence of fibronectin and its receptor

Two Morris hepatoma-derived cell lines, McA-RH7777 (7777) and McA-RH8994 (8994), exhibit different alterations in morphology upon exposure to glucocorticoid. After treatment with synthetic glucocorticoid dexamethasone (DEX), 7777 cells show increased adhesiveness and more flattened shape, while DEX-treated 8994 cells show decreased adhesiveness to substratum and exhibit a marked increase of round and detached cells. Since fibronectin has been thought to play an important role in cell adhesiveness to substratum in hepatoma cell culture, we have also compared the effects of DEX on the biosynthesis of flbronectin (FN) and the functional level of FN receptor in 7777 and 8994 cells. Northern blot analysis and immunofluorescent studies showed that 7777 cells have a high basal expression level of FN synthesis and that DEX treatment induces FN expression two- to threefold with establishment of an extensive fibrillar FN network around the cells. On the other hand, 8994 cells were shown to express little FN and no apparent FN was localized on nonstimulated 8994 cells. However, DEX-treatment drastically increased FN expression in 8994 cells to the level of more than that of DEX-treated 7777 cells and induced a detectable level of cell-associated FN around DEX-treated 8994 cells, which appears to be contradictory to the decreased adhesiveness to the substratum in DEX-treated 8994 cells. Cell attachment assays using FN-coated plates demonstrated that DEX does not exhibit significant effects on the attachment of either 7777 or 8994 cells to FN-coated dishes. Our results suggest that decrease of adhesiveness to the substratum and increase of round detached cells in DEX-treated 8994 cells are independent of changes in the FN expression and the function of FN receptor.

Transforming growth factor-β1 stimulates proliferation of Morris hepatoma cells in serum-free soft agar culture system supplemented with EGF and insulin

The anchorage-independent growth of Morris hepatoma 7777 (MH) cells in serum-free medium was examined. The influence of insulin, epidermal growth factor and transforming growth factor beta 1 (TGF-beta 1) on [3H]thymidine incorporation and colony formation of the investigated cells was described. Contrary to normal rat hepatocytes TGF-beta 1 plus EGF and insulin were found to stimulate MH cells proliferation in presented conditions. A simple, chemically-defined culture system suitable for research on mitogenic peptides was proposed.

Dexamethasone inhibition of rat hepatoma cell growth and cell cycle traverse is reversed by insulin

(1) The growth of 7800 C1 Morris hepatoma cells was inhibited by dexamethasone. The inhibition was detectable at 1 nM and half-maximal effect was obtained with approx. 13 nM dexamethasone. About 80% growth inhibition was obtained with 250 nM of the hormone and the growth rate was normalized on cessation of treatment. (2) These hepatoma cells contain dexamethasone receptors with equilibrium dissociation constant of 0.24 nM and a capacity of 24 fmol/mg cell protein. Treatment of the cells with insulin did not change these dexamethasone binding properties. Binding experiments showed that 2, 10 and 100% of the receptors were occupied when the cells were incubated with 1 nM, 7 nM and 250 nM dexamethasone, respectively. (3) Insulin completely counteracted the growth inhibition by dexamethasone and antagonized the induction of peroxisomal acyl-CoA oxidase and tyrosine aminotransferase caused by the glucocorticoid. (4) Micro-flow fluorometry showed that the cultures had a major diploid DNA stem line and a minor tetraploid stem line. Changes in diploid, tetraploid and S phase cells of the diploid stem line were scored. Dexamethasone reduced the proportion of cells in S phase and of tetraploid cells. Insulin partly reversed the action of dexamethasone in S phase, but prevented the reduction in tetraploid cells caused by dexamethasone. (5) The mitotic rate was significantly reduced by dexamethasone and this effect was reversed by insulin. (6) Continuous [ 3H]methylthymidine labelling showed a growth fraction of unity in all treatment groups. (7) It is concluded that dexamethasone induces growth inhibition by reducing the G 1-S transition. Insulin is able to counteract this effect and increase the rate of DNA synthesis.

