Selective effects of two chloromethyl ketones on amino acid and phosphate uptake in rat liver and tumors.

Injection of L-1-tosylamido-2-phenylethyl chloromethyl ketone (TPCK) at a level of 10 mg/100 g body weight inhibited the incorporation of 3H-labeled amino acids into protein in Morris hepatomas 7777and 9618A2. The degree of inhibition was similar in cytoplasmic proteins and in histone and nonhistone nuclear protein fractions. There was no inhibitory effect on 3H-labeled amino acid incorporation in the livers of the tumor-bearing rats. The inhibitory effect of N-tosyl-L-lysine chloromethyl ketone (TLCK) on incorporation of 3H-labeled amino acids was observed in both the slowly growing hepatoma 7787 and the rapidly growing hepatoma 7777. In hepatoma 7777, TLCK (2.5 mg/100 g body wt) exerted a greater inhibitory effect on incorporation when administered 60 minutes before [3H]leucine injection than when injected simultaneously. Studies on tissue uptake of amino acids, thymidine, and phosphate indicated that inhibitory effects of TPCK and TLCK on active transport may be a major factor in the action of these drugs on macromolecular synthesis. The inhibitory effects of TPCK and TLCK seen in transplanted hepatomas and a colon tumor were not generally seen in normal tissues of the tumor-bearing rats.     

Tumor-selective inhibition of the incorporation of 3H-labeled amino acids into protein by cyanate.

Sodium cyanate at a dose level of 125 or 250 mg/kg i.p. caused an inhibition of incorporation of 3H-labeled amino acids into cytoplasmic and nuclear proteins of the rapidly growing hepatoma 7777 and the slowly growing hepatoma 9618A. There was no inhibitory effect on 3H-labeled amino acid incorporation into protein in the livers of rats bearing these tumors. Studies on the effects of sodium cyanate on incorporation of 3H-labeled amino acids into total acid-insoluble material indicated that a greater than 85% inhibition could be achieved in hepatoma 5123C, hepatoma 9618A2, and the MK3 kidney tumor with either little or no effect in host liver, kidneys, brain, skeletal muscle, intestinal mucosa, and regenerating liver after partial hepatectomy.     

Effect of camptothecin and adriamycin on bleomycin-induced tritiated thymidine triphosphate incorporation in a rat nuclear system.

We investigated the effect of camptothecin and adriamycin on [3H]TTP incorporation and bleomycin-stimulated [3H]TTP incorporation in host liver and hepatoma nuclei of rats. Camptothecin neither stimulated nor inhibited incorporation in the regular nuclear incorporating system. Bleomycin stimulated incorporation to a much greater extent in host liver nuclei and slow-growing hepatomas than it did in the fast-growing hepatoma 7777. Addition of camptothecin to bleomycin stimulated incorporation of [3H]TTP even further. This camptothecin stimulation was slightly greater in hepatoma nuclei than it was in host liver nuclei. Adriamycin inhibited [3H]TTP incorporation in the regular system as well as the bleomycin-induced incorporation. Hepatoma nuclei were more sensitive to this inhibition than were host liver nuclei. Sucrose density gradients indicated that camptothecin caused DNA strand scissions in addition to those produced by bleomycin. Camptothecin alone produced some single-strand but no double-strand scissions. The action of bleomycin was dependent on sulfhydryl-reducing agents. Camptothecin could partially substitute for this requirement. Adriamycin did not produce DNA breaks as determined by neutral or alkaline sucrose density gradients. Despite complete inhibition of bleomycin-induced [3H]TTP incorporation, adriamycin did not prevent bleomycin-induced DNA breaks. The inhibitory effect of adriamycin might have been on the repair system.     

Differentiating and growth inhibitory effects of diallyl disulfide on cancer cells

Diallyl disulfide caused growth inhibition and differentiation of DS19 mouse erythroleukemic cells as judged by hemoglobin synthesis and induction of acetylcholinesterase activity. There was a 50% inhibition of cell division at about 0.25 mM diallyl disulfide which was much more effective than diallyl sulfide. K562 human erythroleukemia cells and mouse melanoma cells were more resistant to the action of diallyl disulfide. Thymidine incorporation into DNA in 7800NJ and 7288CTC rat hepatoma cells and in T47D and MCF7 human breast cancer cells was inhibited by 1-2 mM diallyl disulfide. Administration of diallyl disulfide to rats bearing Morris hepatomas caused marked inhibitory effects on precursor incorporation into DNA and protein in both hepatomas and in livers after a dose of 400 mg/kg body weight, but only small differences were seen at a less toxic dose of 200 mg/kg.     

