Reversible effects of sodium butyrate on the differentiation of F9 embryonal carcinoma cells

We have studied effects of sodium butyrate on embryonal carcinoma F9 cell differentiation. In the presence of sodium butyrate, F9 cells underwent rapid and drastic morphological changes and expressed marked increases in mRNA levels of various differentiation markers. When sodium butyrate was removed from the cultures, all the examined phenotypes of F9 cell differentiation rapidly reverted to the characteristics of undifferentiated stem cells. However, under the same conditions, when cycloheximide or actinomycin D was added to the cultures, such phenotypic reversion was not observed, but high mRNA levels of the differentiation markers as well as altered cell morphology were retained. These results indicated that the effects of sodium butyrate on induction of teratocarcinoma cell differentiation were reversible and that de novo syntheses of some mRNA(s) and protein(s) were necessary for the reversion of differentiated cells to stem cells.     

Evaluation of the antioxidative and genotoxic effects of sodium butyrate on breast cancer cells

Oncogenic stimulation shows a rise in reactive oxygen species (ROS), and ROS can eventually induce carcinogenesis by causing DNA damage. In this context, this study aims to evaluate some biochemical and genotoxic changes in the control of cell death caused by NaBu (Sodium butyrate). treatment in breast cancer cells. NaBu’s impact on cell proliferation was determined via WST-1 assay. The lipid peroxidation (MDA), reduced glutathione (GSH), Nitric Oxide (NO), hydrogen peroxide (H₂O₂), and superoxide dismutase (SOD) enzyme levels were determined biochemically. NaBu-induced genotoxic damage was estimated via single-cell gel electrophoresis (SCGE). NaBu reduced cell viability and potentially induced GSH, but decreased SOD enzyme activity and the level of MDA and NO decreased also H₂O₂ decreased at different times and NaBu concentrations. Higher NaBu concentrations amplified DNA damage in MCF-7 cells compared to the control group. NaBu shows anticancer and genotoxic effects, especially through antioxidant enzymes, one of the oxidative stress parameters in breast cancer. However, the anticancer and genotoxic effects of NaBu is changed in the oxidative stress parameters with time and treatment concentration of NaBu in MCF-7 cells. Furthermore, his oxidative stress-dependent effect changes need to be clarified by further evaluation with molecular and more biochemical parameters.     

Differential effects of sodium butyrate and hyperosmolality on the modulation of alkaline phosphatases of LoVo cells

LoVo cells produce term-placental and intestinal alkaline phosphatases. Hyperosmolality and sodium butyrate increase the levels of both, but the effect of sodium butyrate is more pronounced on the intestinal enzyme. When applied together, induction of term-placental alkaline phosphatase is additive and that of the intestinal enzyme is synergistic. Induction by either stimulus or by their mixture is independent of cell density. However, whereas the effect of hyperosmolality is readily reversible, induction by sodium butyrate is not. No synergistic increase in intestinal alkaline phosphatase activity occurs when cells are sequentially treated with hyperosmolality and sodium butyrate or vice versa. This indicates that only when applied concurrently does one inducer amplify the effect of the other. Since the normal colonic mucosa produces intestinal alkaline phosphatase, its predominant induction by sodium butyrate in LoVo cells may reflect a more differentiated state.     

The expression of glucose regulated protein—94 in colorectal carcinoma cells treated by sodium butyrate

The expression of glucose regulated protein 94 (GPR94) during the treatment of human colorectal carcinoma cell lineClone A cells with codium butyrate was studied.Dodium butyrate (SB) can cause functional and morphological effects on Clone A cells including growth arrest at G0/G1 stage and cell differentiation as observed by morphological changes,MTT and flow cytometry assays,as well as reduced Grp94 gene expression as shown by Northern blot and Western blot assays.The possible mechanism of the correlation between Grp94 gene expression and tumor growth inhibition and cell differentiation is briefly discussed.     

