Reconsider Alzheimer's disease by the 'calpain–cathepsin hypothesis'—A perspective review

Alzheimer's disease (AD) is characterized by slowly progressive neuronal death, but its molecular cascade remains elusive for over 100 years. Since accumulation of autophagic vacuoles (also called granulo-vacuolar degenerations) represents one of the pathologic hallmarks of degenerating neurons in AD, a causative connection between autophagy failure and neuronal death should be present. The aim of this perspective review is at considering such underlying mechanism of AD that age-dependent oxidative stresses may affect the autophagic-lysosomal system via carbonylation and cleavage of heat- shock protein 70.1 (Hsp70.1). AD brains exhibit gradual but continual ischemic insults that cause perturbed Ca2+ homeostasis, calpain activation, amyloid b deposition, and oxidative stresses. Membrane lipids such as linoleic and arachidonic acids are vulnerable to the cumulative oxidative stresses, generating a toxic peroxidation product 'hydroxynonenal' that can carbonylate Hsp70.1. Recent data advocate for dual roles of Hsp70.1 as a molecular chaperone for damaged proteins and a guardian of lysosomal integrity. Accordingly, impairments of lysosomal autophagy and stabilization may be driven by the calpain-mediated cleavage of carbonylated Hsp70.1, and this causes lysosomal permeabilization and/or rupture with the resultant release of the cell degradation enzyme, cathepsins (calpain–cathepsin hypothesis). Here, the author discusses three topics; (1) how age-related decrease in lysosomal and autophagic activities has a causal connection to programmed neuronal necrosis in sporadic AD, (2) how genetic factors such as apolipoprotein E and presenilin 1 can facilitate lysosomal destabilization in the sequential molecular events, and (3) whether a single cascade can simultaneously account for implications of all players previously reported. In conclusion, Alzheimer neuronal death conceivably occurs by the similar 'calpain-hydroxynonenal-Hsp70.1-cathepsin cascade' with ischemic neuronal death. Blockade of calpain and/or extra-lysosomal cathepsins as well as scavenging of hydroxynonenal would become effective AD therapeutic approaches

A putative link of PUFA, GPR40 and adult-born hippocampal neurons for memory

金沢大学医薬保健研究域　医学系Long chain polyunsaturated fatty acids (PUFA) such as docosahexaenoic and arachidonic acids, which are enriched in the brain, are important for multiple aspects of neuronal development and function including neurite outgrowth, signal transduction and membrane fluidity. Recent studies show that PUFA are capable of improving hippocampal long-term potentiation, learning ability of aged rats, and cognitive function of humans with memory deficits, although the underlying mechanisms are unknown. There have been several reports studying physiological roles of G-protein coupled receptor 40 (GPR40) in the pancreas, but no studies have focused on the function of GPR40 in the brain. As GPR40 was recently identified in neurons throughout the brain, it is probable that certain PUFA may act, as endogenous ligands, on GPR40 at their cell surface. However, the effects of PUFA upon neuronal functions are still not clearly understood. Here, although circumferential, a combination of in vitro and in vivo data is introduced to consider the effects of docosahexaenoic and arachidonic acids on brain functions. GPR40 was found in the newborn neurons of the normal and postischemic hippocampi of adult macaque monkeys, while the positive effects of PUFA upon Ca2+ mobilization and cognitive functions were demonstrated in both GPR40 gene-transfected PC12 cells and human subjects with memory deficits. The purpose of this review is to propose a putative link among PUFA, GPR40, and hippocampal newborn neurons by discussing whether PUFA can improve memory functions through GPR40 activation of adult-born neurons. At present, little is known about PUFA requirements that make possible neurogenesis in the adult hippocampus. However, the idea that \u27PUFA-GPR40 interaction might be crucial for adult neurogenesis and/or memory\u27 should be examined in detail using various experimental paradigms. © 2007 Elsevier Ltd. All rights reserved

Еxpression of Transcriptional Factor SOX2 in Populations of Cortical Progenitors in Human Fetal Telencephalon

