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Two boys from separate families presented with hereditary multiple exostoses (EXT) and autism associated with mental retardation. Their fathers both expressed a clinical phenotype of hereditary multiple exostoses milder than those of the patients and without the associated mental disorder. The EXT1 and EXT2 genes from lymphocytes of the affected individuals were analyzed by using denaturing high-performance liquid chromatography and direct sequencing. A novel deletion mutation, 1742delTGT-G in exon 9 of EXT1, causing a frameshift was detected in one boy and his father. Another novel deletion mutation, 2093delTT in exon 11 of EXT1, causing transcription termination was detected in the other affected boy and his father. EXT1 is expressed in the brain, and both EXT1 and EXT2 proteins are associated with glycosyltransferase activities required for the biosynthesis of heparan sulfate, which also has activity in the brain. The coincidental association of mental disorders in the boys was not completely excluded. However, these results suggest the involvement of EXT1 in the development of mental disorders, including mental retardation and autism. [ABSTRACT FROM AUTHOR]

A 10 year old girl with benign childhood epilepsy with centrotemporal spikes showed centrotemporal spikes induced by blinking even in a dark room. Spikes could not be induced by photic stimulation, eye closure, eye movement, eye deviation, or passive blinks. There have been no previous reports of spikes induced by blinks in benign childhood epilepsy with centrotemporal spikes. [ABSTRACT FROM AUTHOR]

Presents a letter to the editor regarding the splice site mutation in the hepatocyte nuclear fator-1β gene associated with maturiy-onset diabetes of the young, renal dysplasia and bicornuate uterus, previously published in "Diabetologia."

Autism spectrum disorder (ASD) is a neurodevelopmental disorder with an unknown molecular pathogenesis. A recent molecular focus has been the mutated neuroligin 3, neuroligin 3(R451C), in gain-of-function studies and for its role in induced impairment of synaptic function, but endoplasmic reticulum (ER) stress induced by mutated molecules also deserves investigation. We previously found two missense mutations, H246N and Y251S, in the gene-encoding synaptic cell adhesion molecule-1 (CADM1) in ASD patients, including cleavage of the mutated CADM1 and its intracellular accumulation. In this study, we found that the mutated CADM1 showed slightly reduced homophilic interactions in vitro but that most of its interactions persist. The mutated CADM1 also showed morphological abnormalities, including shorter dendrites, and impaired synaptogenesis in neurons. Wild-type CADM1 was partly localized to the ER of C2C5 cells, whereas mutated CADM1 mainly accumulated in the ER despite different sensitivities toward 4-phenyl butyric acid with chemical chaperone activity and rapamycin with promotion activity for degradation of the aggregated protein. Modeling analysis suggested a direct relationship between the mutations and the conformation alteration. Both mutated CADM1 and neuroligin 3(R451C) induced upregulation of C/EBP-homologous protein (CHOP), an ER stress marker, suggesting that in addition to the trafficking impairment, this CHOP upregulation may also be involved in ASD pathogenesis.

Objective: The main pathogenic characteristic of Kawasaki disease (KD) is the activation of mononuclear phagocytes. The cytokines produced by activated monocytes/macrophages elicit proinflammatory and prothrombotic responses in endothelial cells. Thus, we speculated that macrophage colony-stimulating factor (M-CSF), derived from monocytes/macrophages or vascular endothelial cells, might play an important role in the pathogenesis of the acute phase of KD. The aim of this study was to investigate the possible role of M-CSF in the pathogenesis of KD and to elucidate the relationship between serum M-CSF levels and clinical features and cardiac lesions., Methods: Using ELISA, we serially assayed M-CSF and several cytokines, including interleukin-6, interleukin-8, tumor necrosis factor-alpha and interferon-gamma in the sera of 32 KD patients aged 2 months to 4 years., Results: The serum M-CSF level during the first week of illness was significantly higher than during the second week or thereafter (first week, median 1710.0; second week, 1121.0; third week, 867.3; fourth week, 909.4 U/ml, p<0.001) in our KD patients. Serum M-CSF levels during the first week of illness were also higher in patients with mitral and/or aortic valvular insufficiency than in patients without cardiac complications. Furthermore, serum M-CSF levels in patients with persistent coronary dilatation were higher than in those with no cardiac complications., Conclusion: M-CSF plays a critical role in the pathogenesis of KD and can be used as an indicator for the risks of valvulitis and coronary arteritis.

