Effects of duration of exposure, co-supplementation with iron and Nippostrongylus brasiliensis infection on accumulation of trichloroacetic acid-insoluble copper in rats given molybdate in drinking water.

Thiolation can convert molybdate (MoO4) into a series of thiomolybdates (MoSxO4-x) in the rumen, terminating in tetrathiomolybdate (MoS4), a potent antagonist of copper absorption and, if absorbed, donor of reactive sulphide in tissues. Systemic exposure to MoS4 increases trichloroacetic acid-insoluble copper (TCAI Cu) concentrations in the plasma of ruminants and induction of TCAI Cu in rats given MoO4 in drinking water would support the hypothesis that rats, like ruminants, can thiolate MoO4. Data on TCAI Cu are presented from two experiments involving MoO4 supplementation that had broader objectives. In experiment 1, plasma Cu concentrations (P Cu) tripled in female rats infected with Nippostrongylus brasiliensis after only 5 days exposure to drinking water containing 70 mg Mo L-1, due largely to an increase in TCAI Cu; activities of erythrocyte superoxide dismutase and plasma caeruloplasmin oxidase (CpOA) were unaffected. Exposure for 45-51 days did not raise P Cu further but TCA-soluble (TCAS) Cu concentrations increased temporarily 5 days post infection (dpi) and weakened the linear relationship between CpOA and TCAS Cu. In experiment 2, infected rats were given less MoO4 (10 mg Mo L-1), with or without iron (Fe, 300 mg L-1), for 67 days and killed 7 or 9 dpi. P Cu was again tripled by MoO4 but co-supplementation with Fe reduced TCAI Cu from 65 ± 8.9 to 36 ± 3.8 µmol L-l. Alone, Fe and MoO4 each reduced TCAS Cu in females and males when values were higher (7 and 9 dpi, respectively). Thiolation probably occurred in the large intestine but was inhibited by precipitation of sulphide as ferrous sulphide. Fe alone may have inhibited caeruloplasmin synthesis during the acute phase response to infection, which impacts thiomolybdate metabolism.

Effects of Molybdate and Tetrathiomolybdate Supplementation of Drinking Water on Immature Rats Infected with Nippostrongylus brasiliensis. 2. Copper Status and Tissue Molybdenum Accretion.

Molybdate (MoO4) and tetrathiomolybdate (MoS4) supplementation of rats via drinking water had opposite effects on the establishment of Nippostrongylus brasiliensis larvae but both induced hypercupraemia, temporarily inhibited activities of superoxide dismutase in liver and duodenum after infection and enlarged the femoral head. Effects of MoO4 and MoS4 on activities of caeruloplasmin oxidase (CpO) in plasma, erythrocyte superoxide dismutase (ESOD) and tissue copper (Cu) and molybdenum (Mo) were compared to test the hypothesis that species lacking a rumen can thiolate MoO4. Three groups of 18 immature Wistar rats were given Mo (70 mg/L as MoO4) or MoS4 (5 mg/L) via drinking water or remained untreated; all received a commercial, cubed diet and 12 from each group were infected with larvae of N. brasiliensis. Rats were killed 7-9 days later and liver, kidney, spleen, heart, muscle (quadriceps), brain and bone (femur) removed for Cu and Mo analysis. Plasma Cu was greatly increased by MoO4 and MoS4, without changing CpO activity, but the effect was more variable with MoO4 and accompanied by a smaller decrease in ESOD. Tissue Cu and Mo were increased by MoS4 in all tissues examined except brain and bone, correlating with plasma Cu and with each other; relationships were strongest in spleen, followed by kidney. MoO4 also increased soft tissue Cu and Mo but increases were generally smaller than those induced by MoS4 and correlations between the two elements and with plasma Cu generally weaker. Since hypercupraemia and correlated increases in liver and kidney Cu and Mo are characteristic of systemic thiomolybdate (TM) exposure, we conclude that MoO4 was partially thiolated to give a different TM profile from that produced by MoS4. The pathophysiological significance of systemic exposure to di- and tri-TM merits investigation in non-ruminants as agents of chelation therapy and in ruminants as agents of short-lived TM toxicity on Mo-rich pastures.

