Exonic deletion of CASP10 in a patient presenting with systemic juvenile idiopathic arthritis, but not with autoimmune lymphoproliferative syndrome type IIa.

Systemic juvenile idiopathic arthritis (s-JIA) is a rare inflammatory disease classified as a subtype of chronic childhood arthritis, manifested by spiking fever, erythematous skin rash, pericarditis and hepatosplenomegaly. The genetic background underlying s-JIA remains poorly defined. To detect copy number variations, we performed single nucleotide polymorphism (SNP) array analysis in 50 patients with s-JIA. We found a 13-kb intragenic deletion of CASP10 in one patient. RT-PCR of the mRNA extracted from the patient's lymphoblastoid cells revealed that CASP10 mRNA was truncated. Sequencing the mRNA revealed that this deletion resulted in a frame shift with an early stop codon. CASP10 is known as a causative gene for autoimmune lymphoproliferative syndrome (ALPS) type IIa, another childhood syndrome of lymphadenopathy and splenomegaly associated with autoimmune haemolytic anaemia and thrombocytopenia. TCR αß(+) CD4/CD8 double-negative T cells in the peripheral blood as a diagnostic marker of ALPS were not high in this patient and lymphocyte apoptosis induced by anti-Fas antibody was normal, denying ALPS in the patient. The father and a sister of the patient showing no symptoms of ALPS or s-JIA, also had the same deletion. Furthermore, we found no other mutations of CASP10 in the other 49 s-JIA patients. These data suggest that the pathogenic significance of CASP10 mutations should be carefully evaluated in s-JIA or even ALPS type IIa in further studies.

Glomerulonephritis in children with mixed connective tissue disease.

AIMS: Mixed connective tissue disease (MCTD) has overlapping clinical features with systemic lupus erythematosus (SLE). Renal biopsy is necessary for all children with SLE to evaluate the prognosis, because they are at a quite high risk of developing renal complications. Furthermore, lupus nephritis and hypocomplementemia usually precede the appearance of clinical manifestations. Immune complex-mediated nephritis is one of the major complications of MCTD. Juvenile MCTD is known to be associated with a higher risk of nephritis than adult MCTD. However, it is uncertain whether all children with MCTD should be subjected to a renal biopsy, and whether most of those with hypocomplementemia present nephropathy, as in patients with SLE. We examined the histopathological characteristics of juvenile MCTD nephritis, the importance of renal biopsy and the implications of hypocomplementemia in our patients and reported cases of MCTD. MATERIAL AND METHODS: We performed renal biopsy in 11 children with MCTD and found 6 patients with glomerulonephritis. In addition, we studied the frequency and the characteristics of glomerulonephritis in 71 cases of juvenile MCTD (our 11 patients and 60 reported cases). We also analyzed the relationship between hypocomplementemia and pathological features in 41 cases of MCTD nephritis (23 adults, 18 children). RESULTS: 6 of our 11 patients had glomerulonephritis, but of them four had no abnormality in urinalysis at the time of biopsy. In 5 patients renal biopsy showed normal findings. Review of 71 cases of juvenile MCTD showed that of them 28% presented latent asymptomatic nephritis at the time of biopsy. Membranous nephropathy (MN) and mesangial proliferative glomerulonephritis (MPG) were common in MCTD. Interestingly, hypocomplementemia was more frequently observed in patients with MN or mixed form of MN and MPG (MPG/MN) than simple MPG based on our review of 41 cases (p < 0.01). CONCLUSION: A more aggressive indication of renal biopsy should be considered in children with MCTD because of the high incidence of non-clinical nephritis. The hypocomplementemia observed in patients with MCTD suggests the high frequency of glomerulonephritis, including membranous lesions.

Split chimerism after allogeneic bone marrow transplantation in Chediak-Higashi syndrome.

Chediak-Higashi Syndrome (CHS) is a hereditary multiorgan disease associated with a lymphoproliferative disorder termed 'accelerated phase' (AP). As AP is often life-threatening, hematopoietic stem cell transplantation has been proposed as the only curative treatment for CHS. Here, we report a 1-year-old Japanese boy with CHS who received an HLA-matched unrelated BMT at the AP stage, which resulted in split chimerism. We evaluated the chimerism status of isolated leukocytes and found that only a limited population of T and NK cells was of donor origin and the majority of these and other hematopoietic cells was of host origin. Clinical outcome was successful, and the patient is currently alive and well, free of AP and serious infections more than 18 months after BMT.