We present here a case of episodic, pure cheiro-oral syndrome caused by a ruptured intracranial dermoid cyst. Cranial magnetic resonance imaging (MRI) using the fat-suppression method revealed a fatty mass lesion in the subarachnoid space of the left parasellar region and multiple lipid droplets in the subarachnoid space over the left perisylvian area. Although no evidence for it pathogenesis was obtained, the patient's cheiro-oral syndrome could have resulted from a transient vasospasm around the left ventral posterior thalamic nucleus or postcentral gyrus.

Background: Gastroschisis is a rare abdominal wall defect. Although the pathogenesis of gastroschisis is unknown, there is some evidence of the genetic etiology of gastroschisis. Recently, a functionally null deletion of the mouse bone morphogenic protein-1 (BMP-1) gene resulted in a phenotype that resembled a human neonate with gastroschisis. BMP-1 thus became the first potential candidate gene for gastroschisis., Methods: To explore this possibility the authors collected blood samples from 11 patients who had gastroschisis. Mutational analysis of exons 2 to 15 of the human BMP-1 gene was performed using genomic polymerase chain reaction, single-strand conformation polymorphism analysis and direct sequencing methods., Results: No mutation of the human BMP-1 gene was observed in any of these patients., Conclusion: Although heterogeneous etiologies might be proposed for gastroschisis, our results provide further evidence of a nongenetic etiology for gastroschisis. J Pediatr Surg 36:885-887., (Copyright 2001 by W.B. Saunders Company.)

Truncated polypeptides containing expanded polyglutamine (polyQ) stretches tend to form cytoplasmic or nuclear aggregates in cultured cells, leading to cell death. Although it has been shown recently that caspase-8 coaggregates with polyQ and is activated during polyQ-mediated cell death, little is known of the location and timing of caspase-8 activation by nuclear polyQ aggregates. Also, the relationship between nuclear polyQ aggregate-mediated cell death and activation of other caspases is unclear. In P19 embryonal carcinoma (EC) cells, which can be made to differentiate into neuronal cells, polyQ72 repeats preferentially aggregate in the nucleus. Nuclear aggregates of polyQ72 induced P19 EC cell death, with a high frequency of cells exhibiting morphology characteristic of apoptosis (i.e., roundness, cell shrinkage, chromatin condensation) and DNA fragmentation. In the present study, we used antisera that specifically recognized the active forms of caspase-8, -3, and -9 but not their proforms, and showed that only caspase-8 and -3 were activated during the generation of polyQ72 aggregates in P19 EC cell nuclei. Furthermore, we showed that the caspase inhibitor z-VAD-fmk inhibited DNA fragmentation, but only partially inhibited the appearance of apoptotic morphology. Thus, caspase activation, including caspase-8 and -3, is necessary for polyQ-mediated DNA fragmentation but not sufficient for polyQ-mediated cell death in P19 EC cells., (Copyright 2000 Wiley-Liss, Inc.)

We report here a case involving a 14-year-old boy who developed acute disseminated encephalomyelitis following live rubella vaccination. The patient became febrile and began to experience nuchal pain 16 days after the immunization. By 22 days after immunization, he experienced difficulty in walking. By 24 days, he had developed tetraparesis with retention of urine, and total sensory loss below the Th1 dermatomal level. He was febrile at this point and showed nuchal rigidity and Lhermitte's sign. Cerebrospinal fluid examination revealed elevated cell counts, protein level, and myelin basic protein. T2-weighted magnetic resonance imaging detected high intensity lesions in the bilateral cerebral white matter and cervical spinal cord. Following the administration of intravenous corticosteroids, the patient's clinical symptoms improved rapidly.