Synergistic actions of tetradecylthioacetic acid (TTA) and dexamethasone on induction of the peroxisomal β-oxidation and on growth inhibition of Morris hepatoma cells: Both effects are counteracted by insulin

(1) The relation between the effects of the sulfur-substituted fatty acid analogue, tetradecylthioacetic acid (TTA), dexamethasone and insulin on enzyme induction and growth rate was studied in Morris hepatoma 7800 C1 cells in culture. (2) The activities of the cynanide-insensitive palmitoyl-CoA oxidase and palmitoyl-CoA hydrolase were induced about 2-fold by 50 μM TTA after 72 h of treatment. Catalase was less induced and NADPH-cytochrome- c 2 reductase, glucose-6-phosphate dehydrogenase and lactate dehydrogenase were unaffected by the fatty acid analogue. (3) Dexamethasone (250 nM) induced the same enzymes as did TTA, but was a less efficient than 50 μM TTA. However, in combination their effects were more than additive, resulting in 4–7-fold increases. (4) Insulin (400 nM) counteracted the inductive effects of both TTA and dexamethasone on all enzymes except for lactate dehydrogenase, which was induced by the combination of all three compounds. (5) TTA inhibited the growth rate of the cells, and this effect was potentiated by dexamethasone and counteracted by insulin. (6) The enzyme inductions were similar in exponential and plateau phases of growth, indicating that these processes were independently affected by the three compounds.

DNA polymerase α from normal rat liver is different than DNA polymerases α from Morris hepatoma strains

To investigate whether DNA replication in rat hepatoma cells is altered compared with that in normal rat liver, the main replicative enzyme, i.e. the DNA polymerase alpha complex, was partially purified from a slow-growing (TC5123) and a fast-growing (MH3924) Morris hepatoma cell strain as well as from normal rat liver. The purified DNA polymerase alpha complexes contained RNA primase. DNA polymerase alpha activities of these complexes were characterized with regard to both their molecular properties and their dNTP and DNA binding sites. The latter were probed with competitive inhibitors of dNTP binding, resulting in Ki values, and with DNA templates, yielding Km values. The sedimentation coefficients of native DNA polymerases alpha from Morris hepatoma cells were found to be lower than that of polymerase alpha from normal rat liver. Consequently, when following the procedure of Siegel and Monty for determination of molecular mass considerably smaller molecular masses were calculated for polymerases of hepatoma strains (TC5123, 127 kDa; MH3924, 138 kDa; rat liver, 168 kDa). Similar differences were found when the dNTP binding site was probed with inhibitors. Ki values obtained with butylphenyl-dGTP were higher for polymerases of the hepatoma strains than for that of normal rat liver. However, Ki values measured with aphidicolin and butylanilino-dATP were lower for DNA polymerase alpha from the fast-growing hepatoma cell strain than for that from normal rat liver, indicating a reduced affinity of the dNTP binding sites for dATP and dCTP. This reduced affinity could be responsible for lowered specificity of nucleotide selection in the base-pairing process which in turn may cause an enhanced error rate in DNA replication in malignant cells. Furthermore, when the DNA binding site was characterized by Michaelis-Menten constants using gapped DNA as a template, Km values were similar for all three DNA polymerases. In contrast, the Km value measured with single-stranded DNA as a template was found to be lower for DNA polymerase alpha from the fast-growing hepatoma MH3924 than for that from normal rat liver. Thus, the DNA-polymerizing complex from MH3924 combines both higher binding strength to single-stranded DNA templates and decreased nucleotide selection, properties which may enhance replication velocity and may lower fidelity.

Activities of enzymes of lipid metabolism in Morris hepatoma 7800 C1 cells

(1) The rate of palmitate oxidation in the 7800 C1 Morris hepatoma cells was about 60% of the activity observed in hepatocytes. The stimulatory effect of glucagon in hepatocytes was not observed in the hepatoma cells. The rate of fatty acid synthesis from [2- 14C]acetate in the hepatoma cells was 1/20 of the activity in hepatocytes. The conversion of [2- 14C]acetate to cholesterol was not different in the two kinds of cell. (2) Acetyl-CoA carboxylase and fatty acid synthetase were significantly decreased in the hepatoma cells. The hepatoma cells had, however, raised activities of malate dehydrogenase (decarboxylating), and glucose-6-phosphate and 6-phosphogluconate dehydrogenases. (3) The activities of the enzymes were not affected by different concentrations of glucose or palmitate in the culture medium. Insulin, dexamethasone, triiothyronine and glucagon had no effect on the enzyme activities. This is in contrast to the adaptation of the peroxisomal β-oxidation system, which is induced by fatty acids and modified by hormones.

