Changes of mitochondrial ultrastructures and function in central nervous tissue of hens treated with tri-ortho-cresyl phosphate (TOCP).

Tri-ortho-cresyl phosphate (TOCP), an organophosphorus ester, is capable of producing organophosphorus ester-induced delayed neurotoxicity (OPIDN) in humans and sensitive animals. The mechanism of OPIDN has not been fully understood. The present study has been designed to evaluate the role of mitochondrial dysfunctions in the development of OPIDN. Adult hens were treated with 750 mg/kg·bw TOCP by gavage and control hens were given an equivalent volume of corn oil. On day 1, 5, 15, 21 post-dosing, respectively, hens were anesthetized by intraperitoneal injection of sodium pentobarbital and perfused with 4% paraformaldehyde. The cerebral cortex cinerea and the ventral horn of lumbar spinal cord were dissected for electron microscopy. Another batch of hens were randomly divided into three experimental groups and control group. Hens in experimental groups were, respectively, given 185, 375, 750 mg/kg·bw TOCP orally and control group received solvent. After 1, 5, 15, 21 days of administration, they were sacrificed and the cerebrum and spinal cord dissected for the determination of the mitochondrial permeability transition (MPT), membrane potential (Δψ(m)) and the activity of succinate dehydrogenase. Structural changes of mitochondria were observed in hens' nervous tissues, including vacuolation and fission, which increased with time post-dosing. MPT was increased in both the cerebrum and spinal cord, with the most noticeable increase in the spinal cord. Δψ(m) was decreased in both the cerebrum and spinal cord, although there was no significant difference in the three treated groups and control group. The activity of mitochondrial succinate dehydrogenase assayed by methyl thiazolyl tetrazolium (MTT) reduction also confirmed mitochondrial dysfunctions following development of OPIDN. The results suggested mitochondrial dysfunction might partly account for the development of OPIDN induced by TOCP.

[Effect of phenylmethylsulfonyl fluoride pretreated on neurofilament subunits in spinal cords of hens administrated with tri-o-cresyl phosphate].

OBJECTIVE: To investigate the dynamic changes of neurofilaments (NFs) proteins in spinal cords of hens with phenylmethylsulfonyl fluoride (PMSF) pretreatment for exploring the mechanism of tri-o-cresyl phosphate (TOCP)-induced delayed neuropathy (OPIDN). METHOD: Adult Roman hens were randomly divided into three groups, control, TOCP and PMSF + TOCP. Birds in PMSF + TOCP set were pretreated with PMSF, 24 hours later, hens in both TOCP group and PMSF + TOCP group were administrated with TOCP at a single dosage of 750 mg/kg. Then all animals were sacrificed on the corresponding time-points of 1, 5, 10, and 21 days respectively after dosing of 750 mg/kg TOCP. The spinal cords were dissected, homogenized, and centrifuged at 100,000 x g. The levels of high molecular neurofilament (NF-H), medium molecular neurofilament (NF-M) and low molecular neurofilament (NF-L) in both pellet and supernatant fractions of spinal cords were determined by SDS-PAGE and Western-blotting. RESULTS: The hens in TOCP group showed paralysis gait at the end of 21-day experimental period. The levels of NFs proteins in spinal cords changed obviously. Compared with control, the NFs in pellet showed a dramatic decrease on day 10 and then followed by a recovery. In the supernatant, the NFs proteins showed similar changes, which decreased significantly on day 10 and almost recovered control on day 21. Such as, NF-L, NF-M and NF-H decreased by 51%, 86% and 38% on day 10. The OPIDN signs were not observed in PMSF + TOCP group, and imbalances of NFs were obviously alleviated. Compared with control, only NF-M in pellet increased by 21% (P < 0.05) on day 21, others remained no changes; The levels of NF-H and NF-M in supernatant respectively increased by 19% and 35% on day 21, others were no significant statistical differences. CONCLUSION: TOCP may induce imbalance of NFs levels in progress of OPIDN, and PMSF pretreatment may protect animals from OPIDN by reducing above changes, which may explain that TOCP-induced imbalance of NFs may be connected with the occurrence and development of OPIDN.

