Biomedical subjects
Publications and source records attributed to K Kikuchi.
We report a Japanese patient with Hermansky-Pudlak syndrome (HPS) who developed systemic lupus erythematosus (SLE). This is the second case report of HPS complicated with SLE. A 1-bp duplication of adenine at codon 441 was found in the HPS gene, namely HPS1, which caused a frameshift. This case serves as evidence indicating that a patient with HPS can be predisposed to SLE.
Matrix metalloproteinase-9 (MMP-9) and its inhibitor, tissue inhibitors of metalloproteinases-1 (TIMP-1) are involved in tissue inflammation and fibrotic processes. We previously reported an elevated serum TIMP-1 level in patients with systemic sclerosis (SSc). We measured serum TIMP-1 and active MMP-9 levels in 62 patients with SSc, 10 patients with rheumatoid arthritis (RA) and 15 normal controls using a modified enzyme-linked immunosorbent assay. The active MMP-9 level in the serum of the patients with RA or SSc was not significantly different from that of controls. Serum MMP-9 activity in patients with diffuse cutaneous SSc was significantly decreased compared with that of limited cutaneous SSc or normal controls. The MMP-9 activity and modified-Rodnan total skin thickness score in patients with SSc were negatively correlated. Serum MMP-9 activity and TIMP-1 level in patients with SSc were not correlated. Serum MMP-9 activity might be a useful indicator of disease activity, especially skin severity, in SSc patients.
Influenza patients show a high incidence of T lymphocytopenia in the acute phase of the illness. Since CD8+ T cells play an important role in influenza virus infection, we investigated which subset of CD8+ T cells was involved in this lymphocytopenia. CD8+ T cells from eight patients with influenza A were studied for lymphocyte count, surface marker, and intracellular IFN-gamma production in the acute (days 1-3) and recovery phases (days 9-12). Total and T lymphocyte counts in the acute phase were approximately three times less than in the recovery phase; however, the CD4/8 ratio was the same in both phases. The cell count reduction in the acute phase was attributed predominantly to the CD28+ CD8+ subset, compared with the CD28- CD8+ subset. The memory/activation marker CD45RO on the CD8+ T cells was assessed. The CD28+ CD45RO- subset, a naive phenotype, was reduced significantly in number in the acute phase compared with the recovery phase. The CD28+ CD45RO+ subset, a memory phenotype, was also reduced in the acute phase, but the reduction was not statistically significant. Intracellular IFN-gamma in the CD8+ subset after mitogenic stimulation was measured by flow cytometry; the percentage of CD28+ IFN-gamma-/CD8+ subset in the acute phase was significantly less than in the recovery phase. These results indicated that the predominant reduction of peripheral CD8+ T cells in the acute phase of influenza was from naive-type lymphocytes, suggesting that these quantitative and qualitative changes of CD8+ T cells in influenza are important for understanding the immunological pathogenesis.
It has been clearly shown that autoimmune diseases can be treated by apheresis by eliminating immune complexes, however, the effects of therapeutic apheresis are not limited to immune disorders. Almost all diseases are associated with immune systems. Immune systems can be regulated by advanced techniques of apheresis, including immunoadsorption and immunocytapheresis, removing immune effector molecules and various immune-associated cells selectively. Therefore, apheresis can be used as a nondrug treatment for many diseases. In addition, disease-associated proteins that cause disease or are produced in the course of diseases and accumulate in the body could be eliminated selectively by apheresis using the extremely powerful ability of the immune system to recognize polypeptide structures specifically and distinguish miniscale differences among molecules. In this article, we discuss the current status of treatment of immune diseases by apheresis and possible treatment approach of a variety of diseases by apheresis based on immune reactions.
The content and bioavailability of selenium (Se) in the dark muscle of tuna (DMT) were studied. Fluorometric analysis showed that DMT contained 2.0-4.7 microg x g(-1) Se. A large part of Se of the DMT was recovered in the dry powder of the defatted fraction prepared by successive treatment with acetone and n-hexane/ethanol. On trypsin digestion of the defatted DMT, release of Se paralleled that of nitrogen and about 70% of Se was released within 4 h. Male weanling ddY mice were fed a Torula yeast-based Se-deficient diet (basal diet) for 3 weeks, and then fed the basal diet or a diet supplemented with a 0.05-0.25 microg x g(-1) Se as either sodium selenite or the defatted DMT for a further 1 week. Se contents and GSHPx activities in the liver increased gradually with increases in the amount of supplemented Se. No differences were observed between selenite and defatted DMT in the effect on Se content. At low Se levels, the effect of Se in the defatted DMT on the liver GSHPx activities was inferior to that of selenite, but at a high Se level, Se in the defatted DMT showed a greater effect. The bioavailability of Se in the defatted DMT, as assessed by slope ratio analysis using selenite as the reference Se, was 87% for the liver Se content and 168% for the GSHPx activity. The results indicate that the defatted DMT contains high levels of Se in a nutritionally available form.