Background: Pulmonary hypertension (PH) causes mortality in some congenital and acquired heart and lung diseases. However, inhalation of NO gas requires complicated and expensive instruments and elaborate preparations to avoid toxic gas administration. We tested the effectiveness and safety of inhaled nebulized nitroglycerin (Neb-NTG) in dogs with experimental PH., Methods: Experimental PH was induced by continuous infusion of a thromboxane analog (U46619). The U46619 infusion rate was adjusted to maintain a systolic pulmonary artery pressure (PAP) at 40 mmHg in 10 anesthetized and mechanically ventilated dogs. Then, 20 micrograms/kg of NTG liquid nebulized by compressed air was inhaled., Results: After infusion of U46619, the systolic, diastolic and mean PAP increased by 119%, 228% and 169%, respectively, and the systolic, diastolic and mean systemic arterial pressures (SAP) increased by 19%, 29% and 23%, respectively. The systolic pulmonary to systemic pressure ratio (Pp/Ps) and mean Pp/Ps increased by 83% and 113%, respectively, and the pulmonary vascular resistance (PVR), systemic vascular resistance (SVR) and pulmonary to systemic resistance ratio (Rp/Rs) increased by 341%, 100% and 145%, respectively. After inhalation of Neb-NTG in dogs with experimental PH, systolic, diastolic and mean PAP and PVR decreased by 25 +/- 4, 26 +/- 11, 25 +/- 9 and 31 +/- 21%, respectively. There were no significant changes in systolic, diastolic and mean SAP, SVR, cardiac output and plasma methemoglobin concentrations. The systolic and mean Pp/Ps decreased by 18 +/- 7 and 20 +/- 7%, respectively. The Rp/Rs decreased by 25 +/- 13%., Conclusions: The results of this study demonstrate that Neb-NTG is an effective and selective pulmonary vasodilator and may offer a new therapeutic option for PH.

The relation between activation of caspase-8 and polyglutamine aggregates has been focused. We prepared an antiserum (anti-m8D387) that recognizes the active form but not the proform of mouse caspase-8. We used immunostaining with anti-m8D387 antiserum to compare the localizations of activated mcaspase-8 in L929 (clone 1422) cells induced by TNF and polyglutamine aggregates. Anti-m8D387 was positive throughout cytoplasm of the TUNEL-positive cells induced by TNF treatment, whereas the anti-m8D387 reactivity was not positive throughout cytoplasm of the cells expressing polyglutamine but was restricted to polyglutamine aggregates. In contrast with TNF-treated cells, cells expressing anti-m8D387-positive cytoplasmic polyglutamine aggregates did not undergo TUNEL-positive apoptosis. Thus activated caspase-8 associated with polyglutamine aggregates alone was not sufficient to induce TUNEL-positive apoptosis of L929 (clone 1422) cells. The distribution of activated caspase-8 associated with polyglutamine aggregates may be essential for the polyglutamine-mediated cell death or downstream of caspase-8 may be different in the TNF-treated cells and cells expressing polyglutamine., (Copyright 2000 Academic Press.)

Two children with Landau-Kleffner syndrome were successfully treated with antiepileptic drugs and a high-dose intravenous corticosteroid. A combination of valproate and a benzodiazepine (clonazepam or diazepam) ameliorated epileptic seizures and electroencephalographic spikes and waves, but speech disturbances persisted. Both patients were treated with an intravenous infusion of high-dose methylprednisolone sodium succinate (20 mg/kg daily) for 3 consecutive days. This infusion was repeated three times with a 4-day interval between treatments, which resulted in a rapid improvement in speech ability. After intravenous therapy, prednisolone was given orally (2 mg/kg daily for 1 month, then gradually withdrawn), which maintained the clinical improvement in speech.

