Terry L. Miller

Effect of hexachlorophene on monovalent cation transport in human erythrocytes a mechanism for hexachlorophene-induced hemolysis

Abstract Hexachlorophene-induced hemolysis, as studied by phase contrast microscopy, appeared to be a result of osmotic swelling. Both swelling and subsequent hemolysis were markedly delayed by addition of the non-penetrating solute sucrose to the incubation mixture. Binding studies indicated that hexachlorophene is associated primarily with the erythrocyte membrane, the remainder being found in the cytoplasm. Hexachlorophane induced a dose-dependent, first-order efflux of Na+ and K+ from red cells. The rates of hemolysis and K+ efflux induced by hexachlorophene were much greater than would be expected if this compound were acting simply as a metabolic inhibitor and/or an inhibitor of (Na+-K+-Mg2+)-ATPase. It is suggested that hexachlorophene induces the efflux of Na+ and K+ from red cells by directly altering the permeability of the cellular membrane. Further, hexachlorophene-induced hemolysis is probably a secondary event resulting from osmotic swelling subsequent to increased membrane permeability.

Effect of hydroxychlorodiphenyl ethers (chlorinated pre-and isopredioxins) on erythrocyte membrane adenosinetriphosphatase activity.

The effect of hydroxychlorodiphenyl ethers (HO-ClX-DPEs; chlorinated pre- and isopredibenzodioxins), contaminants of technical chlorophenol preparations, on human erythrocyte membrane-bound adenosinetriphosphatases (ATPases) has been investigated. Both 2- and 3-HO-Cl9-DPE inhibited the Na+ + K+-activated, Mg2+-dependent ATPase (Na+, K+, Mg2+-ATPase). The Mg2+-dependent ATPase (Mg2+-ATPase) was stimulated at lower concentration of these compounds, but at higher concentrations there was a gradual decrease in the extent of stimulation, 2-Hydroxy-21, 41, 41-trichlorodiphenyl ether (2-HO-Cl3-DPE; Irgasan DP-300; Triclosan) was not as effective as the nonachloro compounds at inhibiting Na+, K+, Mg2+-ATPase and was inactive at stimulating Mg2+-ATPase. Pure pentachlorophenol (PCP) caused both inhibition of Na+, K+, Mg2+-ATPase and stimulation of Mg2+-ATPase, although each effect required a higher concentration of PCP than was needed for the HO-CL9-DPEs. The possible relationship of the effects of HO-CL x-DPEs on human erythrocyte membrane ATPase activities to the potent hemolytic activity of these compounds is discussed.

Chlordecone impairs Na(+)-stimulated L-[3H]glutamate transport and mobility of 16-doxyl stearate in rat liver plasma membrane vesicles.

Chlordecone (CD) treatment of rat liver plasma membranes (LPM) provided in vitro evidence for mechanisms of in vivo liver dysfunction caused by CD. LPM preparations enriched 14- to 19-fold in the bile canalicular markers gamma-glutamyl transpeptidase, alkaline phosphatase, and leucine aminopeptidase were isolated from male Sprague-Dawley rats. CD inhibited the bile canalicular-specific active transport of Na(+)-stimulated L-[3H]glutamate in LPM vesicles. CD (0.08 and 0.5 mumol/mg protein) reduced both the initial velocity and the maximum level of Na(+)-stimulated L-[3H]glutamate uptake without significantly reducing Na(+)-independent uptake. In vitro treatment of LPM with CD (0.2-1.0 mumols/mg protein) also reduced the mobility of a 16-doxyl stearate spin label probe in a concentration-dependent manner. No change in mobility was apparent at CD concentrations below 0.2 mumol/mg protein. These results demonstrated that CD impaired a bile canalicular-specific transport system and induced liver plasma membrane perturbation. Na(+)-stimulated L-[3H]glutamate uptake was more sensitive to CD than was detectable immobilization of the spin label probe.

Thermotropic properties of human erythrocyte membrane proteins as affected by hydroxychloroaromatic compounds

The thermal stability of the anion transport protein (band 3) and other proteins of the human erythrocyte membrane, as influenced by hydroxychloroaromatic (HO-Cl2-Ar) compounds, was studied by differential scanning calorimetry. Various hydroxychlorodiphenyl ethers (HO-Clx-DPEs) and hexachlorophene, but not pentachlorophenol, caused a marked decrease in the thermal stability of band 3. Most of the other calorimetric transitions of the erythrocyte membrane were only slightly affected. The activity of HO-Clx-DPEs toward lowering the transition temperature of band 3 generally increased with the degree of chlorination, and was somewhat dependent on the position of hydroxyl substitution. At higher concentrations of HO-Clx-DPEs, there was a decrease in the enthalpy change and a broadening of the endothermic transition of band 3. The order of effectiveness of these compounds, as determined from band 3 denaturation temperatures, was similar to the order of potency previously observed for hemolysis of human erythrocytes.