Effects of Chronic Exposure to Molybdate and Tetrathiomolybdate Supplementation of Water on Immature Rats Acutely Infected with Nippostrongylus brasiliensis. 1. Effects on Outcome of Infection and Host Health.

Low molybdate (MoO4) exposure via drinking water in mature rats infected with Nippostrongylus brasiliensis raised liver and plasma copper (Cu) concentrations. The possibility that anthelmintic effects were attributable to conversion of MoO4 to tetrathiomolybdate (MoS4) in a non-ruminant species was investigated by giving three groups of 18 immature rats drinking water containing 70 mg Mo l-1 as MoO4 (group A), 5 mg Mo l-1 as MoS4 (group B) or no supplement (group C), while receiving a commercial cubed diet. After 41 days, 12 rats from each group were inoculated subcutaneously with 2,000 L3-stage N. brasiliensis larvae. Subgroups were killed 7, 8 or 9 days post infection (dpi), when adult worms are normally expelled, and enzyme markers for the inflammatory response to infection were measured in plasma or liver. Male rats given MoS4 prior to infection grew more slowly than those given MoO4. Eight dpi, females given MoS4 had lost more bodyweight than those in group C, while those given MoO4 had gained weight. Mean worm counts at 7 dpi were 160, 65 and 250 ± 30.6 (SE), respectively, in groups C, A and B, and differed significantly from each other (P <0.05) but only rats given MoO4 remained infected 9 dpi (mean worm count 52 ± 16.4): Faecal egg counts followed a broadly similar pattern. Both Mo sources pre-empted increases in liver and duodenal superoxide dismutase activity, induced by infection 7 and 9 dpi, respectively, in group C and enlarged the femur: neither source prevented hypertrophy of the small intestine and a rise in serum mast cell protease concentration caused by infection. Since data for plasma Cu concentration and caeruloplasmin oxidase activity, reported separately, indicated MoO4 was thiolated in vivo, differences between Mo sources may be attributable to differences in the degree of thiolation, extent of thiomolybdate exposure and rates of thiomolybdate degradation at critical times in host or parasite development.

Effects of Raising or Lowering Molybdenum Status on Outcome of Acute Infection with Nippostrongylus brasiliensis in Mature Rats.

Molybdate (Mo+) supplements can suppress or enhance nematode infections in ruminants, depending on exposure level, but there have been no investigations in non-ruminants. Three groups of 16 mature rats were each fed a commercial diet and given Mo+ (10 mg Mo/l), tungstate (a molybdenum [Mo] antagonist) (MoO4, 350 mg W/l) or no supplement (C) via drinking water for 40 days before acute infection with 3,600 Nippostrongylus brasiliensis larvae. Group Mo- also received allopurinol (1 g/l), a molybdenoenzyme inhibitor, from 4 days post infection (dpi). Subgroups of four rats from each group were killed at 7-14 dpi. A group of six rats was left untreated and uninfected and subgroups killed 10 or 12 dpi. Infection reduced intakes of food and water but impacts were greatest in group Mo-. Median worm counts in groups C, Mo- and Mo+ were 900, 941 and 510, respectively, at 7 dpi and 9, 40 and 0 (P = 0.05) at 10 dpi. Median faecal egg counts were consistently lowest in group Mo+. Worm weight was reduced (P <0.05), worm tissue protease increased and superoxide dismutase activities increased in worm (P < 0.01) and host duodenal homogenates (P < 0.01) from group Mo+. In group Mo-, liver Mo concentration decreased, duodenal xanthine oxidoreductase activity (DXOR) became totally inhibited and plasma uric acid was barely detectable at 10 dpi. Plasma mast cell protease activity and duodenal malonyldialdehyde concentrations, markers of inflammation, were increased by nematode infection (P <0.001) but unaffected by water treatments. Liver Mo, liver copper (Cu) and plasma Cu concentrations were increased in group Mo+ and plasma Cu concentration was increased in group Mo- suggesting systemic exposure to partially thiolated MoO4 and WO4. Supplementary MoO4 impaired larval establishment and changed parasite biochemistry without affecting the inflammatory response to infection but may have required partial thiolation to do so. Rats did not rely on DXOR activity to expel N. brasiliensis.