Tonic function of nicotinic receptors in stress-induced release of L-DOPA from the nucleus accumbens in freely moving rats.

We investigated whether stress induces the release of L-3,4-dihydroxyphenylalanine (DOPA) and dopamine from the nucleus accumbens in conscious rats and characterized the stress-induced response. Electrical foot-shock stress induced both DOPA and dopamine release, measured by microdialysis, from the nucleus accumbens in freely moving rats. Pretreatment of rats with mecamylamine completely blocked stress-induced DOPA release, but only partially blocked dopamine release. Diazepam did not affect the foot-shock-induced release of DOPA, while the same dose of diazepam partially blocked the stress-induced release of dopamine. These findings suggest a tonic function of central nicotinic receptors in stress-induced DOPA release from the nucleus accumbens in conscious rats.

[Disseminated bone marrow metastases from gastric cancer: detection and monitoring the effectiveness of chemotherapy by bone marrow scintigraphy].

Disseminated bone marrow metastasis of cancer is a critical condition, frequently complicated by disseminated intravascular coagulation (DIC). A 32-year-old man with gastric cancer was diagnosed as having disseminated bone marrow metastases. Bone scintigraphy demonstrated many abnormal radionuclide accumulations in the whole body. Bone marrow aspiration revealed cancer cells. Bone marrow scintigraphy with 111In-Cl3 demonstrated central marrow failure and peripheral expansion. The remission of DIC was observed after sequential methotrexate and 5-FU therapy, then uptake of radionuclide in the central bone marrow was remarkably improved by bone marrow scan. After thirteen anti-cancer chemotherapies, recurrence of DIC was suspected because of the reduction of blood platelet count. Nevertheless, repeated bone marrow scan still demonstrated the central bone marrow clearly. The patient discharged from our hospital without the recurrence of DIC. We considered bone marrow scintigraphy is useful in the detection of disseminated bone marrow metastases of cancer and monitoring the effectiveness of chemotherapy.

Autoradiographic studies using L-[(14)C]DOPA and L-DOPA reveal regional Na(+)-dependent uptake of the neurotransmitter candidate L-DOPA in the CNS.

We previously proposed that L-3,4-dihydroxyphenylalanine (L-DOPA) is a neurotransmitter in the CNS. Receptor and transporter molecules for L-DOPA, however, have not been determined. In the present study, in order to localize the uptake sites of L-DOPA in the CNS, we performed autoradiographic uptake studies using L-[14C]DOPA and L-[3H]DOPA in the uptake study on rat brain slice preparations, and further analyzed the properties of L-DOPA uptake. Image analysis of the L-[14C]DOPA autoradiogram showed a unique heterogeneous distribution of uptake sites in the brain. The intensity was relatively high in the cerebral cortex, the hypothalamus, the cerebellum and the hippocampus, while the density was moderate or even low in the striatum and the substantia nigra. L-DOPA and phenylalanine, but not dopamine (10mM) were able to almost completely inhibit the uptake of L-[14C]DOPA to basal levels. Microautoradiographic studies using L-[3H]DOPA revealed accumulation of dense grains in the median eminence, the supraoptic nucleus of the hypothalamus, the cerebral cortex (layer I) and the hippocampus. In the cerebellum, grains formed in clusters surrounding the Purkinje cells. This grain accumulation was concluded to be in Bergmann glial cells, since the morphological pattern of grain accumulation was similar to that of the immunoreactivity of the glutamate aspartate transporter, a marker protein for Bergmann glial cells. In the hippocampus, the grain density significantly decreased under Na(+)-free conditions. In addition, grain density also decreased in the absence of Cl(-). In contrast, grains in the choroid plexus and the ependymal cell layer, were not affected by the absence of Na(+). These findings indicated that the uptake of L-DOPA occurs via various types of large neutral amino acid transport mechanisms. It appears that neuronal and/or glial cells, which take up L-DOPA in a Na(+)-dependent manner, exist in the CNS. Our finding further supports the concept that L-DOPA itself may act as a neurotransmitter or neuromodulator.

