CD26-mediated signaling for T cell activation occurs in lipid rafts through its association with CD45RO.

CD26 is a T cell activation antigen that contains dipeptidyl peptidase IV activity and is known to bind adenosine deaminase. The mechanism by which CD26 costimulation potentiates T cell receptor-mediated T cell activation, leading to subsequent exertion of T cell effector function, is still not clearly defined. In this article, we demonstrate that CD26 localizes into lipid rafts, and targeting of CD26 to rafts is necessary for signaling events through CD26. Importantly, aggregation of CD26 by anti-CD26 mAb crosslinking also causes coaggregation of CD45 into rafts. Moreover, we show that CD26 directly binds to the cytoplasmic domain of CD45. Our results therefore indicate a mechanism whereby CD26 engagement promotes aggregation of lipid rafts and facilitates colocalization of CD45 to T cell receptor signaling molecules p56(Lck), ZAP-70, and TCRzeta, thereby enhancing protein tyrosine phosphorylation of various signaling molecules and subsequent interleukin-2 production.

Error-related brain potentials elicited by vocal errors.

Error-related negativity (ERN) is observed immediately after an error in choice reaction time tasks performed by hand response. We examined whether the ERN occurs in relation to slips of vocalization in the Stroop color word task. In one condition, the subject's vocal responses were masked by continuous pink noise in order to prevent vocalization-related cortical potentials from contaminating the ERN time window. This masking procedure was successful in inhibiting the vocalization-related cortical potential. More importantly, vocalization errors elicited a frontocentral negative-going deflection followed by a positive component immediately after the error response regardless of the masking condition. The present results suggest that the error detection mechanism may also elicit an ERN-like component in response to vocal slips.

Decreased expression of transcription factor GATA-2 in haematopoietic stem cells in patients with aplastic anaemia.

Aplastic anaemia is characterized by reduced haematopoiesis resulting in pancytopenia. It has been speculated that there is an injury in haematopoietic stem cells in the bone marrow; however, the precise nature of the injury has not been elucidated. In this study, the levels of expression of mRNAs for three transcription factors, GATA-2, SCL and AML1, which function in the early stages of haematopoiesis, were examined by quantitative polymerase chain reaction in patients with aplastic anaemia, idiopathic thrombocytopenic purpura (ITP) and normal subjects. Among these factors, expression of GATA-2 mRNA in purified CD34-positive cells was markedly decreased in aplastic anaemia compared with that in ITP and in normal subjects. The expression levels of SCL and AML1 mRNA in CD34-positive cells in aplastic anaemia were not different from those in normal subjects. When the expression of GATA-2 protein in CD34-positive cells was examined by immunocytochemical analysis, the percentage of GATA-2-positive cells in aplastic anaemia was lower than that in normal subjects. These findings strongly suggest that there is an aberrant expression of transcription factors in stem cells in aplastic anaemia, which may be responsible for the development of the disease.

Expression of gp34 (OX40 ligand) and OX40 on human T cell clones.

gp34, which we previously cloned, is a ligand of OX40 (CD134), a costimulatory molecule involved in T cell activation. To elucidate the role of human OX40 / OX40L interaction, we examined the expression of gp34 (OX40L) and OX40 in normal human hematopoietic cells by using flow cytometry. OX40 expression is observed on activated T cells, while OX40L is expressed in antigen-presenting cells. However, cytotoxic T lymphocyte (CTL) clones specific for Epstein-Barr virus (EBV)-transformed autologous lymphoblastic cell lines (LCLs) induced both OX40 and OX40L expression after antigen or T cell receptor (TCR) stimulation. This study suggests a possible function of OX40L / OX40, through T cell-T cell interaction, in the reactivation of memory T cells in an autocrine manner, with implications for the pathogenesis of viral infections and neoplasms.

A repressor element in the 5'-untranslated region of human Pax5 exon 1A.

