酒井 一由

光学顕微鏡レベルで鍍銀陽性小体(核小体様封入体)を観察するためにグルタールアルデヒド含有固定が必須であるかどうか検討した。正常ddY雄マウスを使用し、10%ホルマリン灌流固定群、2.5%グルタールアルデヒド+5%ホルマリン混合液固定群、グルタールアルデヒド0.1%、0.5%、1.0%、2.5%灌流固定群を設定した。2.5%グルタールアルデヒドと5%ホルマリン混合液固定液で1日固定した標本では鍍銀陽性小体の存在が確認できた。最初に10%ホルマリンで固定された脳組織でも、その後にグルタールアルデヒド含有固定液で再固定すれば、鍍銀陽性小体の検索が可能であることが確認された。パラフィン切片での鍍銀陽性小体である核小体様封入体の染色にはグルタール含有固定が必須であることが証明され、最適な固定液は2.5%グルタールアルデヒドと5%ホルマリンの混合液であった。

<b><i>Introduction:</i></b> Patients with dementia show reduced adaptive, behavioral, and physiological responses to environmental threats. Physical exercise is expected to delay brain aging, maintain cognitive function and, consequently, help dementia patients face threats and protect themselves skillfully. <b><i>Methods:</i></b> To confirm this, we aimed to investigate the effects of the shaking exercise on the avoidance function in the senescence-accelerated mouse-prone strain-10 (SAMP-10) model at the behavioral and tissue levels. SAMP-10 mice were randomized into 2 groups: a control group and a shaking group. The avoidance response (latency) of the mice was evaluated using a passive avoidance task. The degree of amygdala and hippocampal aging was evaluated based on the brain morphology. Subsequently, the association between avoidance response and the degree of amygdala-hippocampal aging was evaluated. <b><i>Results:</i></b> Regarding the passive avoidance task, the shaking group showed a longer latency period than the control group (<i>p</i> < 0.05), even and low intensity staining of ubiquitinated protein, and had a higher number of and larger neurons than those of the control group. The difference between the groups was more significant in the BA region of the amygdala and the CA1 region of the hippocampus (staining degree: <i>p</i> < 0.05, neuron size: <i>p</i> < 0.01, neuron counts: <i>p</i> < 0.01) than in other regions. <b><i>Conclusions:</i></b> The shaking exercise prevents nonfunctional protein (NFP) accumulation, neuron atrophy, and neuron loss; delays the aging of the amygdala and hippocampus; and maintains the function of the amygdala-hippocampal circuit. It thus enhances emotional processing and cognition functions, the memory of threats, the skillful confrontation of threats, and proper self-protection from danger.

本学では、臨床検査技師教育課程において人体解剖実習を導入し、その有効かつ有益な教育方法を考察した。その結果、「心臓の構造を理解できた」と回答した学生は96%であり、他臓器においても80%以上と高率であった。また、医療職として必須の倫理的教育効果についても「ご遺体の尊厳についての理解」に関して96%の学生から肯定的な回答が得られた。一方で、教員の不足等による不満もあったが、今後は大学院生や勉学の意識の強い卒業生の実習参加などを通して、より広くかつ意義深い実習にしていきたいと考えている。(著者抄録)

INTRODUCTION: The disabling effects of dementia, an incurable disease with little effect on mortality, affect society far more than many other conditions. OBJECTIVE: The aim of this study was to stop or delay the onset of dementia using low-cost methods such as physical exercise. METHODS: Senescence-accelerated model-prone (SAMP) 10 mice were made to perform a user-friendly shaking exercise for 25 weeks. The motor function and hippocampal functions (learning, spatial cognition) of the mice were evaluated using behavioral experiments. The degree of hippocampal aging was evaluated based on brain morphology. The association between behavioral performance of the mice and the degree of hippocampal aging was then evaluated. RESULTS: The behavioral test results showed that the shaking group had higher motor coordination (p < 0.01) and motor learning (p < 0.05). Significantly higher performances in the learning ability were observed in the shaking group at a middle-period experiment (p < 0.05); the spatial cognitive functions also improved (p < 0.05). The shaking group showed delayed ageing of cells in the dentate gyrus (DG; area: p < 0.01) and cornu Ammonis (CA; area: p < 0.01) regions of the hippocampus. CONCLUSIONS: The shaking exercise enhances the activity of mice and reduces age-associated decreases in learning and spatial cognitive functions. Regarding hippocampal morphology, shaking exercise can prevent non-functional protein accumulation, cell atrophy, and cell loss. Specifically, shaking exercise protects cell growth and regeneration in the DG area and enhances the learning function of the hippocampus. Furthermore, shaking exercise maintained the spatial cognitive function of cells in the CA3 and CA1 regions, and prevented the chronic loss of CA2 transmission that decreased the spatial memory decline in mice.

