S. Ghatak

Effect of Plasmodium yoelli nigeriensis infection and chloroquine on the hepatic mixed function oxidase system of mice

Plasmodium yoelii nigeriensis infection in albino mice significantly altered the hepatic microsomal mixed function oxidase system. Cytochrome P-450 (the terminal monooxygenase) and other monooxygenases, viz. aniline hydroxylase, aminopyrine-N-demethylase and benzo(a)pyrene hydroxylase were significantly lowered while microsomal heme showed 4-fold increase at 80% parasitaemia. Noticeable impairment in the other components like NADH:cytochrome b5 reductase, NADPH:cytochrome c reductase, cytochrome b5 and glucose-6-phosphatase was also observed. Oral treatment of normal and P. y. nigeriensis infected mice with chloroquine (64 mg per kg body weight for 4 days) caused lowering of mixed function oxidase activities which however showed a recovering trend, a week after cessation of treatment.

Biogenic amines, metabolites and monoamine oxidase in the filarial worm Setaria cervi

Analysis of biogenic monoamines and their metabolites in Setaria cervi adults by reverse phase high performance liquid chromatography (HPLC) revealed dopamine as the major monoamine followed by norepinephrine and 5-hydroxytryptamine (5-HT). 5-Hydroxy indole acetic acid and tryptophan were also detected in significant amounts. A particulate-bound monoamine oxidase (MAO, EC 1.4.3.4.) catalysing the oxidative deamination of several amines was also demonstrated in both microfilariae and adults. The enzyme from the parasites exhibited unusually high Km values for various monoamines. Dopamine was oxidized at the maximum rate while putrescine was not utilized as the substrate. MAO was predominantly associated with the mitochondrial fraction and concentrated mainly in the cuticle-muscle-hypodermis layer of the filariid. The enzyme was most active around pH 7.5 and 37 +/- 2 degrees C, relatively stable in the frozen state but was thermolabile. The specific MAO inhibitors, clorgyline and deprenyl, inhibited the enzyme with Ki values of 2 x 10(-7) M and 5 x 10(-6) M, respectively. Diethylcarbamazine, suramin, levamisole and centperazine significantly inhibited MAO activity. (The characteristics of the enzyme indicated that it may have a role in host-parasite interactions).

Polyamine metabolism in Setaria cervi, the bovine filarial worm*

Spermine and spermidine were found to be the principal polyamines in the bovine filarial parasiteSetaria cervi, whereas putrescine was observed in very low amounts. Studies conducted on the enzymes of polyamine biosynthesis revealed low activity for S-adenosyl-methionine decarboxylase, questionable and negligible activities for the decarboxylation of ornithine and arginine, and appreciable activity for ornithine aminotransferase. Uptake studies with radiolabeled putrescine, spermidine and spermine showed that these amines are rapidly taken up from the medium by an active uptake process. The uptake was temperature-sensitive and abolished at 0–4°C. The questionable presence of biosynthetic enzymes such as ornithine and arginine decarboxylase and, on the other hand, an effective uptake mechanism indicate that the parasite may depend on the host for its polyamine requirement, thereby indicating a possible target for chemotherapy.

Polyamine metabolism in Ancylostoma ceylanicum and Nippostrongylus brasiliensis.

Spermidine was detected as the major polyamine of Ancylostoma ceylanicum as well as Nippostrongylus brasiliensis. Spermine was present in lower amounts whereas the level of putrescine was even less. S-Adenosylmethionine decarboxylase, a rate-limiting enzyme in the biosynthetic pathway of polyamines, was demonstrated at low levels in both parasites. Decarboxylation of lysine and arginine was absent or negligible and that of ornithine questionable, as the enzyme activity was not inhibited by alpha-difluoromethylornithine while RMI 71,645, an irreversible inhibitor of ornithine aminotransferase, strongly inhibited the liberation of CO2 from ornithine. High activity of ornithine aminotransferase was observed in both the parasites and may interfere with the assay for ornithine decarboxylase. Adults of A. ceylanicum were found to rapidly take up spermidine and spermine from incubation medium while uptake of putrescine was very low. These results indicate that hookworms depend on uptake and interconversion rather than de novo synthesis for their polyamine requirement.