Endogenously released DOPA is a causal factor for glutamate release and resultant delayed neuronal cell death by transient ischemia in rat striata.

Glutamate is implicated in neuronal cell death. Exogenously applied DOPA by itself releases neuronal glutamate and causes neuronal cell death in in vitro striatal systems. Herein, we attempt to clarify whether endogenous DOPA is released by 10 min transient ischemia due to four-vessel occlusion during rat striatal microdialysis and, further, whether DOPA, when released, functions to cause glutamate release and resultant delayed neuronal cell death. Ischemia increased extracellular DOPA, dopamine, and glutamate, and elicited neuronal cell death 96 h after ischemic insult. Inhibition of striatal L-aromatic amino acid decarboxylase 10 min before ischemia increased markedly basal DOPA, tripled glutamate release with a tendency of decrease in dopamine release by ischemia, and exaggerated neuronal cell death. Intrastriatal perfusion of 10-30 nM DOPA cyclohexyl ester, a competitive DOPA antagonist, 10 min before ischemia, concentration-dependently decreased glutamate release without modification of dopamine release by ischemia. At 100 nM, the antagonist elicited a slight ceiling effect on decreases in glutamate release by ischemia and protected neurons from cell death. Glutamate was released concentration-dependently by intrastriatal perfusion of 0.3-1 mM DOPA and stereoselectively by 0.6 mM DOPA. The antagonist elicited no hypothermia during and after ischemia. Endogenously released DOPA is an upstream causal factor for glutamate release and resultant delayed neuronal cell death by brain ischemia in rat striata. DOPA antagonist has a neuroprotective action.

DOPA cyclohexyl ester, a competitive DOPA antagonist, protects glutamate release and resultant delayed neuron death by transient ischemia in hippocampus CA1 of conscious rats.

In rat striata, DOPA released is a causal factor for glutamate release and resultant delayed neuron death by four-vessel occlusion. Nanomolar DOPA cyclohexyl ester (CHE), a potent and relatively stable competitive DOPA antagonist, protects these events. We tried to clarify whether DOPA CHE protects these events in hippocampal CA1 pyramidal cell layers most vulnerable against ischemia. Five to 10 min ischemia caused slight to mild glutamate release in 10 min samples during microdialysis and mild to severe neuron death 96 h after reperfusion. DOPA and dopamine were under assay limit in this design, but were basally detected by 20 min sampling and released by 20 min ischemia. In 10 min samples, intrahippocampal perfusion of 100 nM DOPA CHE 10 min before ischemia for 70 min did not inhibit glutamate release by 10 min ischemia, while it abolished glutamate release and protected delayed neuron death by 5 min ischemia. DOPA CHE is neuroprotective under a mild ischemic condition in rat hippocampus CA1.

The usefulness of 99mTc-HMPAO-labeled leukocyte scintigraphy in the diagnosis of skeletal metastases of cancers.

The usefulness of bone marrow scintigraphy with 99mTc-HMPAO-labeled leukocytes (leukocyte bone marrow scintigraphy) in the diagnosis of skeletal metastases of cancers was investigated in 70 lesions in 27 patients with various types of cancer. The final diagnosis of skeletal metastases was based on one or more criteria consisting of histological confirmation, typical findings of metastases by bone radiograph, CT and MRI, or progressive swellings of the lesions with severe pain due to nerve compression. Of the 70 lesions, 55 were finally diagnosed as metastases, and 15 as benign lesions. Leukocyte bone marrow scintigraphy showed photopenic defects in 52 of the 55 metastatic lesions (sensitivity 95%), and the remaining 3 negative lesions were found positive for metastases by MRI. In contrast, MRI could evaluate only 39 of the 55 lesions because 16 lesions in the ribs, scapula and sternum were not visualized. Of these 39 lesions, MRI showed positive findings for metastases in 33 (sensitivity 85%), and negative findings in 6 with photopenic defects found by leukocyte bone marrow scintigraphy. Of the 15 benign lesions, 3 were false positive for metastases on leukocyte bone marrow scintigraphy (specificity 80%). We conclude that 99mTc-HMPAO-labeled leukocyte bone marrow scintigraphy may be useful in the diagnosis of skeletal metastases of cancers, particularly when MRI fails to evaluate the lesions.

