해조류에서 유래한 HS-1580의 항산화작용을 생체에서 확인하기 위하여 파킨슨병 모델동물을 대상으로 실험을 실시하였다. 10주령의 수컷 C57/BL6 생쥐의 복강에 43.6% dimethyl sulfoxide (DMSO), HS-1580 혹은 L-NAME를 주입한 삼투압펌프를 삽입하고, 2일 후에 복강에 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) (20 mg/kg)를 주사하여 파킨슨병 모델동물을 제작하였다. MPTP를 주사하고 2일과 7일 후에 동물을 관류고정하고 뇌를 적출하여 면역조직화학법을 실시하였다. 일차항체로는 rabbit anti-tyrosine hydroxylase와 rat anti-mouse MAC-1를 사용하였다. MPTP에 의한 흑색질의 TH면역반응 양성 세포의 수적인 감소와 줄무늬체의 TH-면역반응 양성 축삭종말의 상대적인 밀도의 감소가 HS-1580 혹은 L-NAME 전처리로 유의하게 억제되었다. 또한 MPTP에 의하여 흑색질에서 MAC-1 면역반응 양성의 미세아교세포의 활성화가 HS-1580 혹은 L-NAME 전처리에 의하여 억제되었다. 이상의 결과는 MPTP에 의해 유도된 도파민성신경세포의 형질발현 양상의 감소가 해양성 신물질인 HS-1580 혹은 NOS 억제제인 L-NAME를 전처리함으로써 억제되는 것을 형태학적으로 확인하였다.
The antioxidant effects of HS-1580 derived from Brown seaweeds in comparison with NOS inhibitor, L-NAME, were tested in the Parkinson’s disease animal model. C57BL/6 mice were implanted with osmotic pump containing vehicle, HS-1580 or L-NAME intraperitoneally 2 days before MPTP injection. The Parkinson’s animal model were prepared by intraperitonal injection of MPTP (20 mg/kg, 4 times with 2 hours intervals in a day). The mice were perfused with Zamboni fixative 2 days or 7days after MPTP injection. The 30 μm cryostat section were immunostained with free-floating method. Rabbit anti-tyrosine hydroxylase (TH) and rat anti-mouse MAC-1 were used as the primary antibodies. The following results were obtained. The number of TH-immunoreactive (ir) neurons in substantia nigra (SN) and the relative density of TH-ir axon terminals in striatum were decreased by MPTP injection. But the decrease was significantly attenuated with 10% HS-1580 or L-NAME (50 mg/kg) pretreatment before MPTP injection. MAC-1-ir activated microglia were observed in substantia nigra 2 days after MPTP injection. Activated microglia were showed as thickened processes with round cell bodies. The morphological changes of MAC-1-ir activated microglia were inhibited by HS-15980 or L-NAME pretreatment before MPTP injction. Above results mean that the damage of nigrostriatal dopaminergic system was rescued by pretreatment of HS-1580 or L-NAME in MPTP-induced Parkinson’s disease animal model.
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