© 2020, The Japanese Society for Clinical Molecular Morphology. In the present study, we examined morphology and function of hippocampus in the APC1638T/1638T mouse. Expression levels of the APC mRNA and protein were both identical in the hippocampus of the APC+/+ and APC1638T/1638T mice. The dentate gyrus of the APC1638T/1638T hippocampus was thicker, and has more densely-populated granule cells in the APC1638T/1638T mouse hippocampus. Immunoelectron microscopy revealed co-localization of APC with alpha-amino-3- hydroxy-5-methyl- isoxazole-4-propionate receptor (AMPA-R) and with PSD-95 at post-synapse in the APC+/+ hippocampus, while APC1638T was co-localized with neither AMPA-R nor PSD-95 in the APC1638T/1638T hippocampus. By immunoprecipitation assay, full-length APC expressed in the APC+/+ mouse was co-immunoprecipitated with AMPA-R and PSD-95. In contrast, APC1638T expressed in the APC1638T/1638T mouse was not co-immunoprecipitated with AMPA-R and PSD-95. In the hippocampal CA1 region of the APC1638T/1638T mouse, c-Fos expression after electric foot shock was decreased compared with the APC+/+ mouse. The present study showed some abnormalities on morphology of the hippocampus caused by a truncated APC (APC1638T). Also, our findings suggest that failure in APC binding to AMPA-R and PSD-95 may bring about less activities of hippocampal neurons in the APC1638T/1638T mouse.

Accumulation of amyloid β protein (Aβ) in the brain causes cognitive impairment in Alzheimer's disease (AD). The nature of the relationship between smoking and AD or dementia has been controversial. However, a recent meta-analysis revealed that smoking is a risk factor for AD. With regard to nicotinic acetylcholinergic receptors (nAChRs), both AD and control patients that smoke have been reported to show an increase in 3H-cytisine (an a4β4 nAChR agonist) binding in the temporal cortex. The α7 nAChR is also a key factor in AD pathology, particularly in relation to internalization of Aβs. Furthermore, there are many reports showing the neuroprotective effects of nicotine. The internalization of Aβ may lead to Aβ clearance in the brain. We hypothesized that an extracorporeal system that rapidly removes Aβ from the blood may accelerate Aβ clearance from the brain. We have reported that (1) several medical materials including hemodialyzers can effectively remove blood Aβ, (2) the concentrations of blood Aβs decreased during hemodialysis, (3) removal of blood Aβ enhanced Aβ influx into the blood (ideally from the brain), resulting in maintenance or improvement of cognitive function, and (4) Aβ deposition in the brain of hemodialysis patients was significantly lower than in controls. Smoking affected blood Aβ removal efficiencies and brain atrophy. We believe this Extracorporeal Blood Aβ Removal Systems (E-BARS) may contribute as a therapy for AD.

Recently, health awareness in Japan has been increasing and active exercise is now recommended to prevent lifestyle-related diseases. Cytokine activities have many positive effects in maintaining the health of a number of organs in the body. Myokines are cytokines secreted by skeletal muscles in response to exercise stimulation, and have recently generated much attention. Around 700,000 patients in Japan suffer from rheumatoid arthritis, making it the most prevalent autoimmune disease that requires active prevention and treatment. In the present study, a mouse model of spontaneous arthritis (SKG/Jcl) was subjected to continuous exercise stimulation, starting before the disease onset, to examine the effects of anti-inflammatory and inflammatory cytokine secretion on arthritis. For this stimulation, we developed a device that combines shaking and vibration. The results revealed that exercise stimulation delayed the onset of arthritis and slowed its progression. Thickened articular cartilage and multiple aggregates of chondrocytes were also observed. Further, exercise stimulation increased the expression of IL-6, IL-10, and IL-15, and inhibited TNF-α expression. From these results, we infer that the anti-inflammatory effects of IL-6 and IL-10, which showed increased expression upon exercise stimulation, inhibited the inflammatory activity of TNF-α and possibly delayed the onset of arthritis and slowed its progression. Novel methods for preventing and treating arthritis under clinical settings can be developed on the basis of these findings.