[Two children with suspected primary vasculitis of mesenteric vessels--a case report].

We reported 2 children with suspected primary vasculitis of mesenteric vessels. Both children were admitted to our hospital with the complaints of abdominal pain, bloody stool or diarrhea. Laboratory examination simultaneously revealed leukocytosis with dominant neutrophils, positive CRP, and hypoalbuminemia. Although prothrombin time and activated partial thromboplastin time were within normal limits, the increased levels of FDP-E, D-dimer, and von Willebrand factor activity were observed, which suggested the endothelial cell activation and the coagulation/fibrinolysis system activation. Abdominal echography and CT scanning demonstrated the edematous thickening of intestinal or colon walls probably due to the vasculitic permeability changes of mesenteric artery. During the disease courses, skin rash, bleeding tendency, arthritis and proteinuria were not observed, and no autoantibodies including anti-nuclear antibody, anti-DNA antibody, and myeloperoxidase-antineutrophil cytoplasmic antibody, were detected. Taken together, we suspected these children as restricted vasculitis of mesenteric vessels. Intravenous prednisolone was administrated, and the clinical and laboratory abnormalities recovered completely within 2 weeks. Thus, we suggested that the leukocyte counts, CRP, and the determination of von Willebrand factor and coagulation/fibrinolysis study accompanied with X-ray, echography, and CT scanning will be useful for the early diagnosis of vasculitis before the pathologic and irreversible vascular damage are demonstrated.

L-DOPA cyclohexyl ester is a novel potent and relatively stable competitive antagonist against L-DOPA among several L-DOPA ester compounds.

We explored L-DOPA esters with chemically bulky structures to find a potent stable competitive antagonist against L-DOPA, compared to DOPA methyl ester (DOPA ME). In anesthetized rats, DOPA cyclohexyl ester (DOPA CHE), DOPA cyclopentyl ester (DOPA CPE) and DOPA cyclopentyldimethyl ester (DOPA CPDME) at 1 microgram microinjected into depressor sites of the nucleus tractus solitarii elicited or tended to elicit more marked antagonism against depressor responses to 60 ng L-DOPA, compared to DOPA ME. At 100 ng, DOPA CHE elicited the most potent antagonism. At 1 microgram, duration of the antagonistic activity of DOPA CHE was approximately three times longer than that of DOPA ME. During microdialysis of the nucleus accumbens, conversion from DOPA CHE at 1 microM perfused via probes to extracellular L-DOPA was the lowest among these compounds and less than one half of that from DOPA ME. Binding studies showed that the recognition site for L-DOPA differs from ionotropic glutamatergic, dopaminergic D1 and D2 receptors. We recently found that L-DOPA evoked by transient ischemia may act as a DOPA CHE-sensitive causal factor for glutamate release and resultant neuronal cell death. DOPA CHE is the most potent, relatively stable competitive antagonist against L-DOPA and is a useful mother compound to develop neuroprotective drugs.

Hypothetical pathophysiology of acute encephalopathy and encephalitis related to influenza virus infection and hypothermia therapy.

BACKGROUND: To establish a treatment strategy for acute encephalopathy and encephalitis associated with influenza virus infection, the pathophysiology of the disease was investigated through manifestations and laboratory findings of patients. PATIENTS AND METHODS: A child with central nervous system (CNS) complications during the course of influenza virus infection was analyzed in view of immunologic abnormalities. In addition, four children with acute encephalopathy and encephalitis were enrolled in the hypothermia treatment for the purpose of stabilizing the cytokine storm in the CNS. RESULTS: The CNS symptoms preceded the systemic progression to the failure of multiple organs (MOF) and disseminated intravascular coagulopathy (DIC). The mild hypothermia suppressed the brain edema on computed tomography (CT) scanning and protected the brain from the subsequent irreversible neural cell damage. CONCLUSION: The replicated viruses at the nasopharyngeal epithelium may disrupt the olfactory mucosa and gain access to the brain via the olfactory nerve system. The direct virus-glial cell interaction or viral stimulation of the glial cells induces the production and accumulation of the pro-inflammatory cytokines, especially tumor necrosis factor (TNF)-alpha, in the CNS. The cytokine storm results in neural cell damage as well as the apoptosis of astrocytes, due to the TNF-alpha-induced mitochondrial respiratory failure. The disruption of the blood-brain barrier progresses to the systemic cytokine storm, resulting in DIC and MOF. Mild hypothermia appears promising in stabilizing the immune activation and the brain edema to protect the brain from ongoing functional, apoptotic neural and glial damage and the systemic expansion of the cytokine storm.

