ヒト剖検脳の，特に黒質におけるチロシン水酸化酵素の免疫組織学的検討

IMMUNOHISTOCHEMISTRY ON TYROSINE HYDROXYLASE IN THE SUBSTANTIA NIGRA OF HUMAN AUTOPSIED CASES

　抄録　モノアミンの螢光組織化学をヒト剖検脳に応用することは困難であるが，チロシン水酸化酵素（TH）を指標としたカテコールアミン（CA）ニューロンの免疫組織化学は剖検脳においても可能と思われた。本研究ではOkaらの方法にほぼ従って牛副腎髄質からTHを精製し，ウサギに免疫して抗TH抗体を作成し，種々の死因を有する40剖検脳で，さまざまな固定条件下にある黒質を中心に，パラフィン切片上での酵素抗体間接法を行った。黒質におけるTHの陽性染色は脳死例を除くほぼ全例で，神経細胞の胞体，樹状突起，軸索に限局して認められ，これらTH陽性細胞はメラニン含有神経細胞によく一致していた。一方，網状層のメラニン非含有神経綱胞は　TH陰性で，また，緻密層にも散在する非含有神経細胞も陰性であり，黒質緻密層にはCAニューロン以外の他の神経細胞も混在していた。さらに興味ある点はTH陰性のメラニン含有神経細胞が20歳代に出現し始め加齢とともにその数が増加する傾向がみられたことで，これは加齢に伴うCAの変動を考える上で重要な所見と思われた。以上より，ヒト剖検脳におけるTHの免疫組織化学は各種変性性神経疾患のCAの病態の解折に有用と思われた。

Despite the numerous data on the distribution of catecholamine (CA) neurons in a wide variety of species, data on that of human are rare because of technical difficulty in applying fluorescent his-tochemical methods. On the other hand, immuno-histochemistry on tyrosine hydroxylase (TH) which was essential for synthesis of CA, was assumed to be applicable to human autopsied brain.

We performed indirect immunoperoxidase study on TH using 10% formalin or buffered formalin fixed, paraffin embedded sections of the substantia nigra from 40 autopsied cases, ranging in age 7 to 70 years. TH was purified from the bovine adrenal medulla using the procedure described by Oka et al. with some modification. Purified TH was injected with Freund's complete adjuvant into rabbits. The specificity of the antiserum was de-monstrated by the inhibition study of CA relating enzymes, double immunodiffusion and immunoelec-trophoresis against tissue homogenate of bovine adrenal medulla.

TH immunoreactive (TH-IR) products were res-tricted to the neuronal soma, axons and dendrities in almost all of the case except for the "dead brain" cases. TH-IR neurons were of large poly-gonal or multipolar cells, and closely correspond to melanin-pigmented neurons, located in the com-pact zone and scattered in the reticular zone in the substantia nigra. On the other hand, mela-nin free neurons in the reticular and compact zone, were of medium or small bipolar cells and were not stained. It was of interest that the non TH-IR melanin-pigmented neurons firstly appear-ed in the substantia nigra at third decade and increased their number with age.

This finding might be related to aging of the melanin-pigmented neuron of the substantia nigra. These result suggest that TH immunohistochemis-try is available for not only identification of the distribution and cytoarchitecture of human CA neuron system, but detection of pathogenesis of CA alteration in the senile dementia and degene-rative disease such as idiopathic parkinsonism.