As a proof of concept that removal of blood amyloid-beta (A beta) can reduce A beta deposition in the brains of patients with Alzheimer's disease, cortices of patients who had undergone hemodialysis (HD), which removes A beta from the blood, were histochemically analyzed; postmortem brain sections were stained with anti-A beta antibodies. Brains from patients who had undergone HD had significantly fewer senile plaques than those of patient who had not undergone HD. This significant difference was also confirmed by silver staining. Our findings suggest that removal of blood A beta by hemodialysis results in lower accumulation of A beta in the brain.

In order to elucidate the function of anti-aromatic acid decarboxylase (AADC)-only-positive cells in the alimentary canal, 5-hydroxy-L-tryptophan (5-HTP) or L-3,4-dihydroxyphenylalanine (L-DOPA) was intraperitoneally injected into the laboratory shrew, Suncus murinus, and immunohistochemical studies were conducted on continuous sections of the alimentary canal using specific antisera against tyrosine hydroxylase (TH), AADC, dopamine (DA), and serotonin (5-HT). AADC-only-positive cells localized to the epithelial layer of the alimentary canal from the stomach to the large intestine. These AADC-only-positive cells became DA- and AADC-positive cells after L-DOPA injection, and 5-HT- and AADC-positive cells after 5-HTP injection. These results strongly indicate that the AADC-only-positive cells in the alimentary canal of Suncus murinus are capable of synthesizing DA and 5-HT simultaneously upon administration of L-DOPA and 5-HTP.

Osteoporosis is leaving bones more fragile and susceptible to fracture. It has a massive impact, both physically and mentally, markedly diminishing quality of life. A new form of therapeutic exercise or physical therapy that mitigates the abrupt decrease in bone density in postmenopausal women must quickly be developed to avoid those problems. In this study, ovariectomy (OVX) mice were used as models to simulate the decrease in bone density observed in postmenopausal women. Physical therapy via a shaking stimulus, in the form of moving a platform that rotates in a roughly circular motion in the horizontal plane, was studied as a way to prevent the decrease in bone density of the lumbar vertebrae by analysis of bone histomorphometry, a feat that the stimulus from conventional therapeutic exercise and physical therapy have failed to achieve. Comparison of the stimulus/ovariectomized (+/+) group with the -/+ group indicated significant increases in ES (P &lt 0.01), N. Mu. Oc (P &lt 0.05), OV (P &lt 0.05), O. Th (P &lt 0.01), and L. Th (P &lt 0.01) in the +/+ group. If this finding is used clinically, we believe that it could lead to therapy that would prevent compression fractures of the lumbar vertebrae.

Background: Loss of adenomatous polyposis coli (APC) gene function results in constitutive activation of the canonical Wnt pathway and represents the main initiating and rate-limiting event in colorectal tumorigenesis. APC is likely to participate in a wide spectrum of biological functions via its different functional domains and is abundantly expressed in the brain as well as in peripheral tissues. However, the neuronal function of APC is poorly understood. To investigate the functional role of Apc in the central nervous system, we analyzed the neurological phenotypes of Apc(1638T/1638T) mice, which carry a targeted deletion of the 3' terminal third of Apc that does not affect Wnt signaling. Results: A series of behavioral tests revealed a working memory deficit, increased locomotor activity, reduced anxiety-related behavior, and mildly decreased social interaction in Apc(1638T/1638T) mice. Apc(1638T/1638T) mice showed abnormal morphology of the dendritic spines and impaired long-term potentiation of synaptic transmission in the hippocampal CA1 region. Moreover, Apc(1638T/1638T) mice showed abnormal dopamine and serotonin distribution in the brain. Some of these behavioral and neuronal phenotypes are related to symptoms and endophenotypes of schizophrenia. Conclusions: Our results demonstrate that the C-terminus of the Apc tumor suppressor plays a critical role in cognitive and neuropsychiatric functioning. This finding suggests a potential functional link between the C-terminus of APC and pathologies of the central nervous system.

