Biomedical subjects
Publications and source records attributed to M Negishi.
The structure of estrogen sulphotransferase has been solved in the presence of inactive cofactor PAP and substrate 17 beta-estradiol. This structure reveals structural similarities between cytosolic sulphotransferases and nucleotide kinases.
BACKGROUND: The actions of prostaglandin F2alpha are mediated by a cell-surface receptor (FP), but little is known about the regulation of FP gene expression. To clarify the mechanisms underlying tissue specific transcription of the mouse FP gene, we isolated and characterized mouse genomic DNA clones encoding FP. RESULTS: Structural analysis revealed that the mouse FP gene is composed of three exons and two introns, and spans more than 11 kilobases. By primer extension and PCR analyses, the major transcription start site was identified as a cytosine nucleotide, but additional sites of transcription initiation were found in the ovary. There was no apparent difference in the FP gene transcription initiation site between the ovary, kidney and stomach. Sequence analysis of the putative promoter region showed only two potential SP-1 binding sites, but no other typical well-known consensus sequences. We generated transgenic mice with the potential promoter region of the FP gene connected to the lacZ reporter gene. Northern blot analysis showed that the pattern of expression of the transgene corresponded to that of FP expression, except in the ovary. Upon analysis by in situ hybridization, the lacZ gene transcripts were found to be expressed in the fundic glands in the stomach, and the cortical tubules in the kidney, in which endogenous FP transcripts were also expressed. On the contrary, expression of lacZ transcripts was not detected in the corpora lutea, where the highest expression of FP mRNA was observed. CONCLUSIONS: These studies suggest that a separate control mechanism exists for FP expression in the ovary, distinct from the expression in the stomach and kidney.
The secretion pattern of putuitary growth hormone (GH) in rodents is sexually dimorphic and regulates the expression of some hepatic genes. We report cloning of mouse glycine N-methyltransferase (GNMT) cDNA. The mouse GNMT mRNA is expressed at much lower levels in male than in female livers. Hypophysectomy in male mice resulted in as a high level of the expression as in female mice. Treatment with GH of hypophysectomized male mice twice a day for 5 days decreased the expression. These results indicate that the sex difference of the mouse GNMT expression is regulated by GH secretion patterns.
The mission of the Tokyo Metropolitan Komagome Hospital is to build a comprehensive system to provide treatment for people infected with HIV. It aims not only to provide HIV clinical care (medical and skilled nursing) but also to build psychological and social support systems for persons with HIV/AIDS. It is also working to organize the medical care system's response to HIV and to help decrease misunderstanding, prejudice, and discrimination against persons with HIV infection.
We examined the relationship of somatic complaints to coping behaviors and mood states among 50 HIV-positive patients without AIDS. Although no patients fulfilled the DSM-III-R criteria for mood disorders including major depression, scores for depressive symptoms were significantly higher in the HIV-positive patients than in healthy persons. Although depressive symptoms in HIV patients may not be strong enough to warrant a psychiatric diagnosis of mood disorders, these patients may be prone to depressive symptoms. The HIV patients indicated a tendency toward somatic complaints more frequently than their healthy counterparts. The scores for depressive symptoms were significantly and positively correlated with scores for avoidance coping responses. The presence or absence of six complaints (i.e., general fatigue, abdominal distress, chest pain or discomfort, and numbness or chills) could be discriminated based on the score of avoidance coping responses. The results of this study suggest that avoidance coping responses associated with depressive symptoms accompany several somatic complaints in HIV patients without AIDS.
We examined the efficacy of relaxation techniques in a sample of HIV patients without AIDS in the early stages after infection, by comparing the three groups: relaxation group (progressive muscle relaxation and modified autogenic training); ordinary supportive psychotherapy group, and finally no psychiatric treatment group. Scores for anxiety, fatigue, depression and confusion, as measured by the profile of mood states (POMS), were significantly lower after relaxation than before. There were no significant differences in the POMS scores (except for anger) among the three groups. These two results suggest that a combination of progressive muscle relaxation and modified autogenic training is a useful method, which can be easily employed in HIV patients without AIDS.
