Kiyotaka Tomiwa

Tentorial cavernous angioma with calcification in a neonate.

A cavernous angioma of the tentorium cerebelli, first disclosed by perinatal serial ultrasonographic studies, was extirpated totally without remarkable neurological deficit in a neonate. The tumor was accompanied by a calcified expansive hematoma in the posterior fossa. To our knowledge, this is the first case not only of cavernous angioma treated surgically and verified histologically in a neonate, but also of a calcified hematoma revealed on a conventional skull x-ray film at birth. This case suggests the possibility of hemorrhage from intracranial cavernous angioma early in life, even prenatally, and emphasizes the necessity for early diagnosis and early treatment of intracranial cavernous angiomas.

Aristaless -related homeobox gene disruption leads to abnormal distribution of GABAergic interneurons in human neocortex: evidence based on a case of X-linked lissencephaly with abnormal genitalia (XLAG)

X-linked lissencephaly with abnormal genitalia (XLAG) is a rare disorder caused by mutations in the aristaless-related homeobox (ARX) gene, located on Xp22.13. Arx-null mice show loss of tangential migration of GABAergic interneurons, presumably being related to caudal ganglionic eminence tangential migration. In the present study, we investigated a subpopulation of GABAergic interneurons in the brain of an infant with XLAG, who had a novel nonsense mutation of the ARX gene, compared with those of age-matched normal controls and Miller–Dieker syndrome. We performed immunocytochemistry for interneuron and migration markers. We found that glutamic acid decarboxylase (GAD)- and calretinin (CR)-containing cells were significantly reduced in the neocortex and located in the white matter and neocortical subventricular zone, while neuropeptide Y- or cholecystokinin-containing cells were normally distributed. Moreover, in the neocortical subventricular region, the GAD- and CR-containing cells expressed the radial migration marker Mash-1 as well as nestin. Our findings suggest that ARX protein controls not only the tangential migration of GABAergic interneurons from the ganglionic eminence, but also may serve to induce radial migration from the neocortical subventricular zone.

NEUROTOXICITY OF VINCRISTINE AFTER THE OSMOTIC OPENING OF THE BLOOD‐BRAIN BARRIER

The direct effect of intravascularly injected vincristine on the rat brain was investigated after the osmotic opening of the blood‐brain barrier. This was opened unilaterally by injecting 2–5 ml of 1–4 molal mannitol solution into the right carotid artery through a cannula inserted in the external carotid artery in retrograde fashion. Vincristine (0–75 mg/kg) was then injected into either the carotid artery or into a vein. Animals, thus treated, developed left hemiparesis and choreo‐athetoid head motion within several days. Microscopical analysis revealed 1 intracytoplasmic eosinophilic inclusions at an early period, 2 neuronal argyrophilic change and axonal thickening with spheroid formation at later times, and 3 glial mitotic arrest, a finding not reported in the previous studies with intrathecal injection of vincristine. Neuronal changes were usually confined to the selected area in the midbrain, whereas glial mitotic figures were seen in the hippocampus, midbrain and cerebellum. These changes are thought to be direct effects of the vincristine binding with cellular microtubules, including mitotic spindle tubules and neurotubules. The transient opening of the blood‐brain barrier seems to be a useful technique for studying the experimental neurotoxicology of drugs to which the blood‐brain barrier is impermeable.

Early Television Exposure and Children’s Behavioral and Social Outcomes at Age 30 Months

Background Previous research has suggested that television (TV) viewing may be associated with increased behavioral and emotional problems in children. However, there are few prospective studies targeted for its association with outcomes of children under 3 years old. The purpose of this study was to exam the association between children’s early TV exposure at ages 18 and 30 months and the behavioral and emotional outcomes at age 30 months. Methods We analyzed data collected prospectively in the Japan Children’s Study. TV exposure was assessed by mothers’ report at infant ages of 18 and 30 months. The outcomes were assessed using the Strengths and Difficulties Questionnaire (SDQ). Analysis of Covariance was used to estimate the effect of TV exposure on behavioral and emotional outcomes. Results The percentage of children who watched TV 4 hours or more per day was 29.4% at age 18 months, 24.5% at age 30 months, and 21% at both ages. Hyperactivity–inattention at age 30 months was positively associated with TV exposure at age 18 months, whereas prosocial behavior was negatively associated with hours of exposure even after adjustment. However, there were no significant differences in SDQ subscales according to daily hours of TV viewing at age 30 months. Conclusions Daily TV exposure at age 18 months was associated with hyperactivity–inattention and prosocial behavior at age 30 months. However, the directly casual relation was not proved in the present study. Additional research considering the TV program content and exposure timing are needed to investigate the causal relation between TV viewing and behavioral outcome.

