Acute metabolic and hormonal effects of intravenously administered sodium n-butyrate in untreated and alloxan-diabetic sheep.

Experimental diabetes was induced in 4 wethers of the Mutton Merino breed by intravenous injection of alloxan (75 mg.kg-1) in order to determine its impact on plasma glucose, immunoreactive insulin, free fatty acids (FFA), cholesterol, phospholipids, triglycerides, D-(-)-3-hydroxybutyrate (D-3-HB), bilirubin and aspartate aminotransferase (ASAT) as well as on the changes of these parameters brought about by an intravenous infusion of sodium n-butyrate (1 mmol.kg-1). Alloxan administration caused a significant elevation of plasma glucose, FFA, triglycerides, cholesterol, phospholipids, D-3-HB and bilirubin and a decrease of the level of immunoreactive insulin. The increase in glucose level brought about by a bolus injection of sodium n-butyrate in untreated sheep did not appear in alloxanized animals. Thus, it is suggested that the lack of hyperglycaemic response in diabetic sheep was due to the absence of liver glycogen stores. Unexpectedly in alloxan-diabetic sheep, a decrease in the plasma level of FFA occurred after the administration of sodium n-butyrate. Therefore, it may be assumed that beside insulin other factors may contribute to the decrease of FFA under these conditions.

The inhibitory action of sodium butyrate on the growth of KB, MMT and RPMI cells.

Three tumor cell lines (KB, MMT and RPMI) established from epithelial tissues were treated for 24 h with sodium butyrate (BU), the BU concentrations giving rise to 50% inhibition of [3H]thymidine incorporation were 2.0, 0.3, 0.2 mmol/L, respectively, for the KB, MMT and RPMI cell lines. Studies with [14C]BU have shown that, at similar degrees of inhibition of [3H]thymidine incorporation, the intracellular concentrations of BU are very close for all three cell types, despite the dissimilarity of the extracellular BU concentrations. These results imply that the BU sensitivity of the cells does not depend on the inhibition of thymidine incorporation, but on their BU uptake. The [3H]thymidine incorporation of MMT cells exposed to 5 mmol/L BU for 72 h returned to normal within the next 48 h. The same treatment accounted for about an 80-90% decrease in the cloning efficiency and tumourigenicity of MMT cells. These findings indicate that BU pretreatment inhibits DNA synthesis temporarily, while other parameters related to the cell growth, such as cloning efficiency and tumourigenicity, are durably influenced by BU pretreatment.

Effects of butyrate and insulin and their interaction on the DNA synthesis of rumen epithelial cells in culture.

Rumen epithelial cells (REC) were incubated in the presence of various concentrations of butyrate or insulin or with both of them, to obtain information on their effect on the DNA synthesis of cultured cells. The 24-h values of 3H-thymidine incorporation into cellular DNA were measured in the presence of butyrate, insulin or butyrate plus insulin. While butyrate reduced DNA synthesis, insulin produced an increase over the control. Combined butyrate plus insulin treatment influenced the incorporation of label in accordance with the relative proportion of these two substances.

Effect of intraruminal butyrate infusion on the plasma insulin level in sheep.

After intraruminal infusion of butyrate to sheep at dose rates of 0.25, 0.5, 1 and 2 g sodium n-butyrate per kg body mass, butyrate concentration of the rumen fluid and total secreted insulin rose in direct proportion to the butyrate dose infused. The half-life of butyrate in the rumen was always longer than that of insulin. At 90 min after the infusion of 1 g butyrate per kg body mass, butyrate concentration in the ruminal papillae reached the level corresponding to an extracellular concentration that reduced cell division by 50% in vitro. It can be concluded that butyrate may be present in the ruminal papillae in concentrations inhibiting cell proliferation, simultaneously with the presence of blood plasma insulin concentrations stimulating the proliferation of ruminal epithelial cells.

The effects of butyrate and glucagon on the proliferation of ruminal epithelial cells in culture.