Background and aims Elderly individuals who suffer a fracture develop a gait disturbance and require prolonged bedrest. A fracture has a massive impact both physically and mentally and markedly diminishes quality of life. A new form of therapeutic exercise that mitigates the abrupt decrease in bone density in postmenopausal women must soon be developed so that those problems can be avoided. Methods The current study used a model of the decrease in bone density in ovariectomized mice to simulate postmenopausal women. The stimulus was provided by a shaking horizontal platform rotating in a circular motion. Results Comparison of the +/+ (ovariectomized/stimulated) group and +/- group indicated a significant decrease in BV/TV (p &lt; 0.01), Tb.Th (p &lt; 0.01), and Tb.N (p &lt; 0.05) in the +/+ group and a significant increase in OV/BV (p &lt; 0.01), OV/ OS (p &lt; 0.01), BFR/BV (p &lt; 0.01), dLS/BS (p &lt; 0.05), MS/BS (p &lt; 0.05), BRs.R (p &lt; 0.01), and Tb.Sp (p &lt; 0.01) in the +/+ group. Physical therapy to prevent a decrease in bone density was studied via stimulus in the form of rotation of a platform. Analysis of bone histomorphometry revealed lessening of the decrease in bone density of the lumbar vertebrae, a feat that the stimulus from conventional physical therapy had failed to achieve. Conclusion The current study delivered a shaking stimulus to mice in a model of postmenopause. Analysis of bone histomorphometry of the lumbar vertebrae suggested lessening of the abrupt decrease in bone density of trabecular bone. If this finding is used clinically, it could lead to physical therapy exercise that would be able to prevent compression fractures of the lumbar vertebrae.

After finding tonsil-like structures near the entrance of vagina of a laboratory shrew (Suncus murinus), which we subsequently designated as vaginal tonsils, we performed detailed immunohistochemical and developmental studies. The location of T and B cells in the vaginal tonsils differed from that in the palatine tonsils or that in the lymphoid nodes of other animals. The boundary between the germinal center region and the region encompassing follicular interfollicular tissue was not clear. B cells were widely distributed and very dense in the parenchyma, but they were scattered in the epithelial area (B cells were present in around 90% of the vaginal tonsil tissue). In contrast, T cells were scattered in the parenchyma and in the epithelial area (T cells were present in around 10% of the vaginal tonsil tissue). B cells were more prominent than T cells throughout the development of these structures and the epithelium was invaded by many immigrating cells. The size of the vaginal tonsils changed during postnatal development. Vaginal tonsils are structurally similar to other tonsils, and they may function to protect the vagina from infection.

We investigated the distribution of T lymphocytes, B lymphocytes, and S-100 protein-immunoreactive dendritic-like in the anal tonsil of the laboratory shrew, Suncus murinus. In adult animals, T lymphocytes were located mainly at the periphery of the anal tonsil, especially around small blood vessels. B lymphocytes were located in the central and subepithelial region of the anal tonsil, which includes primary lymphoid follicles, and in which there are small numbers of scattered T lymphocytes. B and T lymphocytes were distributed over 72.7 and 27.3% of the tonsillar area, respectively. However, their areas of distribution were not clearly distinguished. The areas containing B lymphocytes were enriched in S-100 protein antibody-immunoreactive cells, which exhibited a dendritic shape. These S-100-positive cells appeared to be identical to the follicular dendritic cells (FDC) seen in the follicles of lymphoid organs. These results suggest that the anal tonsils constitute one of the gut-associated lymphoid tissues (GALT), and that a function of the anal tonsil includes the capture of intruding antigens that would generate protective antibody responses. Microsc. Res. Tech. 74: 819-824, 2011. (C) 2010 Wiley-Liss, Inc.