B-cell-specific activator protein (BSAP) encoded by the Pax5 gene plays a critical role during B-cell development. We have analyzed the 5'-flanking region plus the 5'-untranslated region (5'-UTR) of human Pax5 exon1A to clarify its regulatory mechanisms. Functional dissection of these regions by luciferase reporter assays indicated that a cluster of regulatory elements acts as a strong repressor between +320 and +453. Insertion of this segment between the heterologous simian virus 40 (SV40) promoter and the luciferase gene in both the sense and reverse orientation sharply reduced the luciferase activity, but insertion into the upstream of the SV40 promoter did not. This suggests that this segment must be located in the 5'-UTR to function effectively. A search through databases with the sequence of this segment did not reveal any known DNA binding factor site. Electrophoretic gel mobility shift assay (EMSA) experiments demonstrated that unknown factors bound to the fragment +408 to +429. Insertion of this fragment between the SV40 promoter and the reporter gene strongly suppressed the luciferase activity. Competitive EMSA indicated that the region between nucleotides +413 and +427 encompassed the binding site of the unknown factors and was hence regarded as a repressor element. Mutagenesis in this element significantly recovered reporter gene activity. These results suggest that the segment +320 to +453, especially the repressor element +413 to +427, in the 5'-UTR is involved in the regulation of Pax5 gene expression.

Chemically modified ribozyme to V gene inhibits anti-DNA production and the formation of immune deposits caused by lupus lymphocytes.

A variety of autoantibodies is responsible for the tissue injury in autoimmune diseases. We have demonstrated that the human anti-DNA Ab O-81, of which Ids are commonly detected in renal glomeruli of active lupus nephritis, uses the V3-7 gene. We tried to develop a new therapy for lupus nephritis by using chemically modified ribozymes to specifically inhibit the expression of the mRNA of Ig V gene. The transfection of hammerhead ribozyme or the addition of chemically modified ribozyme against the flanking region of V3-7 caused a potent and selective inhibition of anti-DNA production in V3-7-using B cell clones, but not in irrelevant V gene-using clones in vitro. Chemically modified ribozyme was long-acting and resistant to RNase, and nonspecific cytotoxicity of the ribozyme was negligible. To know the efficacy of the ribozyme in vivo, we used a model of immune complex nephritis in SCID mice in which 5 x 10(6) PBLs from patients with active lupus nephritis (lupus PBL) were transferred twice. The injection of lupus PBL in combination with chemically modified ribozyme to increase resistance to RNase significantly reduced anti-DNA Ab levels in blood and decreased levels of urinary protein in the immune deposit models. Immunofluorescence study also revealed a marked decrease in IgG deposits at renal glomeruli in the ribozyme-treated group. These results indicate an efficacy of chemically modified ribozyme therapy for autoantibody-mediated immune diseases.

An electrophysiological study of the locus of the interference effect in a stimulus-response compatibility paradigm.

This study investigated the locus of the interference effect in a stimulus-response compatibility task using event-related potentials (ERPs). Ten participants were instructed to respond to stimulus color with the left or right middle finger. Red or blue arrows pointed in the same direction as the response hand on congruent trials and pointed in the opposite direction on incongruent trials. Neutral trials were red or blue horizontal bars. Reaction times (RTs) to incongruent stimuli were significantly longer than RTs to congruent stimuli. The peak latency of the P300 for incongruent stimuli was significantly longer than that for congruent stimuli. In addition, onset of stimulus-locked lateralized readiness potential (LRPs) was significantly later for incongruent stimuli than for congruent stimuli. However, electromyogram (EMG)-locked LRPs for incongruent stimuli showed incorrect preparation. These results suggest that the interference effect might occur at the stage in which stimulus evaluation processes and response-related processes overlap.

Human movement-related brain potentials preceding voluntary movements in different arousal states monitored with skin potential level.

The present study investigated the readiness potential (RP) preceding a brisk extension of the right middle finger during different arousal states as monitored by skin potential level (SPL). The late component of the readiness potential in the medium arousal state was significantly larger than those in the low and high arousal states. This finding indicates that the RP waveform may vary as a function of arousal states, suggesting the inverse U-shaped relationship proposed in studies of the contingent negative variation.

Enhanced negative slope of the readiness potential preceding a target force production task.

Readiness potentials (RPs) preceding a trigger pulling movement were recorded in 9 right-handed male subjects. We investigated two tasks, non-purposive and purposive movement tasks. In this study we defined simple trigger pull as non-purposive, and target force production by pulling the trigger as purposive. In the non-purposive task, the subjects were instructed to pull the trigger at their own pace and at an easily-exerted force level. After two sessions in the non-purposive movement task, the subjects were submitted to the purposive movement task, and were requested to pull the trigger in an attempt to produce target force, the range of which was decided individually on the basis of mean force level in the second session of the non-purposive movement task. The RP preceding the purposive movement was larger than that preceding the non-purposive movement. In addition, enhancement of the RP was specific to the negative slope (NS'). Since neither peak force nor time to peak force of the movement differed in the two tasks, it was concluded that the increased NS' was due to a psychological change associated with execution of the purposive movement.