Cytotoxic and apoptotic effects of menadione on rat hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is one of the most common cancers, which may lead to death. Menadione shows cytotoxic activity thought affecting redox cycling in cancer cells. The aim of the present study was to investigate the effects of menadione on rat hepatocellular carcinoma (H4IIE) cell morphology, cytotoxicity, apoptosis and DNA damage or repair in vitro. Cell morphology evaluated by microscopy and cell viability was determined using the 3-[4,5-dimethylthiazol-2yl]-diphenyltetrazolium bromide test. Apoptotic cell death was assessed in H4IIE cells treated with menadione by 4′,6-diamidino-2-phenylindole staining. Quantitative real time polymerase chain reaction used to determine the expression level of poly (ADP-ribose) polymerase 1 (PARP1) gene. According to the results of this study menadione has got a cytotoxic activity (IC₅₀ 25 µM) and change the cell fate in H4IIE cells. Menadione treatments lead to PARP1 activation in a dose dependent manner and induce DNA damage and apoptosis, and this may suggest its use as a therapeutic agent in HCC treatment.     

Biochemical and morphological effects of sodium butyrate on Dictyostelium discoideum development

Pretreatment of proliferating D. discoideum amoebae with 10 mM butyrate for at least 8 h (one duplicating time) induced a reversible and dose dependent premature expression of several developmental parameters when the cells were starved in the absence of the fatty acid. The aggregative phase of the morphogenetic cycle was reduced in 2 h and the appearance of mature fruiting bodies and spores took place 4 h earlier as a result of butyrate pretreatment. Some developmentally regulated proteins, such as contact-sites A, cell surface lectins and cyclic AMP phosphodiesterase were also expressed 2 h earlier in butyrate pretreated cells than in controls. The level of extracellular cyclic AMP was reduced in butyrate pretreated cells, while other parameters of cyclic AMP metabolism were not affected. Butyrate also caused a partial inhibition of growth and the hyperacetylation of histone H4 in growing amoeba. These results suggest that butyrate acts as an inducer of differentiation in D. discoideum and can therefore be used as an experimental tool in order to explore regulatory mechanisms operating in slime mold differentiation.Abbreviations MES 2-N-morpholinoethanesulfonate - EDTA ethylendiaminotetracetate - TCA trichloroacetate - DTT dithiothreitol - SDS sodium dodecylsulfate     

Multiple effects of 5 mM sodium butyrate on Physarum polycephalum macroplasmodia

Summary In a Physarum polycephalum macroplasmodium, nuclei naturally divide synchronously. Thus, it offers an opportunity to study growth and mitosis within a true organism. The effects of 5 mM sodium-butyrate on these processes have been examined. When this material is added to the culture medium during mitosis, the butyrate acts like a fixative on condensed chromosomes. During interphase, this short fatty acid stops growth and immediately inhibits DNA synthesis. Furthermore, it prevents differentiation in macroplasmodia induced to spherulate. All these modifications are readily reversible after transfer to a medium lacking butyrate.     

Flow cytometric and biochemical analysis of dose-dependent effects of sodium butyrate on human endometrial adenocarcinoma cells.

Sodium butyrate (SB) treatment was previously shown to produce seven-fold increases in estrogen hormone receptor binding sites of human endometrial adenocarcinoma (IK) cells. Flow cytometric analysis and histone gel electrophoresis were used to examine cell cycle, cell metabolism, and nuclear histone fractions in IK cells treated with different concentrations of SB. SB-treated cells stained with fluorochromes specific for DNA, RNA, or general protein were analyzed by flow cytometry (FCM). Changes in accessibility to three DNA stains and gel electrophoresis were used to analyze rearrangements in chromatin structure. SB caused an accumulation of cells in the G1 phase and inhibited DNA synthesis, but not cellular levels of RNA and protein. Hoechst accessibility to A-T rich regions on DNA was dramatically increased after removal of SB. H1 histones were dephosphorylated and core histones were acetylated during SB-treatment. Information obtained in these studies may be useful for correlating cellular and biochemical events with SB-induced increases in nuclear steroid hormone binding sites.     