When the rate of ruminal epithelial cell proliferation was measured on the basis of 3H-thymidine incorporation into the cellular DNA, butyrate dose-dependently reduced 3H-thymidine incorporation. In contrast, glucagon at 10 and 100 pg/ml had a slight stimulatory effect on the incorporation, but only in the absence of butyrate.

In vitro evaluation of protein degradability in the rumen and digestibility of undegraded protein.

A three-phase laboratory procedure suitable for predicting protein degradability in the rumen and digestibility of undegraded protein is reported. In the first phase the feed was incubated with starch and buffered rumen fluid. In the incubation mixture the viability of protease-active bacteria was checked by anaerobic culturing, whereas changes in protease activity were monitored by azocasein degradation. In the second and third phase rumen undegradable protein (UDP) was digested with pepsin and pancreatin, respectively. The measurements showed that 63.2, 5.2 and 4.7% of the crude protein of green lucerne was decomposed by rumen fluid, pepsin and pancreatin, respectively. Degradability of the crude protein of extracted sunflower meal was 68.3, 17.7 and 5.5% in the three phases, respectively. Repeated determination yielded crude protein degradabilities of 66.7, 27.1 and 5.1% for the three phases, respectively.

Effect of heat treatment and subsequent urea supplementation of sunflower meal on the in vitro ruminal degradability of crude protein content and its postruminal digestibility.

A three-phase laboratory procedure was used for predicting the degradability of the protein of extracted sunflower meal before (SFM) and after heat treatment (HSFM). The rumen fluid degraded 69.1% and 67.1% of the SFM and HSFM protein, respectively. The digestibility values of rumen undegraded protein (UDP) were 57.3% (SFM) and 57.0% (HSFM) with pepsin and 17.6% (SFM) and 15.9% (HSFM) with pancreatin. Urea supplementation practically did not alter the rumen degradability of HSFM protein, while the pepsin digestibility of UDP decreased to 47.2%. Four fractions (NH3, dissolved amino acids, oligopeptides and proteins) of rumen degradable crude protein (RDP) were also determined in vitro: 81 to 92% of the degraded crude protein was found in the fractions tested. Heat treatment reduced free NH3 content but did not alter the other three fractions. Urea supplementation decreased the quantity of NH3, peptides and proteins as well.

Influence of sodium butyrate on HeLa cell morphology and proliferation.

The effect of one-week exposure to sodium butyrate on HeLa S3 cell cultures was studied with special regard to influence on prekeratin synthesis, by comparison to cultures similarly treated with the known proliferation inhibitor hydroxyurea, and not treated. Like hydroxyurea, sodium butyrate inhibited cell proliferation to a considerable degree, but accounted additionally for an increase in membrane-bound alkaline phosphatase activity, cellular prekeratin synthesis, tonofilament number, and filament bundle formation. These phenomena unequivocally indicate that sodium butyrate acted as a specific stimulator of Hela (epithelial) cell differentiation. Similar differentiation phenomena can be observed during early spontaneous keratinization of the stratified horny epithelium.

Immunohistochemical detection of bovine ruminal carbonic anhydrase isoenzyme.

Rabbits immunized with low-activity ruminal carbonic anhydrase (RCA) isoenzyme, extracted from ruminal epithelial cells isolated by digestion with trypsin, yielded anti-RCA sera which reacted specifically with bovine RCA in double agar gel diffusion and immunoelectrophoretic tests, but failed to cross-react with bovine erythrocyte CA. The localization of RCA was identified in histological sections and isolated ruminal epithelial cell preparations by indirect immunofluorescence and immunoperoxidase tests as the basal, spinosum and granulosum layers of ruminal mucous epithelium.

Culture of epithelial cells from bovine ruminal mucosa.

A method is reported for the primary in vitro culture of epithelial cells derived from bovine ruminal mucosa. That the cultures of ruminal epithelial cells consisted exclusively of stratum spinosum, stratum basale and stratum granulosum was confirmed by immunoperoxidase and immunofluorescence staining using carbonic anhydrase isoenzyme as a marker.