Epidermal growth factor and proliferation in rat hepatocytes in primary culture isolated at different times after partial hepatectomy

Primary hepatocyte cultures have been prepared from normal adult rat liver and from rat liver at 4, 8, 12, 24, and 48 h following partial hepatectomy (removal of 70% of the liver). Cells were maintained in minimal essential medium alone or supplemented with hormones. Comparing DNA synthesis in normal adult rat hepatocytes with DNA synthesis in hepatocytes isolated from regenerating livers, we found with minimal essential medium alone little DNA synthesis in normal adult rat hepatocytes and in hepatocytes isolated 4, 8, or 12 h after 70% hepatectomy. In hepatocytes isolated 24 h after partial hepatectomy, however, the incorporation of [3H]thymidine was 3 times the rate of normal hepatocytes. The addition of insulin to minimal essential medium had minimal effect on DNA synthesis in all hepatocytes. Addition of epidermal growth factor alone or in combination with insulin resulted in a dramatic increase in DNA synthesis in hepatocytes from regenerating rat liver. Increased incorporation was detectable as early as 4 h after partial hepatectomy and reached a maximum at 24 h after the operation. Results obtained with [3H]thymidine incorporation were confirmed by autoradiography and by direct DNA determinations in hepatocyte cultures. Epidermal growth factor binding to the hepatocytes was determined and agreed with previously reported binding studies. Binding of epidermal growth factor in hepatocytes isolated at 4 h after partial hepatectomy was the same as in normal hepatocytes but was undetectable in hepatocytes isolated from rats at 12 and 24 h after partial hepatectomy.     

DNA synthesis in membrane-denuded nuclei and nuclear fractions from host liver and Morris hepatomas.

Incorporation of [3H]TTP into membrane-denuded nuclei and fractions of these nuclei from host liver and Morris hepatomas has been compared. Treatment of sucrose nuclei with Triton X-100 removed 95% of the phospholipids and 15 to 20% of the protein. These membrane-denuded nuclei remained physically stable. The Triton X-100-extracted nuclei incorporated label into their DNA in nuclear-incorporating system similar to sucrose nuclei with their membranes intact. Triton X-100-treated nuclei from hepatoma 7777 incorporated six times more label and those from hepatoma 7800 incorporated three times more label than Triton X-100-treated host liver nuclei. Nuclei from the three sources incorporated more label when exogenous DNA was added to the incubation system, but the difference in incorporation between the hepatoma nuclei and the host liver nuclei disappeared. When Triton X-100-treated nuclei, prepared from a tumor-bearing animal given injections of [3H]thymidine for 10 min were fractionated on sucrose gradients after disruption by high Mg2+ concentration, the fractions from hepatoma 7777 nuclei contained six times as much label as the host liver nuclear fractions. Nuclear fractions prepared from unabeled hepatomas or host livers had DNA polymerase activity. The activity, however, is the same in fractions prepared from hepatoma 7777 or host liver nuclei. It is suggested that the nuclear membrane does not play an important role in nuclear DNA synthesis. It is further suggested that the increased incorporation found with hepatoma nuclei is dependent on a physical or chemical arrangement of components within the nucleus and not solely on different enzyme levels.     

Assessment of the anti-tumor potential of glutathione

This study assesses the effect of reduced glutathione (GSH) on regenerating liver, 3'-methyl-4-dimethylaminoazobenzene (3'-MDAB) hepatocarcinogenesis, and normal and transformed hepatocytes in vitro. GSH administered intragastrically caused only a 30% reduction in thymidine incorporation into liver DNA at 24 h after partial hepatectomy; there was no apparent effect on RNA and protein synthesis. Furthermore, in 3'-MDAB induced hepatocarcinogenesis, all GSH-treated animals developed hepatocyte nodules, and serum alpha-fetoprotein (AFP) levels were not reduced. In vitro, GSH was shown to be cytotoxic to both normal and transformed hepatocytes at serum concentrations under 10%. GSH inhibited [3H]thymidine incorporation slightly in 2 transformed hepatocyte lines, but not in normal hepatocytes.     

Effect of bleomycin on [3H]Thymidine 5'-Triphosphate incorporation into host liver and hepatoma nuclei.