Differential effect of sodium butyrate on cyclic AMP phosphodiesterase activities in butyrate sensitive and resistant mastocytoma cells

The effect of sodium butyrate on the intracellular cyclic AMP levels and the activities of cyclic AMP-regulating enzymes were examined in two types of mastocytoma p-815 cells in culture: one type (S cell) was sensitive and the other (R cell) was resistant to the induction of differentiation by sodium butyrate. In the presence of sodium butyrate, adenylate cyclase activity increased in both S and R cells to the same degree, whereas the level of cyclic AMP was elevated only in S cells. Cyclic AMP phosphodiesterase activity increased in R cells but not in S cells. Cyclic AMP phosphodiesterase activities of two cell populations differed in their response to sodium butyrate and they seem to have an important role in regulating cellular level of cyclic AMP that might be an important factor in controlling cell differentiation.     

Effect of sodium butyrate on mammalian cells in culture: A review

Summary Sodium butyrate produces reversible changes in morphology, growth rate, and enzyme activities of several mammalian cell types in culture. Some of these changes are similar to those produced by agents which increase the intracellular level of adenosine 3′,5′-cyclic monophosphate (cAMP) or by analogs of cAMP. Sodium butyrate increases the intracellular level of cAMP by about two fold in neuroblastoma cells; therefore, some of the effects of sodium butyrate on these cells may in part be mediated by cAMP. Sodium butyrate appears to have properties of a good chemotherapeutic agent for neuroblastoma tumors because the treatment of neuroblastoma cells in culture causes cell death and “differentiation”; however, it is either innocuous or produces reversible morphological and biochemical alterations in other cell types.     

Synergistic antitumor effects of rhein and doxorubicin in hepatocellular carcinoma cells

The aim of this study was to investigate the synergistic antitumor activity of rhein and doxorubicin (DOX) and to elucidate the underlying mechanisms in hepatocellular SMMC-7721 and HepG2 cells. Cell growth curves, caspase-3 activity, and intracellular DOX accumulation were observed using an IncuCyte real-time video imaging system. Combination index was used to calculate synergistic potential of rhein and DOX. Cell apoptosis was detected by the Annexin V-FITC/PI apoptosis kit. Lactate dehydrogenase and adenosine triphosphate (ATP) levels were assessed using an assay kit. Oxygen consumption rates (OCR) and extracellular acidification rates were assessed by the Seahorse XFe96 Extracellular Flux Analyzer. Mitochondrial inner membrane potential (ΔΨm) was monitored with JC-1 fluorescence. Western blot analysis was used to detect the level of P-glycoprotein. Synergistic antiproliferative and proapoptotic effects were exerted by the combination of rhein at 10 μM and DOX at 2 μM in SMMC-7721 and HepG2 cells. Rhein could influenced the accumulation of DOX in both cells, which was associated with remarkably decreased mitochondrial energy metabolism and ATP levels. Rhein could reduce ΔΨm in both cells. mPTP, opener atractyloside (ATR) could accelerate the loss of ΔΨm, and further suppress the OCR induced by rhein. In contrast, the mPTP blocker cyclosporin A (Cs A) inhibited the loss of ΔΨm and the OCR induced by rhein. Our data indicate that a decline in mitochondrial energy metabolism was responsible for the synergistic antitumor effects of rhein and DOX in hepatocellular carcinoma cells. Reduction of ΔΨm and opening of mPTP inhibited the exchange of ATP/adenosine diphosphate between mitochondrial matrix and cytoplasm is the important mechanism.     

Induction of carcinoembryonic-antigen-gene expression in human colorectal carcinoma by sodium butyrate.

