Kenji Yoshimi

Caspase-11 Mediates Inflammatory Dopaminergic Cell Death in the 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Mouse Model of Parkinson's Disease

The present study was designed to elucidate the inflammatory and apoptotic mechanisms of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neurotoxicity in a model of Parkinsons disease. Our results showed that mutant mice lacking the caspase-11 gene were significantly more resistant to the effects of acute treatment with MPTP than their wild-type mice. Thus, the neurotoxicity of MPTP seems to be mediated by the induction of both mitochondrial dysfunction and free radical generation. Previously, we showed that overexpression of the Apaf-1 dominant-negative inhibitor inhibited the mitochondrial apoptotic cascade in chronic MPTP treatment but not in acute MPTP treatment. The present results indicate that MPTP neurotoxicity may be mediated via activation of the caspase-11 cascade and inflammatory cascade, as well as the mitochondrial apoptotic cascade.

Cardiac Sympathetic Rejuvenation: A Link Between Nerve Function and Cardiac Hypertrophy

Neuronal function and innervation density is regulated by target organ-derived neurotrophic factors. Although cardiac hypertrophy drastically alternates the expression of various growth factors such as endothelin-1, angiotensin II, and leukemia inhibitory factor, little is known about nerve growth factor expression and its effect on the cardiac sympathetic nerves. This study investigated the impact of pressure overload-induced cardiac hypertrophy on the innervation density and cellular function of cardiac sympathetic nerves, including kinetics of norepinephrine synthesis and reuptake, and neuronal gene expression. Right ventricular hypertrophy was induced by monocrotaline treatment in Wistar rats. Newly developed cardiac sympathetic nerves expressing β3-tubulin (axonal marker), GAP43 (growth-associated cone marker), and tyrosine hydroxylase were markedly increased only in the right ventricle, in parallel with nerve growth factor upregulation. However, norepinephrine and dopamine content was paradoxically attenuated, and the protein and kinase activity of tyrosine hydroxylase were markedly downregulated in the right ventricle. The reuptake of [125I]-metaiodobenzylguanidine and [3H]-norepinephrine were also significantly diminished in the right ventricle, indicating functional downregulation in cardiac sympathetic nerves. Interestingly, we found cardiac sympathetic nerves in hypertrophic right ventricles strongly expressed highly polysialylated neural cell adhesion molecule (PSA-NCAM) (an immature neuron marker) as well as neonatal heart. Taken together, pressure overload induced anatomical sympathetic hyperinnervation but simultaneously caused deterioration of neuronal cellular function. This phenomenon was explained by the rejuvenation of cardiac sympathetic nerves as well as the hypertrophic cardiomyocytes, which also showed the fetal form gene expression.

An immunohistochemical study of MAP2 and clathrin in gerbil hippocampus after cerebral ischemia.

Changes in MAP2 and clathrin immunoreactivity were studied in gerbil hippocampus after transient cerebral ischemia. MAP2 immunoreactivity decreased significantly by 1 h in the subiculum-CA1 and CA2 areas which correspond to reactive change, while no decrease was observed in CA1 until day 4. Before the initiation of delayed neuronal death, MAP2 immunoreactivity was not changed in CA1. On the other hand clathrin immunoreactivity increased in the pyramidal cell layer of CA1 by 3 h after ischemia and remained high for 2 days. Clathrin immunoreactivity in the pyramidal cell layer of CA1 diminished after delayed neuronal death. The transient change of clathrin was noted especially in CA1 in the period prior to delayed neuronal death. These results imply an abnormal change in clathrin turnover after ischemia, which may participate in the pathogenesis of delayed neuronal death.

IgG-immunostaining in the intact rabbit brain: variable but significant staining of hippocampal and cerebellar neurons with anti-IgG.