REVERSIBLE OSMOTIC OPENING OF THE BLOOD‐BRAIN BARRIER Prevention of Tissue Damage with Filtration of the Perfusate

Histological and fluoromicroscopical studies were performed in order to obtain information about reversibility and associated tissue damage of an osmotic opening of the blood‐brain barrier. Three ml of 1.4 M mannitol sulution were perfused through the right carotid artery of a rat, and the opening of the blood‐brain barrier was examined by using Evans blue as a tracer. The barrier was opened for 60–120 minutes and then reestablished without gross neurological defect. Microscopically, however, edematous change and microinfarcts were often observed, which might be due to microembollsm of recrystallized mannitol. With filtration of the perfusate through a millipore filter, the blood‐brain barrier was reversibly opened without any tissue damages. This could be a useful therapeutic technique and an experimental model for neurotoxicology. ACTA PATHOL. JPN. 32: 427∼435, 1982.

Cloning of human cDNAs for Apg-1 and Apg-2, members of the Hsp110 family, and chromosomal assignment of their genes.

In mice, the Hsp110/SSE family is composed of the heat shock protein (Hsp)110/105, Apg-1 and Apg-2. In humans, however, only the Hsp110/105 homolog has been identified as a member, and two cDNAs, Hsp70RY and HS24/p52, potentially encoding proteins structurally similar to, but smaller than, mouse Apg-2 have been reported. To clarify the membership of Hsp110 family in humans, we isolated Apg-1 and Apg-2 cDNAs from a human testis cDNA library. The human Apg-1 was 100% and 91.8% identical in length and amino acid (aa) sequence, respectively, to mouse Apg-1. Human Apg-2 was one aa shorter than and 95.5% identical in sequence to mouse Apg-2. In ECV304, human endothelial cells Apg-1 but not Apg-2 transcripts were induced in 2 h by a temperature shift from 32 degrees C to 39 degrees C. As found in mice, the response was stronger than that to a 37-42 degrees C shift. The human Apg-1 and Apg-2 genes were mapped to the chromosomal loci 4q28 and 5q23.3-q31.1, respectively, by fluorescence in-situ hybridization. We isolated cDNA and genomic clones encompassing the region critical for the difference between Apg-2 and HS24/p52. Although the primer sets used were derived from the sequences common to both cDNAs, all cDNA and genomic clones corresponded to Apg-2. Using a similar approach, the relationship between Apg-2 and Hsp70RY was assessed, and no clone corresponding to Hsp70RY was obtained. These results demonstrated that the Hsp110 family consists of at least three members, Apg-1, Apg-2 and Hsp110 in humans as well as in mice. The significance of HS24/p52 and Hsp70RY cDNAs previously reported remains to be determined.

Improvement in auditory brainstem response of hyperbilirubinemic infants after exchange transfusions

Auditory brainstem response tests were performed before and after exchange transfusions in 6 infants with hyperbilirubinemia. The latencies of Waves I, II, and V decreased significantly after the exchange transfusions (p less than .05, p less than .02, p less than .005, respectively) and I-V interpeak latencies also were decreased (p less than .01). However, the latencies of Wave I both before and after exchange transfusions were within normal limits. The central conduction times were prolonged by hyperbilirubinemia, but the peripheral auditory pathways were not impaired. These auditory brainstem response abnormalities became normal with decreased serum bilirubin concentration. Neonatal hyperbilirubinemia is associated with transient brainstem lesions which are reversible in the early stages. Auditory brainstem response testing is an effective and readily available technique for detecting bilirubin neurotoxicity.