Induction of peroxisomal β-oxidation in 7800 C1 Morris hepatoma cells in steady state by fatty acids and fatty acid analogues

(1) The activities of peroxisomal β-oxidation and palmitoyl-CoA hydrolase in Morris hepatoma 7800 C1 cells were studied. The cells were grown until they reached steady state (constant DNA content per dish) and then were cultured in the presence of fatty acids or alkylthioacetic acids, i.e., S-substituted fatty acid analogues. (2) The fatty acid analogues increased the activity of the cyanide-insensitive palmitoyl-CoA oxidase several-fold. The effect was dose-dependent; 5 μM tetradecylthioacetic acid (TTA) was sufficient to give a significant induction. With 20 μM TTA, the increase in enzyme activity was discernable after 3 h and reached a maximum after 3 days. The inducing effect of the alkylthioacetic acids increased with the length of the hydrophobic alkyl end of the analogue. The inducing ability disappeared when the fatty acid analogue was Ω-oxidized to the corresponding dicarboxylic acid. Oxidation of the sulfur atom resulted in inhibited cellular uptake and abolished enzyme induction. (3) At higher concentrations (0.5–1 mM), normal fatty acids also induced cyanide-insensitive palmitoyl-CoA oxidation. Myristic acid was the most potent inducer, whereas fatty acids with shorter as well as longer carbon chains were less efficient. The inducing effect increased with the number of double bounds in the fatty acid. (4) The normal fatty acids as well as the fatty acid analogues also induced palmitoyl-CoA hydrolase, but the relative changes were much less pronounced than with the palmitoyl-CoA oxidase.

Criteria of sensitivity of Zajdela and morris hepatoma cells to glucocorticoids

Glucocorticoids induce tyrosine aminotransferase synthesis in 7777 Morris hepatoma but fail to do so in Zajdela hepatoma. This internal property indicates the resistance to the hormone. However, both hepatoma cell lines do respond to the triamcinolone acetonide in a similar way, as judged by some other criteria, e. g. interaction with the immobilized hormone on the inert carrier, adhesion to glass and kinetic parameters of alkaline phosphodiesterase I activity. Moreover, both cell types respond to glucocorticoids by modification of synthesis of some proteins, as revealed previously by two-dimensional electrophoresis. The results show that in case of tumour cells which retain their specific receptor apparatus but do not respond to glucocorticoids by usual criteria, the conclusion whether tumour cells are hormone-sensitive or not has to be drawn from the analysis of their multiple response judging by several assays.

An affinity labeling of ras p21 protein and its use in the identification of ras p21 in cellular and tissue extracts.

We have carried out photoaffinity labeling of the ras p21 protein, a ras oncogene product, with [alpha-32P]GTP. Based on our studies, a sensitive, rapid, and specific assay for the detection of multiple forms of ras p21 has been developed. The specificity of this protocol is shown by (a) sensitivity of affinity labeling of ras p21 to known inhibitors of GTP binding and (b) immunoprecipitation of affinity labeled protein with anti-ras p21 serum. Detection and semiquantitation of ras p21 by this method is accomplished in less than 24 h and requires as little as 100,000 cells or about 5 mg of tissue sample from skin tumor, liver, and mammary tumor tissues. Furthermore, using this approach, we were able to detect the selective loss of one species of ras p21 in transplanted Morris hepatoma cells.

Synergistic effect of coumadin and cytoxan in reducing lung metastases

Lung metastasis is a frequent cancer complication resulting in significant mortality. This study evaluates the effect of coumadin and cytoxan alone and in combination on lung metastases in rats challenged with Morris hepatoma 3924A. Seventy-seven male American Cancer Institute (ACI) rats weighing 200 g were studied. Thirty-seven rats received coumadin orally for six days, which resulted in a prothrombin time 2 times that of controls (30 sec). All rats received 1 × 10 5 clumped Morris hepatoma cells via tail vein injection. Animals were divided into four groups: Group I (controls, n = 20) received no antitumor treatment; group II rats (n = 20) received 25 mg/kg cytoxan intraperitoneally at the time of tumor challenge; group III animals (n = 19) received coumadin alone; while group IV (n = 18) received both coumadin and cytoxan. Rats were evaluated for number of lung metastases and lung weight at 3 weeks postinjection. Data was subjected to statistical analysis by the Student's t test. The mean number of lung metastases were 580 ± 45 in group I, 350 ± 310 in group II, 330 ± 263 in group III, and 200 ± 161 in group IV ( P < .005 [IV] v [I], P < .05 [IV] v [II], [III]), ( P < .05 [II] v [I]), ( P < .05 [III] v [I]). Mean lung weights were 2.597 g ± 1.65 in group I, 2.049 g ± 0.75 in group II, 1.898 g ± 0.80 in group III, and 1.677 g ± 0.31 in group IV. ( P < .025 [IV] v [I], P < .05 [IV] v [II]). These data indicate coumadin and cytoxan act synergistically in reducing lung metastases. These observations suggest the synergistic effect of these two agents may prove useful in the prevention of lung metastases in the clinical setting.