Varicella has been thought to be one of the representative infectious disease in childhood, but recently we are under the impression that adults contracting varicella are increasing in number. On the other hand, they say that varicella generally causes a serious illness in adult patients. So we investigated signs and symptoms of varicella, source of infection, occupations of adult patients, except those who were immunologically compromised, by means of medical records, to know the characteristics of varicella in adulthood. According to the varicella severity score proposed by Nagai et al., varicella in the hospitalized adult patient was found to be much severer than that in children. The most remarkable symptoms, were high fever and sore throat, and these were the main reason of hospitalization in most of our patients. Although severity scores were very high in admitted adult patients with varicella, their clinical courses were not serious, and most of them recovered with only supportive therapy. These patients rarely suffered from complications, like pneumonia. If adult patients with varicella hospitalized in the early stage and received supportive care, they could recover without any complications. In most cases of adult varicella the source of infection was unknown. In the case of married persons, however, many of them were infected through their child. When adults contract varicella, not only the patients themselves suffer from high fever and sore throat, but also they act as the source of infection, if they are medical care workers. Furthermore, in public, the contraction of varicella results a socioeconomic loss from suspension of business caused by the illness. Prophylaxis with varicella vaccine, therefore, should be considered, when there are people who have never contracted varicella, whether or not they are medical staff.
Regional and cellular distribution of mRNAs for prostaglandin E (PGE) receptor subtypes was investigated in the mouse gastrointestinal tract by in situ hybridization. Strong signals for EP1 transcripts were detected in cells of the muscularis mucosae layer, especially in the body of the stomach. Intense signals for EP3 transcripts were detected in neurons of the myenteric ganglia throughout the tract. Moderate EP3 mRNA expression was also observed in fundic gland epithelial cells, except for surface mucous cells in the stomach. Expression of EP4 mRNA was moderate in surface epithelial cells of the corpus and in glands from the surface to the base of the antrum. Strong EP4 signals were observed in the epithelium in the duodenum, jejunum, and ileum. In the ileum, signals were only observed in the upper part of the villi. However, no or weak signals for EP2 transcripts were detected. These findings suggest that PGE2 modulates various gastric or intestinal functions via at least three different PGE receptors.
Stereotactic posteroventral pallidotomy was carried out in 13 cases with rigid-akinesia-type Parkinson's disease with the aid of depth microrecording. The outcome of the pallidotomy was classified into four groups: excellent (6 cases), good (3 cases), moderate (3 cases) and fair (1 case). Electrophysiological study during the operation showed continuous high-frequency and high-amplitude spike discharges in the globus pallidus (GPi) in the excellent-response group. Before the operation, a PET study had revealed low regional cerebral glucose metabolism in the prefrontal area in all the patients. Unilateral sequential opposite finger movement induced less increase of regional cerebral blood flow both in the supplementary and primary motor area in the less effective-response groups. The indications of pallidotomy for the treatment of rigid-akinesia-type Parkinson's disease are discussed.
BACKGROUND: Although several studies have indicated the association of mood states including depression with somatic complaints in HIV patients, there has been no report on Japanese HIV patients. The purpose was to examine the relationship between somatic complaints and mood states in HIV patients without AIDS. METHODS: Subjects were 50 HIV patients without AIDS. As controls, we used 47 healthy persons without HIV, matched for age and gender of the subjects. After the Structured Clinical Interview for the DSM-III-R, several inventories including the Profile of Mood States (POMS) were administered. The presence or absence of somatic complaints were examined. RESULTS: No patients fulfilled the criteria for the DSM-III-R Mood Disorders Including Major Depression. However, scores on depression as measured by the POMS were significantly higher for the HIV group than for the healthy group. Although depressive symptoms in the HIV group may not be strong enough to warrant a psychiatric diagnosis of mood disorders, these HIV patients may be prone to a depressive state. The HIV patients indicated somatic complaints more frequently. Among these, the presence or absence of five complaints (i.e., abdominal distress, sleep disturbance, smothering sensations, chest pain or discomfort, and numbness or chills) was significantly discriminated based on scores on depression. DISCUSSION: The results suggest that depressive symptoms accompany several somatic complaints in HIV patients without AIDS. Although a longitudinal follow-up study is required, the assessments of somatic symptoms may bring clinical benefits such as earlier diagnosis and/or intervention for a depressive state.