The effect of bleomycin on [3H]thymidine 5'-triphosphate ([3H]TTP) incorporation into isolated sucrose nuclei from host liver and Morris hepatomas has been compared. Bleomycin stimulates [3H]TTP incorporation 13-fold in host liver and hepatoma 16 nuclei, 8-fold in hepatoma 7800 nuclei, and 3-fold in hepatoma 7777 nuclei. Differences in the nuclear membranes are not responsible for the different response of the nuclei. Nuclei, denuded of their membranes by Triton X-100 treatment, give similar results to sucrose nuclei. Analysis of DNA extracted from liver or hepatoma nuclei incubated with bleomycin indicates that bleomycin produces scissions in the nuclear DNA and that some repair synthesis takes place. Incubation of nuclei with 111indium-labeled bleomycin shows an equal binding capacity of liver and hepatoma nuclei for bleomycin. Bleomycin also stimulates incorporation of [3H]TTP in a system using chromatin or calf thymus DNA as primer. Host liver or hepatoma chromatin incubated with a DNA polymerase extracted from normal rat liver nuclei is stimulated approximately to the same extent by bleomycin. When DNA polymerase extracts from host liver and hepatoma nuclei are assayed with calf thymus DNA as primer, bleomycin has a greater stimulatory effect on [3H]TTP incorporation with host liver DNA polymerase than with hepatoma DNA polymerase in the system. We suggest that a defect in the repair system in hepatoma nuclei is responsible for the relatively lower response to bleomycin.     

Hypermethylation of replicating hepatic DNA following N-methyl-N-nitrosourea administration

Changes in the degree of methylation of cytosine in DNA are considered to be mechanistically important in modulating gene expression. To gain a better understanding of the relationship(s) linking onco-proliferative processes and enzymatic DNA methylation, a study has been carried out on the hepatic DNA methylation pattern during DNA replication following partial hepatectomy (PH), mitogen treatment and N-methyl-N-nitrosourea (MNU) administration in rats. The following results were obtained: (i) DNA hypomethylation was seen during DNA synthesis, with each of the 3 stimuli, namely MNU administration, partial hepatectomy, and hepatomitogen treatment; (ii) the level of DNA hypomethylation was not quantificatively related to the extent of DNA replication as measured by incorporation of [³H]thymidine into hepatic DNA: (iii) MNU administration under conditions conducive to carcinogenic development, i.e. during the S phase of compensatory cell proliferation, caused hypermethylation of replicating hepatic DNA, as shown by Hpall and Mspl restriction patterns.     

Biochemical and morphological changes in hepatic nuclear membranes produced by N-hydroxy-2-acetylaminofluorene.

The effect of N-hydroxy-2-acetylaminofluorene on the ultrastructure and synthesis of hepatic neclear membranes was evaluated in partially hepatectomized rats. The incorporation of L-[4,5-3H]leucine into two nuclear membrane fractions increased within 2 hr after hepatic resection and reached a peak at 20 hr. After partial hepatectomy, the decay of radioactivity in nuclear membrane proteins labeled with L-[4,5-3H]leucine revealed similar half-lives for the two membrane fractions when compared to those obtained from sham-operated animals. The protein concentration of the nuclear membrane fraction of higher density decreased sharply within 2 hr after partial hepatectomy and remained low throughout a 20-hr postoperative period. Polyacrylamide gel electrophoresis of both nuclear membrane fractions showed a similar composition. Nine proteins were resolved, varying from 21,000 to 190,000 daltons. The two major protein bands were in the range of 50,000 and 70,000 daltons, respectively. Treatment of partially hepatectomized animals with N-hydroxy-2-acetylaminofluorene showed marked dilation of the nuclear envelope and rough endoplasmic reticulum in situ upon electron microscopic examination. Vacuolization and evagination of the perinuclear membranes were also noticeable in isolated nuclei obtained from carcinogen-treated rats. Inhibition by N-hydroxy-2-acetylaminofluorene of the incorporation of L-[4,5-3H]leucine into the nuclear membranes was dose-dependent and remained depressed throughout a 60-min labeling period. These results suggest that the inhibitory effects on RNA and protein synthesis previously shown to be produced by this arylhydroxylamine hepatocarcinogen may lead to disruption of the morphology and synthesis of the nuclear envelope.     