We used immunoneutralization of endogenous estradiol to investigate deficiencies in the estradiol-feedback regulation of LH secretion as a primary cause of follicular cysts in cattle. Twenty-one cows in the prostaglandin (PG) F(2alpha)-induced follicular phase were assigned to receive either 100 ml of estradiol antiserum produced in a castrated male goat (n = 11, immunized group) or the same amount of castrated male goat serum (n = 10, control group). The time of injection of the sera was designated as 0 h and Day 0. Five cows in each group were assigned to subgroups in which we determined the effects of estradiol immunization on LH secretion and follicular growth during the periovulatory period. The remaining six estradiol-immunized cows were subjected to long-term analyses of follicular growth and hormonal profiles, including evaluation of pulsatile secretion of LH. The remaining five control cows were used to determine pulsatile secretion of LH on Day 0 (follicular phase) and Day 14 (midluteal phase). The control cows exhibited a preovulatory LH surge within 48 h after injection of the control serum, followed by ovulation of the dominant follicle that had developed during the PGF(2alpha)-induced follicular phase. In contrast, the LH surge was not detected after treatment with estradiol antiserum. None of the 11 estradiol-immunized cows had ovulation of the dominant follicle, which had emerged before estradiol immunization and enlarged to more than 20 mm in diameter by Day 10. Long-term observation of the six immunized cows revealed that five had multiple follicular waves, with maximum follicular sizes of 20-45 mm at 10- to 30-day intervals for more than 50 days. The sixth cow experienced twin ovulations of the initial persistent follicles on Day 18. The LH pulse frequency in the five immunized cows that showed the long-term turnover of cystic follicles ranged from 0.81 +/- 0.13 to 0.97 +/- 0.09 pulses/h during the experiment, significantly (P < 0.05) higher than that in the midluteal phase of the control cows (0.23 +/- 0.07). The mean LH concentration in the immunized cows was also generally higher than that in the luteal phase of the control cows. However, the LH pulse and mean concentration of LH after immunization were similar to those in the follicular phase of the control cows. Plasma concentrations of total inhibin increased (P < 0.01) concomitant with the emergence of cystic follicles and remained high during the growth of cystic follicles, whereas FSH concentrations were inversely correlated with total inhibin concentrations. In conclusion, neutralization of endogenous estradiol resulted in suppression of the preovulatory LH surge but a normal range of basal LH secretion, and this circumstance led to an anovulatory situation similar to that observed with naturally occurring follicular cysts. These findings provide evidence that lack of LH surge because of dysfunction in the positive-feedback regulation of LH secretion by estradiol can be the initial factor inducing formation of follicular cysts.
The aims of this study were to develop a sensitive and specific assay for bovine inhibin A using europium and to investigate the endocrine role of inhibin A in various reproductive conditions by characterizing the relationship between profiles of inhibin A, FSH, and estradiol and follicle growth during the postpartum period, during the intact estrous cycle, and in cows with follicular cysts. The time-resolved immunofluorometric assay (Tr-IFMA) for bovine inhibin A, using purified polyclonal antibodies to alpha and beta(A) subunits, was specific for bovine inhibin A and did not cross-react with bovine activin A, activin AB, activin B, pro-alphaC or human recombinant inhibin B. The detection limit of the IFMA was 3.3 pg/ml expressed in terms of bovine 32-kDa inhibin A. Dose-response curves of plasma samples obtained from intact and FSH-stimulated cows and cystic cows were parallel to the standard without any preassay processing of samples. Plasma inhibin A levels increased (P < 0.01) concomitant with emergence of nonovulatory or ovulatory follicular waves during the postpartum period. In cystic cows, plasma inhibin A was sustained at high levels for a longer period, associated with growth of persistent dominant follicles. The highest levels of inhibin A were noted during the growth phase of normal and persistent dominant follicles; however, inhibin A levels declined (P < 0.01) as these dominant follicles ceased to grow or ovulated. An inverse relationship between patterns of plasma inhibin A and FSH existed during each follicular wave in the three physiologic conditions. Increases in plasma inhibin A levels were associated with increases in plasma estradiol levels during most follicular waves; however, there was no increase in plasma estradiol level and no relationship between patterns of estradiol and FSH during follicular waves observed during the early postpartum period or midluteal phase of the estrous cycle. In conclusion, the Tr-IFMA does not require pretreatment of samples and can be used for precise measurement of bovine inhibin A without interference with free inhibin alpha subunits. Inhibin A, produced primarily during growth of the dominant follicle, functions as a negative feedback regulator for FSH secretion throughout the postpartum period and the estrous cycle, whereas estradiol appears to have a minor role in regulation of FSH compared with inhibin A, especially during the early postpartum period and midluteal phase of the estrous cycle. The results also indicate that a persistent dominant follicle sustains inhibin A production for a longer period than the dominant follicle emerging in the estrous cycle and establishes long-term dominance by suppressing emergence of a new follicular wave.
