Membrane permeabilization by trypanosome lytic factor, a cytolytic human high density lipoprotein.

Trypanosome lytic factor (TLF) is a subclass of human high density lipoprotein (HDL) that mediates an innate immune killing of certain mammalian trypanosomes, most notably Trypanosoma brucei brucei, the causative agent of a wasting disease in cattle. Mechanistically, killing is initiated in the lysosome of the target trypanosome where the acidic pH facilitates a membrane-disrupting activity by TLF. Here we utilize a model liposome system to characterize the membrane binding and permeabilizing activity of TLF and its protein constituents, haptoglobin-related protein (Hpr), apolipoprotein L-1 (apoL-1), and apolipoprotein A-1 (apoA-1). We show that TLF efficiently binds and permeabilizes unilamellar liposomes at lysosomal pH, whereas non-lytic human HDL exhibits inefficient permeabilizing activity. Purified, delipidated Hpr and apoL-1 both efficiently permeabilize lipid bilayers at low pH. Trypanosome lytic factor, apoL-1, and apoA-1 exhibit specificity for anionic membranes, whereas Hpr permeabilizes both anionic and zwitterionic membranes. Analysis of the relative particle sizes of susceptible liposomes reveals distinctly different membrane-active behavior for native TLF and the delipidated protein components. We propose that lysosomal membrane damage in TLF-susceptible trypanosomes is initiated by the stable association of the TLF particle with the lysosomal membrane and that this is a property unique to this subclass of human HDL.

Effects of acute bovine trypanosomosis (Trypanosoma vivax) on plasma kinetics of intravenously administered lactose.

Four calves infected with Trypanosoma vivax and four uninfected control calves were each injected intravenously with repeated doses of 0.5 g lactose kg-1 body weight, thrice daily at intervals of 4 h. Plasma samples were collected at specified time intervals and analysed for lactose. Pharmacokinetic parameters were calculated from the data. T. vivax infection delayed excretion of lactose from the body, thus leading to significantly (P < 0.001) increased biological half life (t1/2) and a significantly (P < 0.001) reduced elimination rate constant for lactose in the body. The apparent volume of distribution and total clearance of lactose were not affected by the infection. T. vivax infection also appeared to cause accumulation of lactose in the plasma after repeated intravenous administration.

Bioavailability, pharmacokinetics, and tissue distribution of 14C homidium after parenteral administration to Boran cattle.

The absorption, distribution and elimination characteristics of 14C homidium have been described in non-infected and Trypanosoma congolense-infected cattle treated with 14C homidium chloride by either intramuscular (i.m.) or intravenous (i.v.) injection at a dose level of 1 mg/kg body weight. Results show that the mean (+/-SD) elimination of the drug from plasma followed a biexponential process, with half-lives of 0.084 +/- 0.006 h and 97.66 +/- 16.28 h for the distribution and elimination phases after intravenous injection, respectively. Bioavailability of the intramuscular dose was 62.5% and 57.8% in non-infected and trypanosome-infected cattle, respectively. Absorption was rapid, with a tmax of 15 min and a mean Cmax (+/-SD) of 268.4 +/- 4.09 ng/mL following the intramuscular dose in non-infected cattle. The major route of excretion was via faeces. Approximately 90% of the total dose given to non-infected i.m.-treated cattle was excreted within 14 days. Following intramuscular administration of the drug, residues remained high in the major excretory organs, with the liver having concentrations of 1411 and 1199 ng/g after 14 and 28 days, respectively. Over the same period, the values in the kidneys were 649 and 448 ng/g. Concentrations in the liver 14 and 21 days following i.v. treatment were 2195 and 2454 ng/g, respectively. These results show that there was no significant difference in liver drug residues between 14 and 21 days, or 28 days depending on the treatment given, suggesting that once the drug is in this organ, it is released back into the circulation at an extremely slow rate.

Disposition of diminazene aceturate (Berenil) in trypanosome-infected pregnant and lactating cows.

Three cows were repeatedly infected with different strains of Trypanosoma congolense and treated intramuscularly each time with a different dose of diminazene aceturate (Berenil). Biphasic decline was observed of the maximal plasma drug levels, which were attained at 15 min after the first treatment and at 30 min after the second and third treatments. The rate constants for the distribution and terminal phases depended on the period of exposure to parasitaemia of the animal at the time of treatment. Maximal diminazene aceturate residue levels were found in milk 8 h post treatment and declined biexponentially to 4.56 ng ml-1 and 8.76 ng ml-1 at 21 days post treatment after 3.5 mg kg-1 and 7.0 mg kg-1 doses, respectively. In the three cows, higher drug residues were found in the kidney (7.04, 3.92 and 7.99 micrograms g-1) than in liver (3.26, 2.87 and 1.24 micrograms g-1) and heart (1.79, 1.25 and 1.03 micrograms g-1). The results of this study indicate that the level of parasitaemia (degree of anaemia) in the animal at the time of treatment affects the distribution, disposition and elimination of diminazene aceturate in the animal. Furthermore, the residue level in milk after treatment depends on the treatment dose and could easily be bioavailable to the consumer.