Cerebral cortex оf mammals is mainly generated during the embryonic period by stem cells and their derivative progenitors in the palium of the developing telencephalon. Various genes in complex interactions are involved in the processes of differentiation of the cerebral neurons. Transcriotional factor Sox2 plays a key role for self renewing and sustaining multipotency of embryonic neural stem/progenitor cells. Data about the expression and function of Sox2 in human fetal brain are insufficient and controversial.In the present sudy tissue samples of spontaneously aborted human fetuses aged between 12th to 28th gestational weeks (g. w.) were examined by a standard histological and immunohistochemical technique for paraffin sections. Sox2 expression was followed in the zones of cellular proliferation and migration in the occipital lobe of human fetal telencephalon mainly during 17th g. w. Within ventricular and outer subventricular zones we detected similar amount approximately 45% Sox2+ cells, whereas in the intermediate zone, cortical plate and marginal zone expression of Sox2 was not found.The data obtained on the location and expression dynamics of Sox2 contribute to a more complete understanding of neural stem/progenitor cell biology during embryonic neurogenesis in the human cerebral cortex

Quantity and Distribution of Proliferating Cells in the Juvenile and Adult Primate Spinal Cord

ÃŒultipotent progenitors exist in the adult mam- malian central nervous system, capable of producing both neurons and glia. Their proliferation in the spi- nal cord is limited. Generation of putative stem/pro- genitor cells has been reported in intact and injured spinal cord of rodents and in a limited number in monkeys with a spinal injury, but not in intact spi- nal cord in vivo. We recently reported de novo gener- ated cells in the intact spinal cord of macaque mon- keys. Here we extend these findings by showing data of intact juvenile and neonatal monkey spinal cord. We used bromodeoxyuridine (BrdU) to label the de novo generated cells in the experimental animals and stud- ied their quantity and distribution at different time- points after the BrdU infusion. As expected, we found a significant elevation of the BrdU-labeled cells at neo- natal stage. However, there was no difference between juvenile and adult stages. These results suggest that the survival of newly born cells in the intact primate spinal cord does not change after juvenile stage and this could be used to further study repair mechanisms in adult primate spinal cord

Phenotype of De Novo Generated Cells in the Spinal Cord of Adult Macaque Monkeys

Neuronal stem and progenitor cells exist in the spinal cord of sexually mature mammals. Òhåy play an important role during repaining processes after in- jury, but their proliferation and differentiation are limited. In the present study we used the proliferative marker bromodeoxyuridine (BrdU) for a short (2 h) and three longer survival periods (2, 5 and 10 weeks) to investigate the quantity, topography and fate of de novo generated cells in intact spinal cord of adult pri- mates (macaque monkeys). We applied as well single or in combinations markers for mesenchymal cells or/ and neuronal stem/progenitor cells to demonstrate the phenotype of the proliferating cells. We found that af- ter the short period of BrdU application (2 h) the num- ber of BrdU+ cells is significantly elevated only in the cervical segments. Most of the cells in the ependymal layer are immunopositive for Vimentin or/and Nestin. This is an indice for their cellular belonging. A con- siderable number of Vimentin+ cells of the ependy- mal layer form long characteristic processes directed to underlying blood capillaries. Òhis indicates their participation as a component of the ependymal cellu- lar niche. The presence of BrdU+/Nestin+ cells in the central canal surrounding zone confirms the existence of dividing neuronal stem/progenitor cells cells in the spinal cord of adult primates

脳腫瘍における細胞間接着因子(カドヘリン)の研究

金沢大学医学部カドヘリンはカルシウム依存性の細胞接着分子で、組織の形態形成や腫瘍形成において重要な役割を果たすとされている。本研究においては、髄膜腫の発生に際しカドヘリンがどのような役割を果たしているかを明らかにするために、髄膜腫とその発生母地であるクモ膜絨毛について、生化学的ならびに免疫組織化学的な研究を行った。対象は3例のクモ膜絨毛と31例の髄膜腫であり、後者の内訳は合胞細胞型(syncytial type)が11例、移行型(transitional type)が12例および線維芽細胞型(fibroblastic type)が8例である。モノクロ-ナル抗体はヒト上皮型カドヘリン(E型カドヘリン)に特異的に反応するHECDー1を用いた。ウエスタンブロッティングでは、クモ膜絨毛および合胞細胞型と移行型の髄膜腫において全例に分子量約124KDのE型カドヘリンが検出されたのに対し、線維芽細胞型では1例も検出されなかった。免疫組織化学的には、クモ膜絨毛ではクモ膜細胞層(arachnoid cell layer)、細胞集簇部(cap cell cluster)および中芯部(central core)のクモ膜細胞にE型カドヘリンの発現がみられたが、線維性被膜(fibrous capsule)ではみられなかった。一方、髄膜腫においては、合胞細胞型と移行型の合胞体を形成する部分にE型カドヘリンの発現が強くみられたが、移行型の束状配列(stream)を形成する部分では弱く、線維芽細胞型ではみられなかった。免疫電顕による検索においては、クモ膜絨毛ではE型カドヘリンは、隣接するクモ膜細胞間に斑点状に凝集して存在していたのに対し、髄膜腫では接着装置を含め細胞間のほぼ全域に分布し、しかもその発現量は増加していた。以上の結果より、カドヘリンの発現様式の差異が髄膜腫の組織学的多様性と密接に関連していること、および腫瘍化に伴いカドヘリンの分布や発現量に変化が生ずることが示唆された。研究課題/領域番号:02670624, 研究期間(年度):1990出典：研究課題「脳腫瘍における細胞間接着因子(カドヘリン)の研究」課題番号02670624（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/ja/grant/KAKENHI-PROJECT-02670624/）を加工して作

