sodium-taurodeoxycholate and Hemolysis

This study was performed to compare the effects of two hydrophilic bile acids, taurohyodeoxycholic acid (THDCA) and tauroursodeoxycholic acid (TUDCA), on HepG2 cells. Cytotoxicity was evaluated at different times of exposure by incubating cells with increasing concentrations (50-800 micromol/l) of either bile acid, while their cytoprotective effect was tested in comparison with deoxycholic acid (DCA) (350 micromol/l and 750 micromol/l)-induced cytotoxicity. Culture media, harvested at the end of each incubation period, were analyzed to evaluate aspartate transaminase (AST), alanine transaminase and gamma-glutamyltranspeptidase release. In addition, the hemolytic effect of THDCA and TUDCA on human red blood cells was also determined. At 24 h of incubation neither THDCA nor TUDCA was cytotoxic at concentrations up to 200 and 400 micromol/l. At 800 micromol/l both THDCA and TUDCA induced a slight increase in AST release. At this concentration and with time of exposure prolonged up to 72 h, THDCA and TUDCA induced a progressive increase of AST release significantly (P<0.05) higher than that of controls being AST values for THDCA (2.97+/-0.88 time control value (tcv) at 48 h and 4.50+/-1.13 tcv at 72 h) significantly greater than those of TUDCA (1.50+/-0.20 tcv at 48 h and 1.80+/-0.43 tcv at 72 h) (P<0.01). In cytoprotection experiments, the addition of 50 micromol/l THDCA decreased only slightly (-5%) AST release induced by 350 micromol/l DCA, while the addition of 50 micromol/l TUDCA was significantly effective (-23%; P<0.05). Higher doses of THDCA or TUDCA did not reduce toxicity induced by 350 micromol/l DCA, but were much less toxic than an equimolar dose of DCA alone. At the concentration used in this experimental model neither THDCA nor TUDCA was hemolytic; however at a very high concentration (6 mmol/l) both bile acids induced 5-8% hemolysis. We conclude that bile acid molecules with a similar degree of hydrophilicity may show different cytotoxic and cytoprotective properties.

Topics: Alanine Transaminase; Aspartate Aminotransferases; Deoxycholic Acid; Dose-Response Relationship, Drug; Erythrocytes; Hemolysis; Humans; Taurochenodeoxycholic Acid; Taurodeoxycholic Acid; Time Factors; Transglutaminases; Tumor Cells, Cultured

The hemolytic activities of sodium deoxycholate (DChol) and its tauro-conjugate (TDChol) and glyco-conjugate (GDChol) were analysed. 50 % hemolysis occurred in 30 min at pH 7.3, at the concentrations of these detergents equal to 0.044, 0.042 and 0.040 % respectively. These values are below their critical micellar concentrations. Based on its kinetics, this hemolysis is classified as being of permeability type. The detergents increase the permeability of erythrocyte membranes to KCl, and colloid osmotic hemolysis occurs. The minimum of hemolytic activity of the three cholates is at about pH 7.5. A very high increase in hemolytic activity occurs at pHs below 6.8, 6.5 and 6.2 for DChol, TDChol, and GDChol, respectively. These values are close to the pK(a) for DChol (6.2), but much higher than the pK(a) for TDChol (1.9) and GDChol (4.8). It is therefore suggested that the increase in hemolytic activity is not a result of the protonation of the anionic groups of the cholates. At acidification below pH 6, the kinetics of DChol induced hemolysis change to the damage type characterised by nonselective membrane permeability. Such a transition is not observed in TDChol and GDChol induced hemolysis. It is therefore suggested that the change in the type of hemolysis depends on protonation of the anionic group of cholates.

Topics: Bile Acids and Salts; Cell Membrane Permeability; Deoxycholic Acid; Detergents; Erythrocyte Membrane; Glycodeoxycholic Acid; Hemolysis; Humans; Hydrogen-Ion Concentration; In Vitro Techniques; Kinetics; Taurodeoxycholic Acid

To evaluate why hemolysis of red blood cells (RBC) by bile acids varies in different mammalian species, we determined the mean corpuscular volume (MCV), lipid content and the concentrations of the conjugates of deoxycholate and of NaCl inducing 50% hemolysis of RBC from healthy humans, pigs, horses, cows, sheep and jaundiced humans. A volume of 0.05 mL of washed RBC at 1% hematocrit, which has the same lipid content but different phospholipid composition and number of erythrocytes (owing to the variable MCV), was incubated in taurodeoxycholate (TDC) solution (0-5 mM) to determine the TDC concentration inducing 50% hemolysis (TDC50). The TDC50 was highest in RBC of sheep and decreased within the series sheep > pig > cow > horse > healthy human > jaundiced human, which have generally increasing MCV. The osmotic resistance followed an inverse order, with jaundiced human > healthy human > horse > cow > pig > sheep. Although we found no correlation between the TDC50 and phospholipid composition of the erythrocytes tested, the extent of bile salt-induced hemolysis seemed to depend on both the MCV and the number of erythrocytes in the incubation medium.

Topics: Animals; Bile Acids and Salts; Cholestasis; Dose-Response Relationship, Drug; Erythrocyte Membrane; Erythrocyte Volume; Glycodeoxycholic Acid; Hemolysis; Horses; Humans; Osmotic Fragility; Phospholipids; Ruminants; Sodium Chloride; Sphingomyelins; Taurodeoxycholic Acid