Comparative pharmacokinetics of diminazene in noninfected Boran (Bos indicus) cattle and Boran cattle infected with Trypanosoma congolense.

The pharmacokinetics of diminazene in five female Boran (Bos indicus) cattle before and then during acute and chronic phases of experimental infections with Trypanosoma congolense were investigated. A 7.0% (wt/vol) solution of diminazene aceturate (Berenil) was used in all three phases of the study and administered as a single intramuscular dose of 3.5 mg of diminazene base per kg of body weight. There were no significant differences between the values of pharmacokinetic parameters for the noninfected cattle and the values for cattle with a chronic T. congolense infection. However, the maximum concentration of the drug in plasma during the acute phase of infection (8.25 +/- 1.72 micrograms/ml) was significantly (P < 0.01) greater than that during chronic infection (5.04 +/- 0.26 micrograms/ml) and that in the noninfected state (4.76 +/- 0.76 micrograms/ml). Similarly, the time to maximum concentration of the drug in plasma when diminazene was administered during the acute phase of infection (18.00 +/- 6.71 min) was significantly (P < 0.02) shorter than that for noninfected cattle (36.00 +/- 8.22 min) and that during chronic infection (33.75 +/- 7.50 min). The volume of distribution at steady state during acute infection (1.01 +/- 0.31 liter/kg) was significantly (P < 0.01) smaller than that in the noninfected state (1.37 +/- 0.17 liter/kg) and that in chronic infection (1.51 +/- 0.24 liter/kg). Eight hours after the drug had been administered, the concentration-time data profiles for each of the three study phases were very similar. Mean concentrations of diminazene in plasma 48 h after administration of the drug were 0.43 +/- 0.07 microgram/ml in noninfected cattle, 0.43 +/- 0.11 microgram/ml during the acute phase of trypanosome infection, and 0.44 +/- 0.09 microgram/ml during the chronic phase of the infection. Results of the present study indicate that the area under the concentration-time curve for diminazene in trypanosome-infected cattle did not differ significantly for noninfected cattle. It, therefore, appears that the total amount of diminazene attained and maintained in the plasma of cattle is not significantly altered during infection with T. congolense.

Ethidium bromide: pharmacokinetics and efficacy against trypanosme infections in rabbits and calves.

[14C]ethidium bromide has been used to determine drug levels in tissues and body fluids of rabbits and calves following intramuscular injection. Uninfected and Trypanosoma brucei- or Trypanosoma congolense-infected animals were studied. Blood and tissue fluid level reached a maximum with 1 h and then fell rapidly; after 96 h 80-90% of the radioactivity injected had been excreted, approximately one third in urine and two thirds in faeces. By 1 h after injection of 1 mg [14C]ethidium/kg into a T. congolense-infected calf, 70-80% of the radioactivity in blood was found to be bound to trypanosomes. Doses of 1 or 10 mg/kg were found not to be curative for T congolense or T. brucei infections in rabbits: drug treatment resulted in a period of sub-patent parasitaemia which was always followed by a relapse. Examination of the prophylactic action of ethidium in rabbits showed that the drug extended the pre-patent period following trypanosome inoculation but provided no absolute protection. A period of "apparent' prophylaxis observed after drug treatment of infected rabbits has been correlated with the presence of anti-trypanosome IgG in the serum.

Experimental bovine trypanosomiasis. Changes in the catabolism of serum immunoglobulins and complement components in infected cattle.

The turnover of serum proteins of calves experimentally infected with Trypanosoma congolense was compared to that of normal uninfected cattle. All proteins examined had much increased catabolic rates in infected animals. In normal animals the average half-lives in days for each protein were: IgG1 17.4, IgG2 22.4, IgM 4.8, IgA 3.4, IgE 1.9, C1 5.6 and C3 2.9. In trypanosome infected cattle the average half-lives were IgG1 1.9, IgG2 1.7, IgM 0.9, IgA 1.2, IgE 0.9, C1 1.2 and C3 1.1 days.