The ability of the insulin-induced phospho-oligosaccharide to stimulate amino acid transport was studied in isolated rat hepatocytes. At low alpha-aminoisobutyric acid concentrations (0.1 mM), both 100 nM-insulin and 10 microM-phospho-oligosaccharide doubled amino acid uptake after 2 h of incubation. This stimulation was prevented by 0.1 mM-cycloheximide or 5 micrograms of actinomycin D/ml, indicating that the phospho-oligosaccharide, like insulin, was acting via the synthesis of a high-affinity transport component. The effects of the phospho-oligosaccharide and of insulin were blocked by Ins2P (2.5 mM), but not by myo-inositol, inositol hexaphosphoric acid or several monosaccharides such as mannose, glucosamine and galactose. Both the temporal effect on amino acid entry and the extent of stimulation of this process by the phospho-oligosaccharide indicate that this molecule mimics, and may mediate, some of the long-term actions of insulin. However, the effects of phospho-oligosaccharide and insulin were not exactly the same, since the effect of insulin, but not of the phospho-oligosaccharide, was additive with that of glucagon.     

Genetic networks responsive to sodium butyrate in colonic epithelial cells

We performed microarray and computational gene network analyses to identify the detailed mechanisms by which sodium butyrate (SB) induces cell growth arrest and the differentiation of mouse colonic epithelial MCE301 cells. Two thousand six hundred four differentially expressed probe sets were identified in the cells treated with 2mM SB and were classified into four groups. Of these, the gradually increased group and the gradually and remarkably decreased group contained the genetic networks for cellular development and cell cycles or canonical pathways for fatty acid biosynthesis and pyrimidine metabolism, respectively. The present results provide a basis for understanding the detailed molecular mechanisms of action of SB in colonic epithelial cells.     

Effect of sodium butyrate on autophagy and apoptosis in Chinese hamster ovary cells

Sodium butyrate (NaBu), which is widely used in recombinant Chinese hamster ovary cell (rCHO) cultures for high-level expression of therapeutic proteins, is known to induce apoptosis in a dose-dependent manner. Lately, the significance of autophagy has increased in the field of CHO cell culture due to the fact that autophagy is related to the programmed cell death mechanism. To determine the effect of NaBu on autophagy as well as apoptosis of rCHO cells, rCHO cells producing erythropoietin were subjected to NaBu treatment. NaBu treatment up to 5 mM increased cleaved forms of PARP, caspase-3, and Annexin V positive population, confirming the previous results that NaBu induces apoptosis. Concurrently, NaBu treatment increased the level of accumulation of the autophagic marker, LC3-II, independently of nutrient depletion, suggesting that NaBu induces autophagy. To elucidate the potential role of autophagy induced by NaBu, a representative autophagy inducer (rapamycin) or an inhibitor (bafilomycin A1) was added to cultures together with NaBu. It was found that autophagy had the potential role of a positive cell survival mechanism under NaBu treatment. Furthermore, gradual reduction in mitochondrial membrane potential/mass and recruitment of a mitophagy protein, Parkin, to the mitochondria were observed under NaBu treatment, suggesting that this positive function of autophagy might be mediated by the autophagic removal of damaged mitochondria. Taken together, autophagy was observed in rCHO cell culture under NaBu treatments and the results obtained here support the positive effects of autophagy induced by NaBu treatments.     

Effect of sodium butyrate on mammalian cells in culture: a review.

Sodium butyrate produces reversible changes in morphology, growth rate, and enzyme activities of several mammalian cell types in culture. Some of these changes are similar to those produced by agents which increase the intracellular level of adenosine 3',5'-cyclic monophosphate (cAMP) or by analogs of cAMP. Sodium butyrate increases the intracellular level of cAMP by about two fold in neuroblastoma cells; therefore, some of the effects of sodium butyrate on these cells may in part be mediated by cAMP. Sodium butyrate appears to have properties of a good chemotherapeutic agent for neuroblastoma tumors because the treatment of neuroblastoma cells in culture causes cell death and "differentiation"; however, it is either innocuous or produces reversible morphological and biochemical alterations in other cell types.