A 4-year-old boy showed two episodes of encephalitis/encephalopathy involving disturbed consciousness, convulsion, and paresis associated with the elevated levels of protein and myelin basic protein of the cerebrospinal fluid. MRI studies of the brain revealed symmetrical lesions in the brain stem and thalami at the first episode, and additional lesions were found in the cerebellum involving both the gray and white matter in the second episode. The intensities of MRI lesions were low in T I and high in T2. These episodes were followed by an elevation of the anti-viral antibody titers, for influenza A virus during the first episode and for adenovirus during the second. In the second episode, intravenous methylprednisolone therapy resulted in rapid improvement of his neurological signs.

dy/dy mice, which carry an unidentified mutation in the Lama2 gene, show dystrophic pathologies similar to those of human congenital muscular dystrophy. Laminin alpha2 deficiency induces apoptosis with DNA fragmentation. Caspases, which are involved in various types of cell death, are sequentially activated through a processing by other members of caspases. By using a cleavage site-directed antibody against caspase-3 that specifically reacts with the active form of caspase-3, we immunochemically demonstrated that caspase-3 is activated in the skeletal muscle fiber of dy/dy mice and that some of the activated caspase-3 muscle fibers are TUNEL-positive. Thus the lack of laminin alpha2 signals activates caspase-3, resulting in the apoptosis of muscle fibers., (Copyright 1999 Academic Press.)

To examine any role of macrophage colony-stimulating factor (M-CSF), in the immune responses in Kawasaki disease (KD), we serially assayed M-CSF and several related cytokines using ELISA. In 10 paediatric patients with KD the level of M-CSF was significantly higher in the acute phase than in the convalescent phase (1476.1 +/- 443.6 v 805.0 +/- 184.7 U/ml). Higher levels of serum granulocyte colony-stimulating factor (G-CSF) and interleukin-6 were also found in the acute phase. These results suggest that M-CSF, G-CSF and interleukin-6, derived from monocytes as monokines or derived from vascular endothelial cells, might play an important role in the acute phase of KD.

P19 embryonal carcinoma (EC) cells undergo apoptosis during neuronal differentiation induced by all-trans retinoic acid (RA). Caspase-3-like proteases are activated and involved in the apoptosis of P19 EC cells during neuronal differentiation.1 Recently it has been shown that growth factor signals protect against apoptosis by phosphorylation of Bad. Phosphorylated Bad, an apoptotic member of the Bcl-2 family, cannot bind to Bcl-xL and results in Bcl-xL homodimer formation and subsequent antiapoptotic activity. In the present study, we demonstrate that this system is used generally to protect against apoptosis during neuronal differentiation. Bcl-xL inhibited the activation of caspase-3-like proteases. Basic fibroblast growth factor (bFGF) inhibited more than 90% of the caspase-3-like activity, inhibited processing of caspase-3 into its active form, and inhibited DNA fragmentation. bFGF activated phosphatidyl-inositol-3-kinase (PI3K) and stimulated the phosphorylation of Bad. Phosphorylation was inhibited by wortmannin, an inhibitor of PI3K and its downstream target Akt. Thus, Bad is a target of the FGF receptor-mediated signals involved in the protection against activation of caspase-3.

To determine the threshold at which rapid atrial pacing brings on fetal circulatory failure, we made a fetal supraventricular tachyarrhythmia model and measured the central venous pressure, aortic pressure, and right and left ventricular outputs in five fetal lambs. Under maternal anesthesia, the uterus was opened, and under local anesthesia, polyvinyl catheters were inserted into the fetal superior vena cava and ascending aorta through a neck incision. Pacing leads (Medtronic model 6492) were then sutured onto the fetal right atrial appendage via right thoracotomy. Ventricular output was estimated using a Toshiba SSH-65A echocardiography by a transuterine approach. Fetal hemodynamics were observed without pacing (control), and at the atrial pacing rates of 200, 300, 350, and 400/min. Central venous pressure (CVP) increased and the aortic pressure decreased when the right atrium was paced at 350/min or more. Right ventricular output decreased when the right atrium was paced at 300/min or more. The left ventricular output, however, remained constant. The right ventricular output was 382 +/- 106 mL/kg/min at control, and 391 +/- 117 mL/kg/min when paced at 200/min, but decreased to 210 +/- 138 mL/kg/min when paced at 300/min, to 223 +/- 102 mL/kg/min when paced at 350/min, and to 186 +/- 86 mL/kg/min when paced at 400/min. Fetal circulatory failure occurred when the right atrium was paced at 300/min or more.