カルパインとカテプシンの動態より見た霊長類の虚血性神経細胞死の病態と治療

金沢大学医学部海馬のCA-1ニューロンは近時記憶を担っているが、一過性の脳虚血に脆弱で短時間血流が途絶えただけでも、その数日後には壊死に陥り個体は記憶障害をきたす。この遅発性神経細胞死に関しては現在なおその病態はベールにおおわれており有効な治療法がない。我々はサルの脳虚血実験で活性型muカルバインの基質はリソソーム膜に存在すること、及びmuカルパインの活性化が加水分解酵素であるカテプシンBのリソソーム外への放出を惹起することを示唆する知見を得た。即ち、活性型muカルパインのみを認識する抗体を用いた免疫染色を行うと、虚血をかけていない海馬のCA-1ニューロンは全く染色されないのに対し、20分間の脳虚血を負荷後のCA-1ニューロンは、核周囲の胞体が陽性所見を呈した。しかも、免疫電顕により検索すると、空胞化したリソソーム膜に活性型カルパイン局在がみられた。従って、虚血後のCA-1ニューロンに特異的にmuカルパインの活性化が生ずること、及びin vivoでの活性型muカルパインの基質はリソソーム膜の膜蛋白かその関連蛋白であることが示唆された。さらに、抗カテプシンB抗体を用いた免疫染色を行うと、一過性の虚血後にはCA-1以外の海馬ニューロンはリソソーム内での染色性の増加を示すものであるのに対し、CA-1ニューロンは特異的にリソソーム外へのカテプシンの放出を示した。以上より、CA-1ニューロンにおいて特異的に活性化されるmuカルパインは、リソソーム膜蛋白を限定分解する結果膜の断裂をきたし、カテプシンBがリソソーム外に漏出するものと推定された。そこで、カテプシンBの特異的拮抗薬であるCA-074を虚血負荷後に投与すると、CA-1領域の約2/3のニューロンは軽度の虚血性変化を示すものの遅発生神経細胞死は免れていた。研究課題/領域番号:09267215, 研究期間(年度):1997出典：研究課題「カルパインとカテプシンの動態より見た霊長類の虚血性神経細胞死の病態と治療 」課題番号09267215（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09267215/）を加工して作

Proliferating cells in the adult primate cerebellar cortex after ischemia

INTRODUCTION: Brain ischemia is a devastating neurological condition with significant medical and social impact. Here we investigated the effect of experimental ischemia of different duration on the ability of the adult macaque cerebellum to produce new cells of specific phenotypes.MATERIAL AND METHODS: We used a well-established model of global brain ischemia in young adult Japanese monkeys applying bromodeoxyuridine (BrdU) for 5 days х 100 mg/kg daily. Animals were distributed into different experimental and control groups depending on postischemic survival periods: 4, 9, 15, 23, 44 days (D) and BrdU starting day. Immunohistochemical detection of BrdU+ cells, Iba1+ microglia and GFAP+ astroglia was performed on cryosections. Statistical evaluation of newly generated cells with phenotyping for microglia and astrocytes in various cortical layers of the cerebellum were done.RESULTS and CONCLUSIONS: The numbers of BrdU+ cells in some ischemic groups were significantly higher compared to control animals. By cerebrocerebellum, there was an increasing value by D4 group compared to the control, then slightly reducing in D9 and D15 groups and increasing again by D23 and D44 groups. In the spinocerebellum, an increase was detected only in D44 group. The newly generated cells were dispersed in all cerebellar cortical layers with highest concentration in Purkinje cell layer. Our data show that ischemia stimulates cellular proliferation in the cerebellum but this effect declines with time after ischemia. We found evidence for generation of new microglia but not for astroglia. Our data may contribute to a better understanding of regeneration in the cerebellum after brain ischemia