Hepatic microsomal cytochrome P450 system during experimental hookworm infection

Experimental infection of golden hamsters with the hookworm, Ancylostoma ceylanicum, caused a profound decline in the hepatic microsomal cytochrome P450 content. Concomitant decrease was also noticed in aminopyrine N-demethylase and benzo[a]pyrene hydroxylase activities. However, aniline hydroxylase activity was only marginally elevated during the infection. Microsomal markers, viz., cytochrome b5, NADH-cytochrome-c reductase, and glucose-6-phosphatase, were not significantly altered. Hepatic tissue exhibited an accumulation of lipids, especially phospholipids, triglycerides, and cholesterol, resulting in fatty necrosis around the central vein region. Isolated hepatic microsomes showed a decrease in phosphatidylcholine content. Impairment in hepatic mixed function oxidase (MFO) activities was further confirmed by prolongation in hexobarbital sleeping time and zoxazolamine-induced paralysis. The hepatic MFO system of A. ceylanicum-infected hamsters responded qualitatively and quantitatively in a manner similar to that of control hamsters, upon stimulation with selective chemical inducers like phenobarbitone and 3-methylcholanthrene. Kinetic and in vitro substrate binding studies revealed that for aminopyrine the substrate affinity and the maximum enzyme activity (Vmax) were decreased, while for aniline the binding affinity was decreased and the binding capacity was enhanced. Results indicate specific/selective impairment of the hepatic microsomal cytochrome P450 system during hookworm infection and may have many practical implications in toxicology and pharmacology.

Biochemical and histopathological alterations in golden hamster during infection with Ancylostoma ceylanicum

Ancylostoma ceylanicum infection in golden hamsters (Mesocricetus auratus) caused marked biochemical and histopathological derangements. Jejunum, the primary site of infection, showed pronounced alterations compared with liver. Though the biochemical composition of jejunum was not significantly altered, activities of a few lysosomal enzymes were enhanced during hookworm infection. Marked damage to mitochondrial and microsomal membranes was reflected in changes in the activities of the marker enzymes from jejunal tissue. Lipid content, especially phospholipids and neutral lipids of hepatic tissue, exhibited marked elevation. Levels of hexokinase, phosphofructokinase, and lactate dehydrogenase were enhanced in jejunal as well as hepatic tissues, indicating activation of the glycolytic machinery during hookworm infection. A decrease in the levels of mucosal disaccharidases indicated damage to intestinal brush border membranes. However, alkaline phosphatase activity was increased in intestinal mucosa during the infection. Light microscopic examination of jejunal tissue revealed peeling off of the upper epithelial layer, activation of the goblet cells, and thickening of muscularis mucosa. However, hepatic tissue did not show gross alterations, except for slight necrosis in the centrilobular region.

Characterization of lactate dehydrogenase of Plasmodium knowlesi

Some properties of purified lactate dehydrogenase, (EC. 1.1.1.27) from schizonts of Plasmodium knowlesi are described. The plasmodial enzyme migrated as single entity on polyacrylamide gel, and resembled rabbit muscle (M4) lactate dehydrogenase in its electrophoretic mobility. The P. knowlesi enzyme consisted of four identical subunits of 31 kDa. Purified lactate dehydrogenase was inhibited almost completely when incubated with 100 microM p-chloromercuribenzoate, Ag+ or Hg+ and such inhibition could be reversed by the addition of beta-mercaptoethanol or L-cysteine. Metal chelators did not show any remarkable effect. Oxalic acid is a competitive inhibitor of pyruvate reduction by this enzyme with apparent Ki of approximately equal to 0.41 mM.

Status of the microsomal mixed function oxidase system in the intestine, lungs and kidneys of hamsters during infection with Ancylostoma ceylanicum☆

Abstract Tekwani B. L., Tripathi L. M., Mukerjee S., Visen P. K. S., Katiyar J. C., Shukla O. P. and Ghatak S. 1988. Status of the microsomal mixed function oxidase system in the intestine, lungs and kidneys of hamsters during infection with Ancylostoma ceylanicum . International Journal for Parasitology 18 : 11–14. Infection of golden hamsters with Ancylostoma ceylanicum causes significant alterations in microsomal cytochrome P-450 and associated components in intestine, lungs and kidneys. In intestinal microsomes, cytochrome P-450, cytochrome b 5 , heme, cytochrome b 5 reductase, NADPH-cytochrome c reductase and benzo(a)pyrene hydroxylase decreased considerably. This generalized damage in jejunal microsomes appears to be due to marked pathological lesions in this tissue. Lung microsomes exhibited a decrease in cytochrome P-450, heme and benzo(a)pyrene hydroxylase, while cytochrome b 5 , and cytochrome b 5 reductase were not significantly affected. Renal microsomes showed elevation in cytochrome P-450 and benzo(a)pyrene hydroxylase, while other associated components remained unaffected.

Suppression of release of microfilariae in vitro from Setaria cervi (Nematoda: Filariodeae) by chlorpromazine.

Chlorpromazine (CPZ) caused suppression in release of microfilariae in vitro from Setaria cervi at 10(-5) M concentration. The inhibitory effect of CPZ was further stimulated by Ca2+ (2 mM), whereas addition of EGTA (2 mM) resulted in partial reversal. Addition of dopamine (1 mM) along with CPZ (10(-5) M) caused similar inhibition as was observed in the presence of CPZ (10(-5) M) alone. CPZ did not effect in vitro oxygen uptake by worm homogenates up to 0.5 mM concentration. The results indicate the Ca2+/calmodulin-dependent anthelmintic action of CPZ on S. cervi.