We previously demonstrated that Caspase-3 is highly expressed in dorsal root ganglia and trigeminal ganglia of mouse embryos [T. Mukasa, K. Urase, Y.M. Momoi, I. Kimura, T. Momoi, Specific expression of CPP32 in sensory neurons of mouse embryos and activation of CPP32 in the apoptosis induced by a withdrawal of NGF, Biochem. Biophys. Res. Commun., 231 (1997) 770-774.]. Since, however, Caspases are processed into active form during apoptosis, it is difficult to examine the involvement of activated Caspases in naturally occurring cell death during development by immunohistochemical staining or in situ hybridization method. We prepared a cleavage site-directed antiserum against Caspase-3 (anti-p20/17). This antiserum reacted with fragment (p20/17) of Caspase-3, but not proCaspase-3 (p32), proCaspase-7 (p34) and its cleaved fragment (p24). We examined the relationship between the activation of Caspase-3 and the appearance of the naturally occurring apoptotic cells in the nervous system during development. In the trigeminal ganglia and dorsal root ganglia, the expression of Caspase-3 mRNA was maximal before the appearance of p20/17-positive cells and apoptotic cells. In the mouse brain, many p20/17-positive cells and apoptotic cells were observed in the neuroepithelium in the early developmental stages, but very few p20/17-positive cells were detected in postmitotic neurons in the cerebral cortex although Caspase-3 mRNA was expressed highly. Caspase-3 is activated mainly during apoptosis of neuroepithelial cells in the early developmental stages but not of mature neurons at postnatal stages., (Copyright 1998 Elsevier Science B.V.)

Serum concentrations of valproic acid (VPA) were reduced to 0% to 40% of the original levels by concomitant use with panipenem-betamipron (PAM-BP) in three patients. The serum VPA level began to decrease 2 days after the administration of PAPM-BP, and it began to increase within 24 hours of the last dose. The rapid change in the serum VPA level suggests the existence of unique and as yet unknown mechanisms of interaction between VPA and PAPM-BP. Epileptic seizures developed in two of the three patients during PAPM-BP use, which signaled the dangers of PAPM-BP administration to patients concomitantly administered VPA. PAPM-BP should not be used in patients administered VPA.

A 16-year-old girl had fully manifested Coffin-Lowry syndrome and drop episodes. Her drop episodes were precipitated by sudden unexpected tactile or auditory stimuli associated with the electrostatic circumstances in her leg muscles immediately after the stimuli. Studies revealed that her drop episode symptom was an unusual type of startle response and that it may be associated with Coffin-Lowry syndrome.

We report on a boy with proximal interstitial deletion of chromosome 4, del(4)(q21.22q23). The patient was born at term with a low birth weight, flat nasal bridge, micrognathia, wide-spaced nipples, clinodactyly of fifth fingers, overlapping fingers, post-axial polydactyly of the right foot, micropenis, hypospadias, a dermal sinus, and cardiac malformations. He developed psychomotor retardation, seizures, and a liver tumor with an increased serum alpha-fetoprotein level and rapid growth. The patient carried a deletion of chromosome 4 involving the 4q21-q22 region that was reported to form a unique syndrome. The absence of central nervous system overgrowth and the presence of a malignant liver tumor are unique to our patient, compared to others with the 4q21-q22 deletion syndrome. The clinical manifestations and relationship between the liver tumor and chromosomal anomaly are discussed.