Microarray analysis of 50 patients reveals the critical chromosomal regions responsible for 1p36 deletion syndrome-related complications

OBJECTIVE Monosomy 1p36 syndrome is the most commonly observed subtelomeric deletion syndrome. Patients with this syndrome typically have common clinical features, such as intellectual disability, epilepsy, and characteristic craniofacial features. METHOD In cooperation with academic societies, we analyzed the genomic copy number aberrations using chromosomal microarray testing. Finally, the genotype-phenotype correlation among them was examined. RESULTS We obtained clinical information of 86 patients who had been diagnosed with chromosomal deletions in the 1p36 region. Among them, blood samples were obtained from 50 patients (15 males and 35 females). The precise deletion regions were successfully genotyped. There were variable deletion patterns: pure terminal deletions in 38 patients (76%), including three cases of mosaicism; unbalanced translocations in seven (14%); and interstitial deletions in five (10%). Craniofacial/skeletal features, neurodevelopmental impairments, and cardiac anomalies were commonly observed in patients, with correlation to deletion sizes. CONCLUSION The genotype-phenotype correlation analysis narrowed the region responsible for distinctive craniofacial features and intellectual disability into 1.8-2.1 and 1.8-2.2 Mb region, respectively. Patients with deletions larger than 6.2 Mb showed no ambulation, indicating that severe neurodevelopmental prognosis may be modified by haploinsufficiencies of KCNAB2 and CHD5, located at 6.2 Mb away from the telomere. Although the genotype-phenotype correlation for the cardiac abnormalities is unclear, PRDM16, PRKCZ, and RERE may be related to this complication. Our study also revealed that female patients who acquired ambulatory ability were likely to be at risk for obesity.

Retroviral Vector-Mediated Gene Expression in Human CD34+CD38- Cells Expanded in Vitro: Cis Elements of FMEV Are Superior to Those of Mo-MuLV

A novel murine stromal cell line, HESS-M28, was established, which supports the expansion of human CD34+CD38- cells more than 300-fold in vitro in the presence of human IL-3 and SCF. These cells were used in an attempt to evaluate cis-acting elements of retroviral vectors in human primitive hematopoietic cells. Cord blood cells were cultured on top of the mixed cell layers of the stromal cell line, HESS-M28, and retroviral vector-producing cells. The FMEV-type vector SF/Lyt contained the spleen focus-forming virus U3 and the MESV primer-binding site (PBS), while MO3/Lyt contained the U3 region and PBS from Mo-MuLV. After transduction by the FMEV-type and Mo-MuLV-based vectors, expression of the marker gene murine CD8 (mCD8) was examined in CD34-, CD34+, and CD34+CD38- cells. In CD34+ and CD34+CD38- cells, expression of mCD8 was higher with the FMEV-type vector, SF/Lyt, compared with the cells transduced by the Mo-MuLV-based vector MO3/Lyt, although the expression was comparable in CD34- cells. Expression of marker genes was also confirmed in long-term culture-initiating cells (LTC-ICs) and SCID-repopulating cells (SRCs).

Efficient Gene Transfer by Hybrid Retroviral Vectors to Murine Spermatogenic Cells

Using murine spermatogenic cell lines GC-1 spg and GC-2 spd(ts) as target cells, an attempt was made to design a retroviral vector that would transduce genes efficiently. Promoter activities of various retroviral long terminal repeats (LTRs) were examined by using chloramphenicol acetyltransferase (CAT) as a reporter. The U3 region of spleen focus-forming virus (SFFVp) showed higher enhancer activity than that of Moloney murine leukemia virus (Mo-MuLV) in both cell lines. The U3 region of myeloproliferative sarcoma virus (MPSV) showed higher activity only in GC-1 spg cells. Expression was suppressed by the repressor element of the primer-binding site (PBS) of the Moloney-related virus. The efficiency of transduction of the multidrug-resistance gene (mdr-1) by an Mo-MuLV-based vector was compared with hybrid vectors consisting of the murine embryonic stem cell virus (MESV) PBS and the LTR of either SFFVp or MPSV. Rhodamine efflux assays and colchicine-resistant colony-forming assays demonstrated higher gene expression by the hybrid vectors. Amphotropic and ecotropic receptors were found to be expressed and functional in both cell lines. Thus, these hybrid vectors represent a powerful tool by which to transfer genes into spermatogenic cells.