HTLV-I Tax trans-activation and cell growth signaling.

We have cloned two genes for cell surface molecules, capable of delivering the intracellular signals, which are modulated for their expression by Tax. One is the gamma chain of the interleukin-2 (IL-2) receptor which is suggested to be critical for IL-2-dependent growth of human T-cell leukemia virus type I (HTLV-I) infected cells. The gamma chain is upregulated by Tax, like the IL-2 receptor alpha chain. This upregulation may compensate the gamma chain downregulation after IL-2 binding, presumably resulting in more frequent growth of HTLV-I infected T cells. The other is gp34 that was initially identified as a molecule specifically expressed on HTLV-I-infected T cells. gp34 has been demonstrated to bind OX40 which belongs to the tumor necrosis factor (TNF) receptor family. We found that HTLV-I Tax induces expression of gp34 and OX40, and that normal T cell transiently express both gp34 and OX40 upon antigenic stimulation. Collectively, it may be possible that HTLV-I-infected T cells are in a predisposition to growth due to modulated expression by HTLV-I Tax of gp34/OX40 and the gamma chain.

Induction of OX40, a receptor of gp34, on T cells by trans-acting transcriptional activator, Tax, of human T-cell leukemia virus type I.

gp34, which we had identified as a target molecule of the trans-activation by Tax of human T-cell leukemia virus type I (HTLV-I), has been found to bind OX40, a member of the tumor necrosis factor receptor family, resulting in growth stimulation of activated T cells. We here demonstrate that not only gp34 (OX40L), but also OX40 can be transcriptionally activated by Tax. Three Tax-producing human T-cell lines carrying the HTLV-I genome expressed OX40 on their surfaces. Furthermore, Tax-induced transcriptional activation of OX40 was shown in Tax-inducible JPX-9 cells. These results demonstrate that both OX40 and its ligand (gp34) are constitutively expressed on the surfaces of Tax-expressing T lymphocytes, suggesting that the OX40L/OX40 system contributes to growth stimulation of the virus-infected T cells.

Isozyme selective induction of mouse pulmonary superoxide dismutase by the exposure to mercury vapor.

Alterations in lung superoxide dismutase (SOD) isozymes after exposure of mice to mercury vapor were examined. Inhalation of mercury vapor (10 mg/m(3)) for 1 h by mice resulted in a higher accumulation of mercury in the kidney and lung compared to other organs, at 1 h after exposure. Under these conditions marked enhancement of protein content in bronchoalveolar fluid (BALF), attributed to lung injury, was observed. Exposure to mercury vapor caused a significant increase in the pulmonary Cu,Zn-SOD activity (1.32-fold at 48 h) whereas Mn-SOD activity was suppressed to 82% of the control level, suggesting different sensitivity to the metal inhalation. The selective induction of Cu,Zn-SOD protein (1.79-fold at 48 h) was confirmed by immunoblot analysis with polyclonal antibodies against these isozymes. These observations suggest that the selective induction of Cu,Zn-SOD at the translational level appears to occur as an initial defense against mercury-promoted oxidative stress.

Selective inhibition of the mouse brain Mn-SOD by methylmercury.

Changes in mRNA levels, protein contents and enzyme activities for brain Cu,Zn- and Mn-SOD by methylmercury chloride (MMC) administration, were examined, over a period of 12 days in ICR male mice. After subcutaneous administration of MMC (10 mg/kg) to mice, brain mercury content reached a maximum at 2 days and remained at that level for at least 5 days. MMC exposure resulted in a time-dependent decrease in the Mn-SOD activity: the enzyme activity at 5 days after exposure to MMC was about 60% of control level whereas this exposure was without effect on the Cu,Zn-SOD activity, indicating differential sensitivity of SOD isozymes to the metal. However, levels of mRNA and protein synthesis for Mn-SOD were unaffected by MMC administration. The direct effect of MMC on the both SOD activities were further examined with purified enzyme preparations. After each SOD isozyme (10 U) was incubated with 0.2 mM MMC for 24 h at pH 7.8, the enzyme activities for Cu,Zn- and Mn-SOD were 90% and 37% of control, respectively. Incubations at a ratio of SOD to MMC (1 : 600) for 24 h resulted in a substantial decrease in the enzyme activity of the Mn form; this isozyme-selective inactivation was noted at alkaline pH. A combination of isoelectric focusing-agarose gel electrophoresis (IEF-AGE) and synchrotron radiation X-ray fluorescence (SR-XRF) analysis revealed that Mn-SOD rather than Cu,Zn-SOD underwent modification. Furthermore, a decrease in native form of Mn-SOD protein after MMC exposure was confirmed by gel filtration chromatography. These results indicate that Mn-SOD, but not Cu,Zn-SOD, is susceptible to modification by MMC and the resulting alteration in structure appears to cause a loss of enzyme activities.