We prepared a cleavage site-directed antiserum against Caspase-3 (anti-p20/17), which reacts with the p20/17 fragment (p20/17) activated by cleavage but not proCaspase-3 (p32), and examined the relationship between the activation of Caspase-3 and apoptosis. We identified p20/17-positive cells where cell death occurs naturally: interdigits of the forelimbs, small intestine epithelium, thymus, trigeminal ganglia, and dorsal root ganglia of mouse embryos. Withdrawal of nerve growth factor induced the appearance of p20/17-positive cells with DNA fragmentation in the culture of dorsal root ganglia neurons, while DNA fragmentation was detected in both p20/17-positive and -negative neurons in dorsal root ganglia of mouse embryos. These results suggest that not only activation of Caspase-3 but also other molecular mechanism play a role in the naturally occurring dorsal root ganglia apoptosis. Cleavage site-directed antisera against Caspases will be useful for the analysis of the molecular mechanism of naturally occurring apoptosis during development.

Involvement of the cardiac conduction system is a common clinical feature in myotonic dystrophy, whereas the association of primary myocardial abnormalities has rarely been reported. A patient with a severe form of congenital myotonic dystrophy who developed fatal left ventricular hypertrophy at 3 months of age and died at 2 years of age is reported. Serial ultrasonographic studies revealed progressive left ventricular hypertrophy accompanied by outflow obstruction of the left ventricle. Southern analysis for the myotonin kinase gene revealed a 5.8 kb expansion of CTG repeats in addition to a fragment of normal length. The degree of expansion was much greater than those of other reported patients with congenital myotonic dystrophy. These findings suggest that left ventricular hypertrophy represents an extreme level of myocardial damage in myotonic dystrophy and that this damage may be related to the larger size of the CTG repeats.

A 12-year-old girl developed ataxia that gradually progressed. At age 18 the patient presented with mental retardation, cachectic dwarfism, microcephalus, and a progeroid appearance but no photosensitive skin lesions or deafness. On analysis of fibroblasts, unscheduled DNA synthesis was reduced to 50% of normal, but colony-forming ability after ultraviolet irradiation was normal. The symptoms and phenotype of the patient were distinguished from those in Cockayne syndrome and xeroderma pigmentosum. This case is interesting because the defect in DNA repair after ultraviolet irradiation was detected in a patient with neurologic disturbances but without photosensitive skin lesions.

We present five different types of dentatorubral-pallidoluysian atrophy in one Japanese family. Two siblings and their paternal uncle manifested the juvenile type dentatorubral-pallidoluysian atrophy, the siblings' father had the late-adult type, and another paternal uncle had the early-adult type. Gene analysis confirmed the diagnosis for the proband and her sibling. By following the clinical courses and electroencephalographic changes, we found that the types of epileptic seizures and the electroencephalograms of the juvenile dentatorubral-pallidoluysian atrophy patients changed as the illness progressed. The siblings exhibited different levels of clinical severity despite the similar deoxyribonucleic acid expansion detected in their lymphocytes.

We present the clinical and laboratory effects of dichloroacetate (DCA) in three children with mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes (MELAS) who had not responded to other medications. Administration of DCA lowered the elevated levels of lactate and pyruvate in the serum and CSF. DCA ameliorated abdominal pain, headache, and strokelike episodes, and improved cognitive function and fatigability in the three patients during the study period. Some transient liver dysfunction, hypocalcemia, and peripheral neuropathy were observed. The use of DCA in MELAS merits further study.

The enzyme oligosaccharyltransferase (dolichyl-diphosphooligosaccharide-protein glycosyltransferase; EC 2. 4.1.119) (DDOST) catalyzes the transfer of a high-mannose oligosaccharide (GlcNac2Man9Glc3) from a dolichol-linked oligosaccharide donor (dolichol-P-GlcNac2Man9Glc3) onto the asparagine acceptor site within an Asn-X-Ser/Thr consensus motif in nascent polypeptide chains across the membrane of the endoplasmic reticulum. We isolated mouse and human DDOST cDNAs from retinoic acid-treated mouse P19 EC cells and human NT-2 cells, respectively. DDOST mRNA is expressed intensely in heart and pancreas, but at lower levels in brain. Here we show that the human DDOST 48-kDa subunit gene (HGMW-approved symbol DDOST) is organized into 11 exons expanding about 9 kb. This DDOST subunit gene is localized on chromosome 1p36.1 by fluorescence in situ hybridization analysis., (Copyright 1997 Academic Press.)