As an initial step to clarify the mechanisms of various uterine actions of PGE2, expression patterns of the messenger RNAs (mRNAs) for four subtypes of PGE receptors, EP1, EP2, EP3, and EP4, were investigated in the mouse uterus during pseudopregnancy. Relative expression levels were investigated by Northern blot analysis of mRNA levels in uteri obtained on days 0, 1, 3, 5, 7, and 9 of pseudopregnancy (day 0 = 48 h after PMSG injection), and cellular localization was determined by in situ hybridization in uteri obtained on days 0 and 5. EP2 mRNA was specifically expressed on day 5, and its expression was confined to the luminal epithelium. On the other hand, the level of the EP3 mRNA expression progressively increased until day 5. Cell populations expressing the EP3 mRNA were confined to the longitudinal smooth muscle on day 0, but they changed to the circular smooth muscle on day 5. The expression level of EP4 mRNA was low on days 0 and 1, but it became high on days 3 and 5. On day 0, EP4 mRNA was localized to the luminal epithelium. On day 5, diffuse, but significant, EP4 expression was observed over the endometrial stroma and epithelium. No EP1 mRNA signals were observed. Transient expression of EP2 on day 5 of pseudopregnancy in the luminal epithelium suggests its involvement in blastocyst implantation signaling. EP4 in the endometrial stroma is suggested to be involved in decidual transformation of the stromal cells, whereas EP3 in the myometrium is believed to be involved in regulation of myometrial activity.
Estrogen sulfotransferase (EST) is a cytosolic enzyme that catalyzes the specific sulfonation of estrogens at the 3-hydroxyl position using 3'-phosphoadenosine-5'-phosphosulfate as an activated sulfate donor. Sulfated estrogens no longer bind to the estrogen receptor and are, therefore, hormonally inactive. Although liver has been considered a primary site for steroid sulfotransferase activities, we previously have cloned the mouse EST complementary DNA and found the enzyme to be expressed abundantly in the testis of normal mice. In this study we show by reverse transcription-PCR that EST is also expressed in the testes of rat and man, suggesting that testicular expression of EST may be a common phenomenon among different species. Using a purified polyclonal antibody raised against the bacterially expressed mouse EST protein, we demonstrate by immunohistochemistry that EST is localized selectively to the androgen-producing Leydig cells within the mouse testis. Additionally, we show that Leydig cell expression of EST is under the control of the pituitary hormone LH and is regulated differentially during development. In contrast to the high level of expression in mature intact animals, EST is not present in Leydig cells of hypophysectomized mice or in Leydig cells of fetal and prepubertal (day 5 or 17) mouse testes. Administration of hCG to hypophysectomized mice restored the testicular expression of EST. Together, these results suggest that testicular expression of EST may play an important role in male reproduction, conceivably by modulating the activity of locally synthesized estrogen in the testis of a sexually mature animal.
The role of the ventral projections of the dorsal raphe nucleus (DRN) in regulating lordosis was examined in female rats. The ventral neural fibers of the DRN were transected by the horizontal circle-cut (VC), anterior (AVC) or posterior half-circle cut (PVC) at the same time as ovariectomy. Three weeks after the surgery, all the rats were implanted with a Silastic tube containing estradiol (E2). The behavioral test was started one day after E2-implantation, and 5 tests in total were carried out every other day. The results showed that the VC females had a higher lordosis quotient (LQ) than control rats. Furthermore, as well as females with the VC, a high score LQ was observed in the AVC rats but not in the PVC females, compared to those in the control animals. These results suggest that the anteroventral neural fibers of the DRN are involved in the lordosis inhibiting mechanisms in female rats.