Characteristics and specificity of the glucocorticoid "carrier" of rat liver plasma membrane.

The uptake of corticosterone by highly purified plasma membrane vesicles of rat liver was studied by a rapid-centrifugation technique which allows uptake measurements within 5 s. The vesicles are free of soluble cytoplasmic constituents. Therefore, association of hormone with the vesicle is attributed entirely to components of the vesicle-membrane. Half maximal uptake is reached at 8 s at 21 degrees C. At 15 degrees C transition of the lipid state in the membrane leads to a decrease of uptake, a characteristic property common to membrane mediated processes. The uptake of corticosterone is saturable and reversible but does not follow normal saturation kinetics. The apparent dissociation constants of three uptake systems bear direct relation to the concentration of free corticosterone in rat plasma (4-16 nM) supporting a physiological role for the system. Uptake of corticosterone decreases with decreases in vesicular volume; about 50% of the hormone is bound specifically and 50% is transported to the lumen of the vesicle. Since outflow of intravesicular hormone also occurs readily, the uptake and transport is proposed to be mediated by putative "carriers". The "carrier" preferentially transports glucocorticoids; dexamethasone is not taken up by this putative molecule. Steroids with 5 alpha conformation are more potent inhibitors of the "carrier" for corticosterone than 5 beta-steroids. Androgens and estrogens are weak competitors of corticosterone. The affinity of the "carrier" for several hormones differs considerably from that of the cytoplasmic receptor. Morris hepatoma cells (MH 3924) do not take up corticosterone. Our results prompt us to propose the hypothesis that the transport function of the "carrier" and the binding of the hormone by the cytoplasmic receptor are two different entities; perturbation of the "carrier" may lead to steroid unresponsiveness. Normal expression of steroid hormone activity is manifested in the concerted action of the functionally sound cell membrane "carrier" and the intracellular receptor.

Study of carbohydrate metabolism in glycogen storing cell lines derived from cultured rat hepatocytes

1. 1. Some aspects of carbohydrate metabolism were investigated in three non-malignant, glycogen storing, cell lines derived from a primary culture of rat hepatocytes, and in the Morris hepatoma 3924 cells. 2. 2. The three cell lines show biochemical alterations which are, to a large extent, similar to those found in the hepatoma cells: increased activity of glycolytic enzymes and decreased activity of gluconeogenetic enzymes. 3. 3. An increase of glucose-6-phosphate dehydrogenase activity is also found. 4. 4. The three cell lines, as the Morris hepatoma cells, actively convert glucose into lactate under the in vitro conditions of culture. Fructose is not taken up as quickly as glucose and galactose is not metabolized. 5. 5. As compared with normal hepatocytes, the three cell lines have altered metabolism and growth behaviour. They largely resemble the preneoplastic cells appearing in rat liver at the early stages of experimental carcinogenesis.

Protamine Sulfate Inhibition of Serum-Induced Mitogenic Responses: Differential Effects on Normal and Neoplastic Cells&lt;xref ref-type="fn" rid="FN2"&gt;2&lt;/xref&gt;

Protamine sulfate reversibly inhibits serum-induced mitogenic stimulation of several nontransformed and neoplastic cell types in vitro. Fifty percent inhibition was induced by approximately 120-150 micrograms protamine sulfate/ml. Cells were affected directly, and inhibition depended on the duration of cell exposure. Heparin, chondroitin sulfate, heparan sulfate, and dextran sulfate neutralized protamine sulfate effects during the early stages of treatment. Nontransformed cells [bovine aortic endothelial cells, adult human gingival fibroblasts (strains 423 and 1101), fetal rat skin (strain 921-K) and muscle fibroblasts] required longer exposure to induce inhibition than did neoplastic cells [rat 3-methylcholanthrene-induced fibrosarcoma cell lines (MCA-6 and MCA-9), a macrophage-like cell line (NCTC-3749), Walker 256 rat carcinoma cells (ATCC-CCL-38), rat Morris hepatoma cells (ATCC-CCL-144), murine melanoma cells (B16), and rat bladder squamous cell carcinoma cells (804-G)]. Other polycationic compounds, including histone type VIII-S, poly-L-lysine, poly-L-arginine, and protamine (free base), were also effective inhibitors, whereas the basic proteins cytochrome c and lysozyme had no effect. Poly-L-histidine, poly-L-glutamic acid, poly-L-aspartic acid, and dextran blue also had no inhibitory effect.