Mutation of the Patched gene has been detected in human inherited basal cell nevus syndrome (BCNS) and sporadic basal cell carcinomas (BCC), suggesting a strong relation between a Sonic hedgehog-Patched signal and cell proliferation. In the present study, we demonstrate that Sonic hedgehog is expressed in human lung squamous carcinoma (LK-2 and EBC-1) and some adenocarcinoma cell lines. The expression of Sonic hedgehog is also detected in the human lung squamous carcinoma tissues, but not in the normal lung tissue of the same patient. The N-terminal region of Sonic hedgehog stimulates the incorporation of BrdU into LK-2 cells and stimulates their cell growth, while anti-Shh-N inhibits their cell growth. These results suggest that a Sonic hedgehog signal is involved in the cell growth of LK-2 cells., (Copyright 1997 Academic Press.)

P19 EC cells undergoes apoptosis during neuronal differentiation induced by retinoic acid. Two CPP32-like proteases, CPP32 and Mch-3, are expressed in untreated and retinoic acid-treated P19 EC cells. CPP32-like activity is remarkably increased in apoptosis during neuronal differentiation of P19 EC cells. Inhibition of CPP32-like proteases prevents apoptosis, suggesting that activation of CPP32-like proteases play central roles in the apoptosis during neuronal differentiation of P19 EC cells. Wortmannin, PI-3K inhibitor, enhances the CPP32-like activity of the retinoic acid-treated P19 EC cells. PI-3K may be involved in the apoptosis during neuronal differentiation as negative regulator.

We isolated mouse CPP32/apopain cDNA, a mammalian homologue most closely related to Ced-3 in C. elegans, and examined the involvement of CPP32 in the apoptosis of nervous system during development. CPP32 is specifically expressed in the trigeminal (V) ganglia, facio-acoustic (VII-VIII) ganglion complex, and dorsal root ganglia (DRGs) of mouse 10.5-day embryos. CPP32-like proteases are activated during apoptosis of DRG neurons induced by deprivation of NGF and serum. Ac-DEVD-CHO, an inhibitor for CPP32-like proteases, prevents apoptosis of DRG neurons, but Ac-YVAD-CHO, an inhibitor for ICE-like proteases, does not. These results suggest that CPP32 or CPP32-like proteases play a role as central mediator in the apoptosis of DRG neurons induced by lack of neurotrophin signals.

Sonic hedgehog (Shh), a homologue of Drosophila hedgehog, was specifically expressed in lung epithelium during branching morphogenesis, but was not uniformly expressed in lung epithelium. Shh was intensely expressed in the distal tips of the bronchial tubes during branching morphogenesis, and Shh was localized on the apical side of the epithelium. On the other hand, Bmp-4, one of the target genes of Shh, was also specifically expressed in the epithelium at the branching point. These results suggest that Shh and Bmp-4 are involved in the branching morphogenesis of lung epithelium.

Clinical and pathological observations of a 6-month-old-boy with Costello syndrome are reported. The main clinical findings were loose skin of the neck, hands, and feet, deep palmar and plantar creases, typical "coarse" face with thick lips and macroglossia, relative macrocephaly, mental retardation, short stature, arrhythmia, large size for gestational age, and poor feeding. At age 6 months he died of rhabdomyolysis. The major pathological findings were fine, disrupted, and loosely-constructed elastic fibers in the skin, tongue, pharynx, larynx, and upper esophagus, but not in the bronchi, alveoli, aorta, or coronary arteries. Hyperplasia of collagen fibers in the skin, hyperplasia of the mucous glands in the bronchus, narrowing of the pulmonary artery, degeneration of the atrial conduction system, calcification and ballooning of skeletal muscle fibers with infiltration of macrophages, and myoglobin depositions in the collecting ducts in the kidney were also observed. The degeneration of elastic fibers was confirmed in the skin of a second Costello syndrome patient. Expression of elastin mRNA in the patient's fibroblasts was normal in size and amount. Given that elastic fiber degeneration was observed in the tissues with clinical symptoms, we speculate that a defect of elastic fibers, possibly relating to alternative splicing in the elastin gene or to defects in elastin microfibrils, might be involved in the pathogenesis of Costello syndrome.