The relationship between the expression of mRNA encoding the prostaglandin F2 alpha (PGF2 alpha) receptor and luteal cell apoptosis as determined by oligonucleosome formation was determined in mouse corpora lutea on days 2 (early phase), 6 (mid-phase), and 11 and 13 (late phase) of pseudopregnancy. No signals for mRNA encoding the PGF2 alpha receptor were detected in the ovary as shown by RNA blot analysis of pregnant mares' serum gonadotrophin (PMSG)-treated mice. The expression of the mRNA that could be detected in the corpora lutea on day 2 after hCG treatment was low, but it was increased on day 6 and reached a plateau on days 11 and 13. On days 11 and 13, in situ hybridization signals for mRNA encoding the PGF2 alpha receptor were localized to large luteal cells of the corpora lutea, especially the cells in the superficial layer. However, ethidium bromide staining revealed marked oligonucleosome formation in the corpora lutea on days 11 and 13. Similarly, positive signals of in situ nick-DNA-end labelling were also detected in the corpora lutea on days 11 and 13. Analysis of the adjacent sections of the corpora lutea on day 13 showed that both signals for DNA strand breaks and mRNA encoding the PGF2 alpha receptor were co-expressed in the corpus luteum. These results suggest that apoptosis during structural luteolysis closely associates with the increased expression of mRNA encoding the PGF2 alpha receptor in luteal cells of pseudopregnant mice.
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Cardiac myxoma cells produce large amounts of interleukin (IL)-6 and IL-8. To determine whether immunosuppressive agents could be used to treat cardiac myxoma, we tested the effects of dexamethasone and three of the newer second-generation immunosuppressive drugs, cyclosporin A, tacrolimus, and deoxyspergualin, on the production of IL-6 and IL-8 in these cells. Cultured cardiac myxoma cells were used as in vitro model of cardiac myxoma. Cells were tested for 24 hours with 10(-7) M dexamethasone, 10(-6) M cyclosporin A, 10(-8) M tacrolimus, and 10(-6) M 15-deoxyspergualin, with aliquots of conditioned medium being assayed for cytokine levels at 0, 6, 12, and 24 hours. Cardiac myxoma cells isolated from 4 patients all produced quantities of IL-6 and IL-8. The concentrations of IL-6 in the medium after 7 days in culture ranged from 79,000 to 2,740,000 pg/ml, and the concentrations of IL-8 ranged from 40,000 to 1,000,000 pg/ml. Exposure of cyclosporin A and dexamethasone almost completely inhibited the production of IL-6 and IL-8 after 24 hours of treatment. Tacrolimus inhibited the production of both cytokines by 55%, while 15-deoxyspergualin reduced IL-6 levels by 24% and IL-8 levels by 48% after separate 24 hour treatments. These results suggest that these newer immunosuppressive agents may be useful in reducing the production of IL-6 and IL-8 in patients with cardiac myxoma.
Prostaglandin E receptor EP3 subtype is widely distributed in the nervous system and is specifically localized to neurons, suggesting that the EP3 receptor plays important roles in the nervous system. We established a PC12 cell line that stably expresses the EP3B receptor isoform isolated from bovine adrenal chromaffin cells and examined the effect of agonist stimulation on the neuronal morphology of the PC12 cells. In the differentiated cells, M&B28767, an EP3 agonist, caused neurite retraction in a pertussis toxin-insensitive manner. 12-O-Tetradecanoylphorbol-13-acetate (TPA) also induced neurite retraction. However, when protein kinase C was down-regulated by long term exposure to TPA, TPA failed to induce neurite retraction, while the EP3B receptor-mediated retraction occurred normally. Clostridium botulinum C3 exoenzyme completely inhibited both EP3 agonist- and TPA-induced neurite retraction when microinjected into the cells, indicating that the morphological effect of the EP3B receptor is dependent on Rho activity. Thus, the activation of the EP3B receptor induced neurite retraction through a protein kinase C-independent Rho-activation pathway.
We studied the heterogeneity and intactness of the vif gene in six HIV-1-infected individuals at various clinical stages. The proviral vif sequences in peripheral blood mononuclear cells were amplified by PCR, followed by cloning and sequencing of 45 vif clones. The intraindividual diversity of the vif genes ranged from 0.45 to 3.3% and was not correlated with disease stage. Although the vif gene has been shown to be essential for infection of HIV-1 in vitro, a high frequency (31%) of defective vif genes was observed. In one patient, six vif clones carried double nonsense mutations at the same positions, five of which were clustered in the phylogenetic tree, suggesting that these vif-defective viruses may have replicated in vivo. Phylogenetic analysis revealed that the vif sequences from each individual were clustered into a separate group and that all of them belong to subtype B.