Involvement of rBAT in Na(+)-dependent and -independent transport of the neurotransmitter candidate L-DOPA in Xenopus laevis oocytes injected with rabbit small intestinal epithelium poly A(+) RNA.

Although L-3,4-dihydroxyphenylalanine (L-DOPA) is claimed to be a neurotransmitter in the central nervous system (CNS), receptor or transporter molecules for L-DOPA have not been determined. In an attempt to identify a transporter for L-DOPA, we examined whether or not an active and high affinity L-DOPA transport system is expressed in Xenopus laevis oocytes injected with poly A(+) RNA prepared from several tissues. Among the poly A(+) RNAs tested, rabbit intestinal epithelium poly A(+) RNA gave the highest transport activity for L-[(14)C]DOPA in the oocytes. The uptake was approximately five times higher than that of water-injected oocytes, and was partially Na(+)-dependent. L-Tyrosine, L-phenylalanine, L-leucine and L-lysine inhibited this transport activity, whereas D-DOPA, dopamine, glutamate and L-DOPA cyclohexylester, an L-DOPA antagonist did not affect this transport. Coinjection of an antisense cRNA, as well as oligonucleotide complementary to rabbit rBAT (NBAT) cDNA almost completely inhibited the uptake of L-[(14)C]DOPA in the oocytes. On the other hand, an antisense cRNA of rabbit 4F2hc barely affected this L-[(14)C]DOPA uptake activity. rBAT was thus responsible for the L-[(14)C]DOPA uptake activity expressed in X. laevis oocytes injected with poly A(+) RNA from rabbit intestinal epithelium. As rBAT is localized at the target regions of L-DOPA in the CNS, rBAT might be one of the components involved in L-DOPAergic neurotransmission.

[Why there were few isolations of tubercle bacilli in young children with pulmonary tuberculosis?].

We retrospectively evaluated the factors influencing the isolation of tubercle bacilli in 51 children under 14 years of age with pulmonary tuberculosis who were admitted to Yokohama City University Hospital from 1975 to 1998. Young children (0-6 years of age) with pulmonary tuberculosis were significantly less positive by smear and culture than elder children (7-14 years of age) with pulmonary tuberculosis. According to the Japanese Society for Tuberculosis classification of finding on chest X-ray film for pulmonary tuberculosis, the culture-positive patients with type II (cavitary lesions) were found in all, the culture-positive patients with type III (non-cavitary lesions) in 39.3%, and the culture-positive patients with type H (hilar and mediastinal lymphadenopathys) in 35.3%. Patients with cavitary lesions (type II) were significantly more positive by smear and culture than patients with non-cavitary lesions (type III + type H). Only 15.8% of the young children with pulmonary tuberculosis had received BCG vaccine and all had non-cavitary lesions (type III and type H). But, 84.6% out of the older children had received BCG vaccine and half had cavitary lesions. Taken together, the result was that there were few isolation of tubercle bacilli in young children with pulmonary tuberculosis because they had non-cavitary tuberculosis without delayed-type hypersensitivity to tubercle bacilli.

[Two infants with classical polyarteritis nodosa but not Kawasaki disease].

We experienced two infants with polyarteritis nodosa (PN). The symptoms started with high fever and skin rash, which were similar to those of Kawasaki disease (KD). However, the involvement of central nervous system and lung, such as distension of large fontanel, stridor and mild disturbance of consciousness, occurred and the systemic vasculitis resulted in hypoalbuminemia and severe generalized edema. They flared up twice or three times during the long clinical courses. Finally, both had multiple giant coronary aneurysms and lung fibrosis. The clinical courses of these patients were different from those of KD in that: (1) the severity of vasculitis, (2) the wide-spread nature of the vasculitis, and (3) the chronic and recurrent clinical course. It is very difficult to distinguish PN from severe KD in the early stage of the diseases especially in infancy. But in the cases intractable to high-dose gamma-globulin therapy and plasma exchange, it is needed to suspect PN and to induce more aggressive immunosuppressive therapy, such as methylprednisolone pulses and cyclophosphamide pulse therapy as soon as possible.