The expression and distribution of three retinoic acid receptors, alpha, beta, and gamma, were investigated in the CNS of mouse embryos during development. mRNAs and protein of RAR-beta that were expressed in the spinal cord of the 12.5-day mouse embryo decreased during development but they were not decreased in the brain. The RAR-beta-positive cells were already present in the ventral region of the spinal cord of 10.5-day mouse embryos, gradually appeared in the dorsal region during development and then disappeared from the spinal cord after birth. In the brain, RAR-beta-positive cells were detected in the mesencephalon and rhombencephalon but not in the telencephalon of the 12.5-day mouse embryos. RAR-beta-positive cells were present in the hippocampus and cingulum but not in the neocortex of 14.5-day mouse embryos. Most neurons in the hippocampus of 16.5-day mouse embryos and the cortex of newborn mice were RAR-beta-positive. In the spinal cord, RAR-alpha mRNAs and proteins also decreased during development but more gradually than RAR-beta mRNAs and proteins. During development, the distributions of RAR-alpha and -beta in the spinal cord and brain did not differ substantially. The main difference was the appearance of a subtypes of RAR-alpha, a 52-kDa protein, in the brain of newborn mice. On the other hand, RAR-gamma proteins were only faintly detected in the spinal cord and the brain of the mice during the embryonal stages but these increased after birth. The distribution of RAR-alpha- or -beta-positive cells were consistent with the neurogenesis during development in the spinal cord and brain.

We report a patient with Miller Fisher syndrome who was treated with an intravenous high-dose of immunoglobulin. This syndrome is considered to be a benign variety of acute inflammatory demyelinating polyneuropathy (Guillain-Barré syndrome). However, there have been several reports of the need for ventilatory support and a few cases have had a fatal outcome. We observed a case of progressive Miller Fisher syndrome in a 3-year-old boy. Following 2 episodes of apnea lasting about 50 s each, he was treated with intravenous immunoglobulin (400 mg/kg/day) for 5 consecutive days. His respiratory state, general muscle strength, truncal ataxia and emotional state improved remarkably after this therapy.

MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes) is a major subgroup of heterogeneous mitochondrial diseases. For identifying a mutation in the mitochondrial gene, we isolated, from the same muscle tissue from a patient with MELAS, cell lines with distinctly different phenotypes: one was respiration-deficient, and the other was apparently normal. Compared with the normal cells, only one A-to-G nucleotide transition at nucleotide 3243 in the tRNA-Leu (UUR) gene was found in whole mtDNA of the respiration-deficient cells. This mutation was also found in eight patients, from unrelated families, who had MELAS in a heteroplasmic manner but was not found in control individuals. Therefore, the single point mutation causes the functional abnormality in the respiratory chain of mitochondria.

Mitochondrial myopathy, encephalopathy, lactic acidosis and strokelike episode (MELAS) is a major group of heterogeneous mitochondrial disorders. To identify the defective gene, mitochondrial DNA from a patient with MELAS was sequenced by using amplified DNA fragments as sequencing templates. In 14.1 kbp determined out of 16.6 kbp of the whole mitochondrial gene, at least 21 nucleotides were different from those of a control human mitochondrial DNA. One of the substitutions was a transition of A to G in the tRNA(Leu) (UUR) gene at Cambridge nucleotide number 3,243. This nucleotide is conserved not only in many mitochondrial tRNAs but in most cytosolic tRNA molecules. An Apa I restriction site was gained by the substitution of this nucleotide. The Apa I digestion of the amplified DNA fragment revealed that all independent 6 patients had G at nucleotide number 3,243 in their mitochondrial DNAs, but none of 11 control individuals had G at this position. This result strongly suggests that the mutation in the mitochondrial tRNALeu gene causes MELAS.