実験的脳血管攣縮が惹起する中膜筋細胞壊死の病態と治療

金沢大学医学部付属病院・脳神経外科脳血管攣縮は刺激物質による血管の機能的収縮(第1相), 中膜筋細胞壊死による血管の麻痺性拡張(第2相)および内膜肥厚による血管内腔の形態学的狭容(第3相)の3相性変化を示した. 即ち, 致死的な脳血管攣縮を合併したクモ膜下出血, 髄膜炎および被殻出血手術の計10症例より採取された攣縮血管を光顕的, 電顕的に検索した結果.(1)視床下部ことにlaterl hypo-thalamicに虚血性変化がみられた. また, くも膜下腔の小血管より多数の炎症細胞や血液物質が漏出していた.(2)脳血管攣縮は原因の如何を問わず同様な血管病変を惹起した.(3)内膜は光顕的には内皮下層の水腫と結合組織の増殖を示し, 血管内腔を形態学的に狭容していた. 電顕的には内皮細胞間にopen juncetonがみられ, 内皮下層ではmyofibroblastが増殖し多量の結合組織や基底膜様物質を産生していた.(4)中膜は光顕的に筋細胞の好酸性の凝固壊死により絶対数が減少し, 血管壁の菲薄化と血管内腔の拡張がみられた.(5)電顕的には中膜節細胞壊死の基本的特徴は筋線稚の融解であり, これらは微細顆粒状物質に置換していた.(6)細胞内器室は腫大した粗面小胞体と糸粒体のみが残存し, 胞体内には多数のライソゾーンがみられた.(7)細胞間隔では基底膜様物質の増加と多量の細胞残〓がみられた. さらに, 視交又槽に血液やエピネフリンを注入して製作した実験的脳血管攣縮モデル犬30頭より採取された攣縮血管を組織学的に検索した結果, (1)犬の視交又槽に1CC/kgの血液を注入しても臨床剖検例に似た中膜筋細胞壊死を作製し得なかった.(2)しかし, 0.2mg/kgのエピネフリンを注入すると, 実験2日目より90日目にかけて中膜細胞壊死が観察された.(3)中膜細胞壊死は視床下部の軟化巣とクモ膜下腔の小血管の達過性亢進を合併した犬で顕著であった.(4)エピネフリンの視交又槽内注入により作製された実験的脳血管攣縮は臨床剖検例によく似た中膜筋細胞壊死を〓した.Myonecrosis following cerebral vasospasm associated with subarachnoid hemorrhage, meningitsis and trans-sylvian surgery was ultrastructurally studied. The basic feature of myonecrosis was dissolution of myofilaments with resultant fine granular or filamentous material. The disintegrating cytoplasm often contained numerous glycogen granules, dense bodies, autophagic vacuoles and myelin-like membranous bodies. A well-developed sarcoplasmic reticulum was preserved despite myofilament dissolution, while mitochondria showed marked swelling. The nuclei showed either dilution of chromatin or pyknotic change. The basal lamina was remarkably thickened and maintained an irregular outline of the necrotic smooth muscle cells. Enlarged intercellular space contained abundant cellular debris, vesicular structures and connective tissue fibers. Furthermore, myonecrosis following the injection of epinephrine into the canine chiasmatic cistern was studied. Microscopically, the circle of Willis showed coagulation necrosis and fibrosis of the media. The fine structure of myonecrosis was characterized by six dynamic chenges of vacuolation, dissolution of myofilaments, focal cytoplasmic necrosis, fragmentation, coagulation necrosis and intercellular fibrosis. Despite a simple experimental procedure, the present models disclosed myonecrosis with a marked similarity to humans and contained all of the previously reported ultrstructural features of experimental myonecrosis.研究課題/領域番号:60570665, 研究期間(年度):1985 – 1987出典：研究課題「実験的脳血管攣縮が惹起する中膜筋細胞壊死の病態と治療」課題番号60570665（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/ja/report/KAKENHI-PROJECT-60570665/605706651987kenkyu_seika_hokoku_gaiyo/）を加工して作