A morphological and immunohistochemical study of the ultimobranchial body of reptiles Japanese lizard and snake was carried out. The ultimobranchial body of the Japanese lizard was located adjacent to the left arch of the aorta between the trachea and esophagus. It was found as a cluster or group of cells with no capsule. Grimelius' silver impregnation and lead-hematoxylin staining produced positive reactions in some of the clustered cells and follicular cells. The same reaction pattern was observed with anti-calcitonin using the PAP method. The PAP reactions were positive to antiserum against pig calcitonin, but negative to antiserum against synthesized human calcitonin. Furthermore, the PAP reactions were negative to antiserum against tyrosine hydroxylase. The immunofluorescent study of the snake ultimobranchial body revealed that most of the clustered cells and some of the follicular cells were calcitonin-immunoreactive but none was tyrosine hydroxylase-immunoreactive. Certain histological similarities exist between the Japanese lizard ultimobranchial body and snake ultimobranchial body, but the distribution of calcitonin-positive cells were slightly different. In the Japanese lizard, the positive cells were scattered between the folicles and the number was small. However, most of the cells which formed the cluster in the ultimobranchial body of snake were positive. The findings suggest that the configuration of amino acid in the Japanese lizard calcitonin and snake calcitonin are similar to that of pig calcitonin, and the reptile and the birds is a boundary of the tyrosine hydroxylase existence.

Circulating immune complex (CIC) is known to play a role in pathological glomerular alterations in malaria. However, the nature of the antigens comprising the CIC is still not fully understood. We report here the isolation of the antigen in CIC and its localisation in mice infected with Plasmodium berghei NK65. The antigen was successfully isolated from CIC extracted from the blood of mice infected with P. berghei, by using Clq-coated microplates. The molecular mass of the antigen separated from CIC bound to Clq was found to be 78 kDa. Furthermore, localisation of the antigen was examined by the fluorescent antibody technique and immunoelectron microscopy. The antigen was detected in the parasitised erythrocyte and the mesangial matrix by both methods. These results suggest that the 78 kDa protein might be associated with the glomerular alterations in malaria infection. (C) 2000 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.

Changes in the parafollicular cells in the thyroid gland of orariectomized rats were investigated to clarify the relationship between the secretory function of sex hormones and that of parafollicular cells. Compared with control rats, the ovariectomized rats exhibited decreases in 1) the number of parafollicular cells in the thyroid glands, 2) the number of secretory granules in the parafollicular cells, and 3) the area occupied by the Golgi complex. These results suggest that the lack of estrogen caused by ovariectomy reduces the synthesis and release of calcitonin in parafollicular cells, which may be one of the causes of osteoporosis.

We detected the transient coexistence of tyrosine hydroxylase (TH) and choline acetyltransferase (CAT) in the neurons of the dorsal motor nucleus of the vagus (DMV) and in cholinergic fibers of the myenteric plexus of the stomach of the laboratory shrew (Suncus murinus) by immunohistochemistry. The DMV neurons showed no reactivity to antiserum against GTP cyclohydrolase I, DOPA, aromatic L-amino acid decarboxylase or dopamine. After birth, the intensity of TH-immunoreactivity of the DMV neurons reached a maximum on postnatal days 7-9 and then decreased until postnatal day 20, when almost no TH-immunoreactivity was detected. These results suggest that TH may function not as a catecholamine-synthesizing enzyme but as a regulatory protein necessary for release acceleration or inhibition in the postnatal development of cholinergic neurons of the DMV.

The catecholaminergic neurons of senescence-accelerated mice (SAM-P8) were analyzed by immunohistochemical microphotometry in terms of immunoreactivities to aromatic L-amino acid decarboxylase (AADC), dopamine (DA), or noradrenaline (NA). Accelerated senescence-resistant mice (SAM-RI) were used as control mice. The immunoreactivities to AADC, DA, and NA of the catecholaminergic neurons of the SAM-Pa mice were weaker than those of the SAM-RI mice in all the brain regions. Immunoelectron microscopy revealed progressive degeneration of dopaminergic neurons and their terminal fibers in the substantia nigra as well as in noradrenergic neurons and their proximal dendrites in the locus coeruleus of the SAM-P8 mice. In contrast, there was no difference between the SAM-P8 and SAM-RI mice in the distribution of AADC-only positive neurons (designated as D neurons in the rat brain by Jaeger et al.) nor in their immunoreactivities. These results may indicate that DA neurons in the substantia nigra and NA neurons in the locus coeruleus degenarate more rapidly during aging in SAM-P8 mice than in control SAM-RI mice and that D neurons may function as a part of a compensatory system for the decreases in catecholaminergic neurons during aging.