Mechanism of potentiation of antitumor activity of 5-fluorouracil by guanine ribonucleotides against adenocarcinoma 755

The effect of various guanine ribonucleotides on the antitumor activity of 5-fluorouracil (FUra) was investigated by its action on adenocarcinoma 755, 5′-GDP and 5′-GMP were both equally effective in potentiating the antitumor activity of FUra without increasing toxicity. 5′-GTP and 5′-IMP also potentiated the activity but not as much as 5′-GMP. 2′-GMP and 3′-GMP did not enhance the antitumor activity. In contrast, cGMP antagonized the effects of FUra.The incorporation of ^3H-labeled FUra into RNA or DNA showed there was no obvious association between the incorporation and antitumor activity after any treatment with guanine ribonucleotides. The combination of FUra and 5′-GMP produced the greatest inhibition of RNA synthesis. The combination of FUra and 2′-GMP had no effect on RNA synthesis. The inhibition of RNA synthesis may be the result of decreased pyrimidine pool size and increased incorporation of FUra into RNA. Potentiation of the antitumor activity of FUra by 5′-GMP was reversed by the injection of cytidine. Moreover, the combination of 5-fluorocytidine (FCyd) and 5′-GMP showed greater antitumor activity than FCyd alone. These results indicate that a decreased CTP pool potentiates the antitumor activity of FUra.Thus, 5′-GMP or 5′-GDP strongly enhanced the antitumor activity of FUra, and the potentiation resulted from the inhibition of RNA synthesis caused by reduction of the CTP and UTP pool sizes and increased incorporation of FUra into RNA.     

Inhibition of tritiated thymidine incorporation in cultured cells by rat kidney extract.

KCl extract from rat kidney, rat liver, and Morris hepatomas inhibited [3H]thymidine incorporation into cultured cells. Tissues came from male inbred BUF rats. The most pronounced inhibition was achieved with the kidney extract. Protein synthesis was not inhibited during a 24-hour exposure of the cells to the inhibitor. Incorporation of [3H]deoxycytidine was inhibited, as was cell growth, when the kidney KCl extract was present for several days. [3H]thymidine incorporation was inhibited almost immediately after the addition of the extract. The inhibition was reversible. Regular [3H]thymidine incorporation was restored 24 hours after removal of the inhibitor, which was neither arginase nor a thymidine-degrading enzyme. The inhibitor was stable to heat (80 degrees C for 10 min) and resistant to trypsin, pronase, DNase, and RNase. Exposure of the extract to proteolytic enzymes, hyaluronidase, and neuraminidase resulted in a loss of inhibitory activity only after extensive dialysis of the treated extract. The inhibitor appeared to be a mucoprotein in which the carbohydrate moiety may be responsible for the inhibition. The KCl extract also inhibited RNA synthesis and DNA synthesis by the de novo pathway. The inhibition of phosphorylation of thymidine, however, appeared to be the primary action of the inhibitor.     

A single-dose protocol for azaserine initiation of pancreatic carcinogenesis in the rat

Previously, the induction of pancreatic carcinogenesis in the rat using azaserine has involved a multiple-dose treatment protocol. The objective of the present study was to determine the effect of multiple azaserine treatments on pancreatic DNA synthesis and to develop a protocol for a single-dose initiation of pancreatic carcinogenesis by azaserine in the rat. Pancreatic DNA synthesis in young rats, which was determined by measuring the amount of [³H]-thymidine incorporation into DNA, was found to be elevated at 4.3 weeks of age and to decrease to a baseline level by 6.3 weeks. Treatment of 4-week-old rats with azaserine resulted in a dose-dependent inhibition of [³H]-thymidine incorporation into both pancreatic and liver DNA. Maximum inhibition was seen at 10 mg/kg body weight. This inhibition was followed by a gradual return of incorporation to normal values over a 48 h period. One week following pretreatment with four weekly injections of azaserine at 30 mg/kg, [³H]-thymidine incorporation into pancreatic and liver DNA was significantly elevated, suggesting that multiple injection protocols cause enhanced DNA synthesis which could have a co-carcinogenic and/or promotional effect. Single-doses of azaserine (10, 30 and 60 mg/kg) given at 7 weeks of age caused the appearance of more atypical acinar cell nodules (AACN) than when given at 5 weeks of age. The most effective dose was 30 mg/kg. Using alkaline elution, we determined that this response was due to the occurrence of more DNA damage in the 7-week-old animals. Thus, these results demonstrate a rationale for the use of single-dose initiation protocols in the pancreas. An effective single-dose protocol for induction of AACN in azaserine-treated rats fed semi-synthetic diet is presented.     