A significant number of brain neurons in the rabbit brain were immunostained with anti-rabbit gamma-immunoglobulin (IgG). IgG-positive neurons were often found in the cerebellum, lower brainstem and motor nuclei. Similar IgG-positive neurons were occasionally found in the hippocampus, cerebral cortex and midbrain, but not in the striatum and thalamus. These neurons showed very clear Golgi-like staining of soma and dendrites but IgG staining was absent from the cell nuclei and axons. In particular, groups of Purkinje neurons in the rabbit cerebellum showed strong IgG-positive staining. To confirm whether the staining reflected the existence of IgG molecules in these neurons, staining specificity was carefully evaluated. Staining was specifically eliminated by pre-absorption of the antibodies with the purified rabbit IgG. An antibody to the neural cell adhesion molecule (NCAM or CD56), a member of the immunoglobulin superfamily, exhibited a completely different pattern of staining as that for IgG. To determine whether IgG-like immunoreactivity was a general feature of mammalian brain, brain sections of rabbits, rats, and mice were immunostained with antibodies to IgGs of each of the three species. Similar IgG-positive neurons were observed in all three species, although the distribution and frequency was characteristic for each species. In rabbit brain, anti-rabbit IgG stained-neurons were more abundant compared to rat and mouse brain. IgG-positive microglia-like cells were evident in mouse brain, but less frequent in rabbit and were hardly observed in rat brain. To evaluate whether stained neurons could synthesize IgG, in situ hybridization was carried out using an antisense oligonucleotide probe to rabbit IgG DNA. No significant label was observed in cerebellum. These results suggest that a significant number of neurons in the intact rabbit brain take up IgGs and concentrate them in their cytoplasm, although the molecular uptake mechanism is retained for future studies. Our results also suggest that the rabbit may be a suitable animal to study the function(s) of IgG in brain neurons.

Involvement of clathrin light chains in the pathology of Alzheimer's disease

Clathrin, which constitutes coated vesicles, plays important roles in neuronal functions. In the brains of the patients with Alzheimers disease, distribution of clathrin was immunohistochemically investigated using four monoclonal antibodies against clathrin light chains, LCB.1, LCB.2, X-16 and CON.1, to study the involvement of clathrin in the pathology of Alzheimers disease. LCB.1, LCB.2, X-16, and CON.1 bind to the aminoterminus of the clathrin light chain b(LCb), to the neuron-specific insert of LCb, to the light chain a(LCa), and to LCa and LCb, respectively. In Alzheimer brains, granular staining of LCB.2 around neurons in the hippocampus was weaker or patchily defected in comparison with control brains. Some neurofibrillary tangles and neurons were intensely stained in Alzheimer brains by LCB.2, whereas neurons were weakly stained in control brains. Crowns of some senile plaques in the brains of early onset Alzheimers disease were positively stained by LCB. 2. LCB. 1 supported the observations of LCB.2. Reactive astrocytes in Alzheimer brains were intensely stained by X-16. On the other hand, Western blot analysis using LCB.2 and X-16 demonstrated no apparent differences in protein amounts and molecular weights of LCa and LCb between control and Alzheimer brains. These observations demonstrated abnormal distribution of clathrin in Alzheimer brains, implying impairment of axonal transport in this disease.

Involvement of clathrin light chains in the pathology of Pick's disease; impilication for impairment of axonal transport

Clathrin, which constitutes coated vesicles and plays important roles in neuronal functions, has been reported to be involved in the pathology of Alzheimers disease. In the brains of the patients with Picks disease, distribution of clathrin was immunohistochemically investigated using monoclonal antibodies binding to different epitopes of clathrin light chain a and b. All the antibodies intensely labeled Picks body and some perikarya of neurons, indicating impairment of slow axonal transport b (SCb). Antibodies against neurofilament, kinesin and synaptophysin also labeled Picks body. These observations suggested impairment of axonal transport in the brains with Picks disease, and might contribute to elucidating the pathology of Picks body forming. It is implied that common pathological processes might lie in Alzheimers disease and Picks disease.

Ischaemia-induced change in clathrin preceding delayed neuronal death.

The mechanism underlying the change in clathrin immunohistochemistry preceding delayed neuronal death (DND) was studied in gerbils. The ischaemic change observed with chc5.9 anti-clathrin antibody in hippocampal CA1 was initially ameliorated by pentobarbital, which blocks DND, but 1 day after ischaemia, no change in the immunoreactivity of the SDS-denatured clathrin molecule was detected by Western blotting and no change in the clathrin content in CA1 was detected by SDS-PAGE. No ischaemia-induced change in immunohistochemistry was observed with another monoclonal anti-clathrin antibody, X-22. The above results imply that some modifications that affect the structure of clathrin molecules around the chc5.9 specific epitope may be a crucial step in the course of DND.