The distribution of monoamine oxidase (MAO)-containing neuronal somata was studied in the grass parakeet (Melopsittacus undulatus) brain by using a histochemical technique involving the coupled peroxidatic oxidation method, and compared with the distribution of the tyrosine hydroxylase-, aromatic L-amino acid decarboxylase-, serotonin-immunoreactive cells in the grass parakeet brain. MAO-containing neurons were located in the nucleus locus coeruleus, around the nucleus n. facialis (nVII), lateral region to the nucleus olivaris inferior and nucleus tractus solitarius, where noradrenaline neurons exist. Another group was located in the ventral region of nervi oculomotorii, caudal region of nucleus n. oculomotorii (nIII), the nucleus raphe and ventral to the nucleus olivaris superior, where serotonin neurons exist. A third group was the non-monoaminergic neurons which were located in the lobus parolfactorius, pareostriatum augmentatum and ventrolateral part of formatio reticularis mesencephali.

We investigated the localization of APUD (amine content and amine precursor uptake and decarboxylation) cells and tyrosine hydroxylase (TH)-negative, aromatic L-amino acid decarboxylase (AADC)-positive neurons (D neurons) in the hypothalamus of the bird (Melopsittacus undulatus) and the lizard (Varanus exanthematicus), and compared the differences between the two species, L-DOPA was demonstrated in the paraventricular organ (PVO) which is a characteristic region in the hypothalamus of the bird and the lizard. In the PVO of the grass parakeet, there were a small number of dopamine neurons in addition to APUD cells. However, in the PVO of the lizard, there were APUD cells but no dopamine neurons, Furthermore, TH-negative, AADC-positive neurons (D neurons) were found in the medial part of nucleus arcuatus of the bird, but not of the lizard.

After 5-hydroxy-L-tryptophan (5-HTP) and L-3,4-dihydroxyphenylalanine (L-DOPA) were injected i.p. in the laboratory shrew Suncus murinus, immunocytochemical and immunofluorescence studies were conducted on continuous or same sections of the brain, using specific anti-tyrosine hydroxylase (TH), anti-aromatic L-amino acid decarboxylase (AADC), anti-dopamine (DA) and antiserotonin (5-HT) antisera which were produced in our laboratory. The results of double-staining by the immunofluorescence method as well as immunoelectron microscopy strongly indicate that the cells of the premammillary nucleus of the laboratory shrew brain (AADC-only-positive neurons) are capable of synthesizing DA and 5-HT simultaneously upon simultaneous administration of L-DOPA and 5-HTP.

L-DOPA immunoreactivity was demonstrated in neurons of the house-shrew brain, using an immunocytochemical method in conjunction with a specific anti-L-DOPA serum. The neurons in the ventrolateral area of arcuate nuclei were tyrosine hydroxylase- and L-DOPA-immunoreactive, but aromatic L-amino acid decarboxylase- and dopamine-negative. Most of these neurons were small, round or oval in shape, and ranged from 10 to 20 mum in diameter with their long axonal processes extended within the third ventricular wall. By immunoelectron microscopy, L-DOPA-positive products were observed throughout the cytoplasmic matrix and its terminal processes which contained small (60 to 80 nm) or large (140 to 160 nm) vesicles.

A morphological study of parafollicular cells in the thyroid gland and parathyroid gland of the house shrew (Suncus murinus) was made. The results indicated that (1) there were two pairs of parathyroid glands which were located in the upper part of the house shrew thyroid gland, (2) the volumes of the house shrew parathyroid glands ranged from 0.014 to 0.079 mm3, (3) the number of parafollicular cells along the follicles was largest in the upper part of the thyroid lobe, while no parafollicular cells were present in the isthmus, and (4) there were about 20 parafollicular cells per 100 follicular cells and 1.519 parafollicular cells per follicle. The number of parafollicular cells per 100 follicular cells was thus about 5 times larger than that in rats and the number of parafollicular cells per follicle about 2.5 times larger than that in rats. © 1992, Editorial Board of Okajimas Folia Anatomica Japonica. All rights reserved.