[An infant of autoimmune hepatitis (type I) with cirrhosis].

A 6 year-old boy with autoimmune hepatitis accompanied with cirrhosis was reported. He was admitted to our hospital because of abdominal distention, high fever, and diarrhea. Laboratory examination revealed abnormalities in hepatic function, cholestasis, anemia, thrombocytopenia, hypoalbuminemia, hypocomplementemia, and low concentration of coagulation factors. Abdominal MRI, and asialoglycoprotein receptor-mediated liver scintigraphy strongly indicated liver cirrhosis. Viral hepatitis, Wilson's disease, and antitrypsin deficiency were excluded serologically. Instead, hypergammaglobulinemia, and positive antinuclear antibody suggested autoimmune hepatitis, and the survey of anti-mitochondrial antibody, anti-smooth muscle antibody, and anti-LKM-1 antibody was negative, indicating type I autoimmune hepatitis. Finally, the histology of liver biopsy specimen indicating the destruction of hepatic lobular architecture, dense mononuclear cell infiltrates, and severe fibrosis confirmed the diagnosis. He was treated firstly with methylprednisolone pulses, and then prednisolone p.o. + azathioprine p.o. All of the abnormal laboratory parameters improved to normal levels, indicating that the immunosuppressive therapy will be effective for the severe AIH with cirrhosis.

[Analysis of tuberculin reaction of tuberculous children below 4 years of age].

To clarify whether the size of tuberculin reaction could be used as an useful index of the severity of tuberculosis, we analyzed the sizes of tuberculin reaction (TR) of 60 children below 4 years of age with active tuberculosis at the time of diagnosis. Of 60 patients, 53 (88.9%) had positive reactions to tuberculin. The mean size of TR of 60 patients was 24.0 +/- 13.9 mm and maximum size was 60 mm. Seven patients who had no reaction to the tuberculin skin test consisted of three primary complex and four serious tuberculosis (two miliary tuberculosis and two tuberculous meningitis). The patients without BCG vaccination showed significantly smaller TR than the patients with BCG vaccination (p < 0.05). The patients less than 1 year of age showed significantly smaller TR than the patients of 4 years of age (p < 0.05). The patients with serious tuberculosis showed significantly smaller TR than the patients with primary complex (p < 0.05). Of patients with primary complex, there were no difference of the size of TR between the patients with pulmonary tuberculosis (III) and hilar lymphadenopathy (H). Together with, it did not necessarily mean that negative TR showed no infection with tuberculosis and the sizes of TR depended on the severity of tuberculosis in infantis and young children.

[Effective combination therapy of plasma exchange and subsequent cyclophosphamide pulses for catastrophic antiphospholipid antibody syndrome: a case report].

A 7-year-old girl with catastrophic antiphospholipid antibody syndrome was described. She firstly admitted to the local hospital with the complaints of persistent fever and abdominal pain, and was diagnosed as systemic lupus erythematosus with the laboratory findings as follows; positive for antinuclear antibody, anti-DNA antibody, and platelet-associated IgG, thrombocytopenia, and hypocomplementemia. 10 days after the initiation of oral prednisolone, she suddenly manifested tonic convulsion and unconsciousness accompanied by high fever. Because of the unresponsiveness to the methylprednisolone pulse therapy for supposed CNS lupus, she was transferred to our hospital. Her unconsciousness persisted, and pulsation on dorsalis pedis was not palpable on admission. Laboratory investigation revealed the falsely positive VDRL, a prolonged aPTT, positive for lupus-anticoagulant and antiphospholipid antibody. The magnetic resonance image demonstrated multiple spotty hyperintensity (T2) in the brain consistent with multiple hemorrhagic infarcts. Arteriogram demonstrated the infarct of dorsalis pedis, and coronary aneurysms. These findings were compatible with the criteria of catastrophic antiphospholipid antibody syndrome, she was diagnosed as catastrophic antiphospholipid antibody syndrome. The plasma exchange and subsequent cyclophosphamide-pulse therapy, which was given once a month for first 6 months, and later, at 3 months intervals, was effectively administered. This combination and oral anti-thrombotic therapy revealed effective for this kind of fatal disorder.