Functional analysis of the human interleukin 2 receptor gamma chain gene promoter.

The third component of the interleukin (IL) 2 receptor, gamma chain, is essential not only for IL-2- but also for IL-4-, IL-7-, IL-9-, and IL-15-induced proliferation of lymphocytes. To elucidate the mechanisms by which the gamma chain is expressed, we have analyzed the promoter region of the gamma chain gene. The 633-base pair fragment upstream of the initiation codon showed the promoter activity in human hematopoietic cell lines, Jurkat and THP-1, when linked to the luciferase gene. With a series of 5'-deletion mutants, the basal promoter activity was found in a fragment from nucleotide 80 to 58 upstream from the RNA start site, including an Ets binding sequence. Treatment of cells with either 12-O-tetradecanoylphorbol-13-acetate or phytohemagglutinin but not forskolin induced transcription from the gamma chain gene promoter. A viral trans-acting transcriptional activator, Tax, of human T-cell leukemia virus type I elevated expression of the gamma chain gene. In contrast, IL-2 decreased transcription from the IL-2 receptor gamma chain promoter. These results suggest that expression of the gamma chain is regulated at the transcription level by extracellular stimuli and may be implicated in immune response.

Compressed heart and mitral valve prolapse in a case of Duchenne's progressive muscular dystrophy with thorax deformity.

We report on a 15 year old patient with Duchenne's progressive muscular dystrophy who demonstrated a narrowing of the left ventricular inflow and outflow tracts due to compression by a highly deformed thoracic spine. A systolic murmur (4/6) with thrill and a diastolic murmur (2/6) were heard, with these murmurs being louder in the expiratory phase as compared with the inspiratory phase. The second heart sound showed a paradoxical splitting. Echocardiograms revealed a compressed and narrowed left ventricle and a prolapsed mitral valve. The intensities of the heart murmurs changed synchronously with the chamber's narrowing due to respiration. A narrowed left ventricle occurring as a result of the compression by the deformed thoracic spine is thought to be the cause of these cardiac findings.

Ultrastructural studies on the neuromuscular junctions of Becker's muscular dystrophy.

Ultrastructural studies on muscle biopsies from three patients with Becker's muscular dystrophy showed that the i.m. nerves presented loss or disarrangement of the neurofilaments and an increased number of glycogen granules and/or myelin figures not infrequently in the myelinated and unmyelinated nerve fibers. The neuromuscular junctions showed markedly widened sole-plate areas, and several terminal axons frequently abutted and formed neuromuscular junctions on the same fiber. The secondary synaptic clefts were markedly decreased in number and short in length in type I fibers but not in type II fibers. Most terminal axons showed no degenerative changes. Therefore, the participation of a neural factor might be suggested as the cause of Becker's muscular dystrophy, although it does not mean denervation in the conventional sense of an axonal degeneration.

The causative mechanisms of mitral valve prolapse in progressive muscular dystrophy in reference to thorax and thoracic spine deformities and left ventricular dysfunction.

The causative mechanisms of mitral valve prolapse (MVP) were evaluated in 58 patients with progressive muscular dystrophy (PMD). Two possible causes, 1) left ventricular (LV) dysfunction and 2) thoracic spine and thorax deformities were assessed. Patients were classified into three groups by echocardiographic findings. Group 1: 31 patients without MVP, group 2: 11 patients with MVP confirmed only by M-mode echocardiogram, group 3: 16 patients with MVP confirmed by both two-dimensional and M-mode echocardiograms. LV functions evaluated by systolic time intervals and fractional shortening showed no significant differences among the three groups. Scoliosis of the thoracic spine was not related to the incidence of MVP. Lordotic or straight spines were found in 32.3%, 100%, 93.8% of cases in group 1, group 2 and group 3, respectively, and the incidences of MVP in cases with kyphosis, straight spine and lordosis were 4.8%, 66.7% and 77.8%, respectively. The shape of the thorax as evaluated by the ratio of anteroposterior internal diameter to transverse diameter was more flattened in groups 2 and 3 than in group 1. From these results, we concluded that LV dysfunction was not related to the incidence of MVP and that the lordotic or straight spine and the flattened thorax were supposed to be the major factors in the occurrence of MVP in PMD.