Hideaki Ishiguro

Changes of growth inhibitory factor after stab wounds in rat brain

The growth inhibitory factor (GIF) is a new metallothionein (MT)-like protein that is downregulated in Alzheimers disease (AD) brain. The biological function of GIF has not been fully clarified yet. We have raised an antibody to the synthetic polypeptide that is specific for rat GIF. The purified antibody reacted to recombinant GIF and native rat GIF but not to MT or maltose-binding protein. Using the antibody and GIF cDNA probe, we investigated changes of GIF and GIF mRNA by Western and Northern blotting techniques in rat brains after stab wounds. The levels of GIF and GIF mRNA began to increase 4 days postoperation, reached a maximum at 14-21 days and sustained the increased level at least through 28 days. While both glial fibrillary acidic protein (GFAP) and GIF were recognized in astrocytes, the increases of these 2 proteins after stab wounds showed different patterns. The results indicated that GIF could play an important role in the repair after brain damage and also produce new insights into the mechanism of gliosis investigated mainly from the viewpoint of GFAP.

Expression of growth inhibitory factor (GIF) in normal and injured rat brains

Immunohistochemical study on growth inhibitory factor (GIF) in rat brain has revealed that a glial cell layer on the surface of cerebral cortex and the cells surrounding Purkinje cells has been reported. In addition, neurons in gray matter were weakly immunostained for GIF. In situ hybridization using digoxigenin-labeled single-strand RNA probes also demonstrated that most of the neurons and small round cells, which were presumably astrocytes, expressed GIF mRNA in the cerebral cortex of rat brain. These findings indicate that GIF is produced in neurons as well as in astrocytes. The most prominent findings in this study are, a very strong reaction of GIF and GIF mRNA in the reactive astrocytes around the site of injury induced by stab wound or kainic acid injection. These results raised the possibility that GIF may act as an acute-phase protein in reactive astrocytes and have a role in tissue repair.

Immunoreactivity of growth inhibitory factor in normal rat brain and after stab wounds — an immunocytochemical study using confocal laser scan microscope

The growth inhibitor factor (GIF) is a new member of the metallothionein family that is downregulated in Alzheimers disease brain. Using a confocal laser scan microscope with polyclonal and monoclonal antibodies to GIF, and monoclonal antibodies to glial fibrillary acidic protein (GFAP) and MAP-2, we demonstrated that GIF immunoreactivity was expressed primarily in astrocytes and much less in neurons. In astrocytes of normal rat brain GIF immunoreactivity was detected mainly in the cell bodies, while GFAP immunoreactivity was detected mainly in the processes. GIF immunoreactivity was more strongly expressed in reactive astrocytes. These findings were confirmed with both polyclonal and monoclonal antibodies. Following stab wounds, a number of GIF-positive reactive astrocytes were detected around the wounds at 3 days postoperation. After 7 days GIF immunoreactivity was detected in cell bodies and processes of reactive astrocytes. The number of GIF-positive astrocytes and the intensity of the immunoreactivity remained elevated over the control levels at least through 28 days. These immunocytochemical findings correlated well with changes in GIF protein and mRNA levels. Not only changes in GIF protein and mRNA levels but also intracellular localization of GIF in normal rat brain and after stab wounds in rat brain were different from those of GFAP. These results support the concept that GIF plays an important role in the processing of reconstruction after brain damage.

Expression of the large myelin-associated glycoprotein isoform during the development in the mouse peripheral nervous system

The developmental maximum expression of the large myelin-associated glycoprotein isoform (L-MAG) protein prior to that of the small myelin-associated glycoprotein isoform (S-MAG) in both the central and peripheral nervous systems (CNS, PNS) in mice was shown by immunoblotting techniques using specific antibodies to the L-MAG protein and the S-MAG protein. Both the L-MAG protein and the S-MAG protein were expressed earlier in the PNS than in the CNS, which reflects earlier myelination in the PNS. The peak of the L-MAG protein concentration was 8 days in the sciatic nerve and 15 days in the brainstem. The concentration of the S-MAG protein in the sciatic nerve reached a peak at 15 days, whereas in the brainstem it increased rapidly between 15 and 20 days and gradually thereafter. Thus, the preceding maximum expression of the L-MAG during active myelination in the PNS demonstrated here as well as in the CNS strongly suggests an important role for L-MAG in myelin formation.

