Objective: To explore whether the upregulation of inducible nitric oxide synthase (iNOS) is involved in lipopolysaccharide (LPS)-induced neurodegeneration.
Methods: 108 SD rats were randomly divided into 2 groups: experimental group and normal control group, and each group was sub-divided into 5 subgroups of 18 rats to undergo examination at different time points (6 h, 12 h, 1 d, 3 d, and 7 d). LPS was stereotaxically infused into the substantia nigra (SN) of left side of the experimental rats and PBS was used instead for the control rats. At different time points different numbers of rats from each subgroup were killed to take out the SN. Biochemical method was used to test the activity of NO and iNOS in 6 rats from each subgroup, iNOS mRNA expression was tested by RT-PCR in 3 rats from each subgroup, and iNOS protein expression was tested by Western blotting in 4 rats from each subgroup. Immunohistochemistry was used to detect the iNOS positive cells.
Results: iNOS positive cells were found since 6h after the intranigral injection of LPS, peaked 1d after, began to decrease 3d after, and basically disappeared 7d after; and were not found in the control group and the SN at the opposite side of the experimental rats. The percentage of iNOS-positive neurons 1d after the injection was 45.30 +/- 4.63, significantly higher than that of the control group (0.11 +/- 0.04, P < 0.01). RT-PCR and Western blotting showed that expression of iNOS mRNA and expression of iNOS protein at all time points were all higher than those of the normal controls and PBS controls (all P < 0.01). iNOS activity and NO amount in the LPS-injected SN began to increase 6 h after the injection, significantly higher then that of the control group (P < 0.05), peaked 1d after, (P < 0.01), began to decrease 3d after, and basically returned to normal level.
Conclusion: Up-regulation of iNOS may be one of the crucial mechanisms in LPS-induced degeneration of DA neurons.