Destruction of erythroleukemia,myelocytic leukemia and burkitt lymphoma cells by photoactivated protoporphyrin

The effect of protoporphyrin on erythroid, myeloid and lymphoid leukemic cells and their destruction induced by the photoactivated porphyrin was studied. Friend erythroleukemic cells (FL) and myelocytic leukemic cells (ML) accumulated protoporphyrin in a cap or patch-like pattern observed by fluorescence microscopy. Photoactivated protoporphyrin induced the appearance of “holes” on the cell membrane demonstrated by scanning electron microscopy. On the other hand, Burkitt lymphoma (BL) and mastocytoma (MS) cells accumulated porphyrin intracellularly around the nuclear envelope and as circular profiles, respectively. Photoactivated protoporphyrin induced development of multiple blebs on the cell membrane, and even complete cell destruction. Cytotoxicity of protoporphyrin at short-term incubation periods was determined by [³H]thymidine and [³H]uridine incorporation. Protoporphyrin, unexposed to light, reduced the incorporation of both precursors only to a moderate extent. On the other hand, porphyrin-treated cells exposed to light showed complete inhibition of RNA and DNA synthesis. Long-term exposure of ML and BL cells to porphyrin in the dark induced a nearly 50 % inhibition of RNA and DNA synthesis. Although the cytotoxic effect of protoporphyrin in the dark was lower than that of photoactivated porphyrin, this may possess a potential activity in vivo even without illumination.     

Metabolism of nucleic acids in normal, regenerating and neoplastic tissues of the liver]

Under study was the kinetic growth of three hepatomas-22A, 60 and 46, characterized by a various degree of differentiation. There was found a correlation between the degree of differentiation and parameters of the hepatomas growth. For the hepatomas a correlation between the DNA content and rate of growth was observed, for hepatoma 46 a value of the DNA content proved to be near to that of normal liver. Moreover, in a regenerating liver the DNA content is the same as normal, that indicates the difference between normal and neoplastic actively proliferating tissues. The processes of DNA synthesis, studied by thymidine-C14 incorporation, showed a linear correlation with the rate of growth in regenerating, normal and neoplastic tissues. The ratio RNA/DNA in tumors is regularly decreased with respect to normal liver (a linear correlation between RNA/DNA and the maximum rate of growth). In a regenerating liver the ratio is high. Variations in the ratio RNA/DNA reflect changes in the functional activity and related disorders in the metabolism of nucleic acids in hepatomas. There was found a suppressed DNA decomposition in autolysis of homogenates from hepatoma tissues. It is essential to note that in a minimum deviated hepatoma some inhibition of the DNA decomposition is the only observed disorder in the nucleic acids metabolism. Spin-labelled DNA preparations of tissues under study were obtained. The presence of structural differences between separate DNA samples has been demonstrated.     

Effect of testosterone and estradiol-17beta on synthesis of DNA, RNA and protein in human breast in organ culture.

Effects of testosterone (T) and estradiol-17beta (E-2) on the synthesis of DNA, RNA and protein were studied in explants of the following types of malignant and non-malignant human female breast grown in organ culture: cystic mastitis (7 cases), fibroadenoma (8), carcinoma (17) and uninvolved tissue from cancer-bearing breast (8). In cultures of systic mastitis T uniformly inhibited the incorporation of thymidine-3H into DNA, uridine-3H into RNA and L-amino acids-14C (AA-14C) into protein. E-2 inhibited the incorporation of thymidine-3H, but had a variable effect on the incorporation of uridine-3H and AA-14C. Cultures of fibroadenoma, with no steroid added, were highly proliferative especially at the outer surface, but the incorporation of thymidine-3H, uridine-3H and AA-14C was much lower than in cystic mastitis. Whereas in fibroadenoma T uniformly inhibited the synthesis of DNA, RNA and protein, the effect of E-2 was variable: in most cases it inhibited the synthesis of DNA,but in one it stimulated it appreciably; in the majority of cases E-2 stimulated RNA and protein synthesis. In cultures of cancerous tissue T depressed, in most cases, the incorporation of thymidine-3H, uridine-3H and AA-14C, but it stimulated it in 4 out of 17. E-2 inhibited the synthesis of DNA, RNA and protein in 6 cases (2 of them were inhibited by T), stimulated them in 9 (one stimulated by T) and had no clear effect in 2. The effect of the steroids on the explants of uninvolved tissue was variable and did not always parallel their effect on the cancerous tissue from the respective patients.     