The Large Isoform of Myelin-Associated Glycoprotein Is Scarcely Expressed in the Quaking Mouse Brain

Abstract: Two polypeptide isoforms of myelin‐associated glycoprotein (MAG) with molecular masses of 72 and 67 kDa are produced by alternative splicing of the exon 12 portion. Our previous work has demonstrated that in the quaking mouse brain this alternative splicing is lacking and that the mRNA coding the large MAG isoform (L‐MAG) is scarcely expressed, whereas that of small MAG isoform (S‐MAG) is overexpressed. In the present study, we prepared antisera specific to the S‐MAG and L‐MAG amino acid residues, respectively. Immunoblots showed that the L‐MAG band was scarcely detectable in the quaking mouse brain, whereas the S‐MAG band had an apparently higher molecular mass than in the normal control. Our immunohistochemical study also showed that L‐MAG was scarcely stained in the quaking mouse brain. These results seemed to reflect a reduction in content of L‐MAG mRNA and abnormal glycosylation in the quaking mouse brain.

Induction of myelin-associated glycoprotein mRNA in experimental remyelination

Two forms of myelin-associated glycoprotein (MAG) mRNA are produced by alternative splicing of the exon 12 portion. Expression of the two forms of MAG mRNA was studied here in experimentally introduced demyelination and remyelination by Cuprizone intoxication. During the demyelinating stage, both forms of MAG mRNA decreased markedly. When feeding with Cuprizone was stopped, MAG mRNA began to increase. One form of MAG mRNA without the exon 12 portion, which appears in normal development at the period of active myelination, was characteristically induced during the remyelinating stage. The other form containing the exon 12 portion was also induced but recovered only to the level in normal development.

Immunohistochemical localization of myelin-associated glycoprotein isoforms during the development in the mouse brain ☆

The developmental changes in localization of myelin-associated glycoprotein (MAG) isoforms in the mouse brain were demonstrated by an immunohistochemical method using antisera specific to two MAG isoforms. The antiserum to the large isoform of MAG (L-MAG) stained the myelin sheaths and the cytoplasm of oligodendroglia in the active myelinating stage in the mouse central nervous system. However, the antiserum to the small isoform of MAG (S-MAG) stained only myelin sheaths in the adult stage. These findings suggest that L-MAG plays an important role in active myelination.

Cerebral Vasculopathy Showing Moyamoya-Like Changes in a Patient with CREST Syndrome

Cerebral Vasculopathy Showing Moyamoya-Like Changes in a Patient with CREST Syndrome Kenshi Terajima a, c, Takayoshi Shimohata a, c, Masatoshi Watanabeb, Takashi Suzuki a, Akira Hasegawa b, Hideaki Ishiguro a, Takashi Minakawa b, Kouichi Hirota a Departments of aNeurology and bNeurosurgery, Akita Red Cross Hospital, and cDepartment of Neurology, Brain Research Institute, Niigata University, Niigata, Japan

Expression of myelin-associated glycoprotein isoforms after sciatic nerve crush injury in mice

The large myelin-associated glycoprotein isoform (L-MAG) protein and small myelin-associated glycoprotein isoform (S-MAG) protein were demonstrated after sciatic nerve crush injury in mice by an immunoblotting technique using specific antibodies to the L-MAG protein and the S-MAG protein, respectively. Immunoblots indicated a rapid decrease in expression of both isoform proteins in the crushed sciatic nerves to < 10% of the control side. By 13 d after injury, L-MAG protein expression had quickly recovered to 100% of the control level. Following the increase in L-MAG protein expression, S-MAG protein expression recovered to 100% by 20 d after injury. It has been reported that the developmental maximum expression of L-MAG protein precedes that of S-MAG protein in both central and peripheral nervous system (CNS and PNS). Our previous work demonstrated that L-MAG mRNA was characteristically induced at the time of most active myelination, including remyelination in the CNS. We here have shown the expression of L-MAG protein precedes that of S-MAG protein during active remyelination in the PNS. This suggests that it plays an important role in the early stage of myelin formation.

A new Japanese amyotrophic lateral sclerosis family with TARDBP (TDP-43) mutation

To date, more than 35 mutations of the transactivation response element DNA-binding protein gene (TARDBP) have been identified in autosomal dominant familial amyotrophic lateral sclerosis (FALS) and simplex cases of ALS. Here, we describe a new Japanese ALS family with a novel nucleotide substitution of TARDBP (Fig. 1a). A 62-year-old Japanese man (individual III-9) presented with three-month history of difficulty in moving his right hand. Neurological examination revealed dysarthria, rightdominant muscle weakness with muscular atrophy, and hyperreflexia in both the upper and the lower limbs. He was diagnosed with probable ALS according to the Awaji criteria. Although he developed bulbar palsy and muscle weakness of the extremities, he remains free from respiratory failure 22 months after the onset. The proband’s first cousin (III-1) noticed dysarthria and dysphasia at the age of 72 years. Four months later, she was diagnosed with progressive bulbar palsy. Subsequently, she developed right-dominant muscle weakness and died of respiratory failure 11 months after the onset of symptoms. Cognitive impairment, behavioral symptoms, and frontotemporal atrophy on brain MRI were not observed in both patients. The proband’s mother (II-5) had remained free from any neurological