[Effects of tacrolimus (FK506) on heat-shock proteins 70, Bcl-2 and Bax expression in nervous tissue of acrylamide-induced rat].

OBJECTIVE: To investigate the effects of tacrolimus (FK506) on behavioral function and heat-shock proteins (HSP70) expression in nervous tissues of acrylamide (ACR)-induced rats. METHODS: Totally 40 health Wistar rats were randomly divided into control group, model group, low and high doses of FK506 groups. All four groups were treated five times per week for four weeks. Gait score was measured every week. And rats were sacrificed on day 28, the cerebrum, spinal cord and sciatic nerve were dissected, and homogenized in ice bath, then the levels of HSP70 and Bcl-2, Bax were analyzed by western bloting. RESULTS: Compared with the ACR model group, the gait score in low and high doses of FK506 groups decreased by 30.1% and 47.7% respectively in the 4th week. In the cerebrum and sciatic nerve pellet, the level of HSP70 in the FK506 groups increased by 11.6%, 33.3% and 56.3%, 58.5% (P < 0.01), but no significant changes existed in spinal cord. The level of Bcl-2 in the sciatic nerve pellet increased by 39.1% (P < 0.01) but no significant changes existed in the cerebrum and spinal cord from low dose of FK506 group. And the level of Bax in the spinal cord pellet markedly increased by 46.8% but not in cerebrum and sciatic nerve pellet; Whereas in the tissues mentioned above, the levels of Bcl-2 were enhanced remarkably by 16.3%, 14.8% and 56.0% (P < 0.01) in the high dose of FK506 group. And the level of Bax in the cerebrum and spinal cord pellet markedly increased by 16.4% and 40.2% but not in sciatic nerve. The values of Bcl-2/Bax in low and high doses of FK506 groups clearly increased by 15.9%, 33.3%, 36.9% and 30.1%, 49.1%, 60.1% (P < 0.01). CONCLUSION: The administration of FK506 has dramatically neuroprotective effects against the development of ACR neuropathy, which may be related to up-regulating the expression of HSP70 and Bcl-2 with down-regulating the expression of Bax.

The anti-fatty liver effects of garlic oil on acute ethanol-exposed mice.

The protective effects of single dose of garlic oil (GO) on acute ethanol-induced fatty liver were investigated. Mice were treated with ethanol (4.8 g/kg bw) to induce acute fatty liver. The liver index, the serum and hepatic triglyceride (TG) levels and the histological changes were examined to evaluate the protective effects. Hepatic malondialdehyde (MDA), glutathione (GSH) levels and superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GSH-Px), glutathione-S-transferase (GST) activities were determined for the antioxidant capacity assay. Acute ethanol exposure resulted in the enlargement of the liver index and the increase of the serum and hepatic TG levels (P<0.01), which were dramatically attenuated by GO pretreatment in a dose-dependent manner (P<0.01). GO treatment (simultaneously with ethanol exposure) exhibited similar effects to those of pretreatment, while no obviously protective effects were displayed when it was used at 2h after ethanol intake. Histological changes were paralleled to these indices. Beside this, GO dramatically prolonged the drunken time and shortened the waking time, and these effects were superior to those of silymarin and tea polyphenol. In addition, GO dose-dependently suppressed the elevation of MDA levels, restored the GSH levels and enhanced the SOD, GR and GST activities. Compared with the ethanol group, the MDA levels decreased by 14.2% (P<0.05), 29.9% and 32.8% (P<0.01) in GO groups 50, 100 and 200 mg/kg, respectively. The GST activity increased by 9.97%, 19.94% (P<0.05) and 42.12% (P<0.01) of the ethanol group in GO groups 50, 100 and 200 mg/kg, respectively, while the GR activity increased by 28.57% (P<0.05), 37.97% (P<0.01), 50.45% (P<0.01) of the ethanol group in GO groups 50, 100 and 200 mg/kg, respectively. These data indicated that single dose of GO possessed ability to prevent acute ethanol-induced fatty liver, but may lose its capacity when used after ethanol exposure. The protective effects should be associated with its antioxidative activities.