Inhibitory effects of anti-tumor platinum compounds on DNA, RNA and protein syntheses in mammalian cells in virtro

The inhibitory effects of anti-tumor, bacterial filament forming platinum compounds, Cis-Pt(II) (NH₃)₂Cl₂ (A), Cis-Pt(IV) (NH₃)₂Cl₄ (B), and Pt(II) (NH₂)₂(CH₂)₂Cl₂(C) on DNA, RNA and protein syntheses was measured by the incorporation of ³H-thymidine, ³H-uridine, and ³H-L-leucine, respectively, into an acid-insoluble polymer in human amnion AV₃ cells. Compound A, the most effective tumor-inhibiting platinum compound, was shown to selectively inhibit DNA synthesis below 5μM and to inhibit ³H-thymidine incorporation more rapidly than ³H-uridine or ³H-leucine incorporation at 25μM. Like A, compounds B and C were also shown to inhibit all three processes after a 24-hour period of treatment at 25μM. A correlation was established between the relative anti-tumor effectiveness of compounds A, B, and C and the extent of their inhibitory effects. On the other hand, two non-tumor-inhibiting platinum compounds, [Pt(II) (NH₃)₄]Cl₂ (D) and Trans-Pt(IV) (NH₃)₂Cl₄ (E) had no inhibitory effects, except compound E, which exhibited a rapid initial inhibition of ³H-L-leucine incorporation at 100μM. The inhibition of ³H-thymidine incorporation into cells pretreated 4 h with 5μM of compounds A, B, and C was shown to be irreversible. Finally the inhibition of the incorporation of ³H-thymidine into DNA by A was shown not to be caused by the inhibition of the uptake of the tracer into the acid-soluble pool. A number of possible explanations for the greater sensitivity of DNA synthesis and the sequential inhibition of DNA, RNA and protein synthesis at higher concentrations are suggested.     

Effects of luzopeptins on protein B23 translocation and ribosomal RNA synthesis in HeLa cells

Localization of protein B23 in HeLa cells after treatment with luzopeptin A and its analogues was studied using indirect immunofluorescence. Bright nucleolar fluorescence was observed in control HeLa cells. After treatment with luzopeptin A (50 ng/ml), luzopeptin B (500 ng/ml), or luzopeptin D (10 ng/ml) for 2 h, uniform nucleoplasmic rather than specific nucleolar fluorescence was observed. Luzopeptin C had no effect on protein B23 translocation. Luzopeptin D, A, and B inhibited [3H]uridine incorporation into the trichloroacetic acid insoluble fraction of HeLa cells with 50% inhibitory concentration values of 3.7 +/- 1.1 (SD), 10.8 +/- 2.1, and 122.0 +/- 34.0 ng/ml, respectively. Less than 10% inhibition of [3H]uridine incorporation was found with luzopeptin C (500 ng/ml and 2 h incubation). Ribosomal RNAs (28 and 18S) were isolated from HeLa cells treated with luzopeptin D (50 ng/ml; 2 h). They were then separated and analyzed in 1% agarose gel electrophoresis. There were 90.1 +/- 1.38 and 95.0 +/- 1.04% inhibition of [3H]uridine incorporation into 28 and 18S ribosomal RNA, respectively. The order of potency for the loss of nucleolar fluorescence and the concurrent increase in nucleoplasmic fluorescence was luzopeptin D greater than luzopeptin A greater than luzopeptin B much greater than luzopeptin C, which correlates with the order of their 50% inhibitory concentration values for inhibition of [3H]uridine incorporation. With 34-55% inhibition of RNA synthesis, both nuclear and nucleolar B23 immunofluorescence were observed. With 70-85% inhibition of RNA synthesis, a uniform nucleoplasmic fluorescence was observed. These results indicate that translocation of protein B23 as observed by indirect immunofluorescence may be a rapid and simple screening test for the selection of antitumor agents which inhibit ribosomal RNA synthesis.     

Metabolic alterations of liver regeneration. 8. Enhanced synthesis of DNA in the liver of 5-azacytidine-treated rats subjected to partial hepatectomy

Following 5-azacytidine, the incorporation of thymidine into DNA in regenerating rat liver is modified depending on the time of drug administration. When given shortly before or after partial hepatectomy, 5-azacytidine leads to the inhibition of DNA synthesis, while its administration 16–40 h prior to hepatectomy results in its enhancement in 24-h regenerating liver. The observed alterations in the synthesis of DNA are in agreement with the formation of thymidine 5′-triphosphate in cell-free liver extract. The significance of altered DNA synthesis and cytologic changes as revealed with autoradiography is discussed.