A 65-yr-old female developed cough, fever and dyspnoea following repeated exposure to a home ultrasonic humidifier. High-resolution computed tomography showed ground-glass opacity in both lung fields. Arterial blood gas analysis gave an oxygen tension of 8.38 kPa (63 Torr). Pulmonary function testing revealed restrictive ventilatory impairment with a reduction in the diffusing capacity. The diagnosis of extrinsic allergic alveolitis (EAA) was confirmed by radiographic findings, pathological evidence of alveolitis and reproductive development by a provocation test to the humidifier water. The yeast Debaryomyces Hansenii was the only microorganism cultured from the water of the humidifier. The double diffusion precipitating test and lymphocyte proliferative response was positive for an extract of D. Hansenii, providing evidence to incriminate this fungus. This is the first described case of EAA caused by D. Hansenii.
Sperm-specific nuclear protamines are dissociated before decondensation of sperm nuclei during fertilization in pigs. In the present study, replacement of nuclear protein by histone in boar spermatozoa during in vitro fertilization was evaluated by immunohistochemistry using anti-histone antibody. First, the specificity of the antibody used in this study was examined. Immunohistochemistry of the testes and epididymides indicated that somatic nuclei, but not elongated spermatids or maturing spermatozoa, were immunoreactive. Furthermore, immunoreaction was diminished after the antibody had been preincubated with unfractionated histone, indicating that the antibody was specific for the somatic nuclear histone. Immunohistochemistry of serial sections of oocytes, which were matured and co-cultured with boar spermatozoa for 2 to 6 h indicated that, at 2 to 3 h after insemination, penetrating sperm nuclei in the condensed state were not immunoreactive. At 4 to 5 h after insemination, some of the condensed sperm nuclei were immunoreactive in part or over the whole area of the nucleus, and all of the decondensing nuclei and male pronuclei were immunoreactive. At 6 h after insemination, the decondensing sperm nuclei and well-developed male pronuclei were immunoreactive. These results imply that, in pigs, remodelling of sperm nuclear protein from protamine to histone is initiated at the time of sperm penetration, before onset of decondensation and male pronuclear formation.
Structure-based in-silico screening was carried out for the Syk C-terminal SH2 domain. Fragments that could interact with the pY or pY+1 pockets were selected by our in-silico screening. After tethering two fragments bound to these pockets, we have designed and synthesized new compounds that show favorable interaction with the pY+3 pocket. One such compound, having a cyclohexylmalonic acid moiety identified as a novel potent phosphotyrosyl mimetic, exhibited an affinity comparable to that of the monophosphorylated ligand peptide.
Cortical silent period (SP) of the limb muscles is thought to reflect the cortical excitability. However, the lingual SP has not been examined precisely even in normal subjects. We investigated SP in the tongue induced by transcranial magnetic stimulation (TMS) in 18 controls. Surface electrodes were placed on the lingual dorsum using a bipolar technique. A round coil (13.5 cm in outer diameter) connected with Magstim 200 stimulator was placed on the motor cortex of the tongue, and the intensity of the stimulation was increased stepwise to maximum. SP was detected in all subjects especially at the contralateral side to the stimulated side, without contamination of peripheral SP. The duration of SP depended on the stimulus intensity, while the degrees of muscle contraction did not influence SP. SP of the tongue showed similar characteristics to that of limb muscles. This suggests that SP of lingual muscles can be clinically useful for the evaluation of corticobulbar excitability.
Protein tyrosine phosphatase (PTP) epsilon (PTPepsilon) exists as 2 forms generated by alternative promoter usage. It has recently been reported that a cytosolic isoform of PTPepsilon (PTPepsilonC) when over-expressed in murine M1 myeloid cells inhibits interleukin-6 (IL-6)- and leukemia inhibitory factor-induced activation of Janus kinases (JAKs), thereby suppressing STAT3 tyrosine phosphorylation and STAT3 signaling. This study characterizes an inhibitory action of PTPepsilonC on IL-6 signaling and also reveals that PTPepsilonC inhibitory activity is independent of other potential negative regulators, such as SHP-2 and SOCS family proteins. Furthermore, it analyzes the selectivity of PTPepsilonC action toward several cytokines. On IL-6 stimulation, expression of PTPepsilonC-DA, a catalytically inactive mutant of PTPepsilonC, results in an earlier onset of STAT3 tyrosine phosphorylation, suggesting different modes of action between PTPepsilonC and other negative regulators. In addition, the study shows PTPepsilonC-DA enhances activation of STAT1 by IL-6 as well. In terms of specificity to cytokines, over-expressed PTPepsilonC also inhibits IL-10-induced tyrosine phosphorylation of STAT3 in M1 cells, whereas PTPepsilonC does not affect either interferon-beta- and interferon-gamma-induced tyrosine phosphorylation of STATs or expression of STAT transcriptional targets. Among cytokines tested, the inhibitory effect of PTPepsilonC is selective to IL-6- and IL-10-induced JAK-STAT signaling.
We investigated the role of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 in the growth regulation. Primary (PM-WK and KHm-4), recurrent primary (RPM-EP and RPM-MC), lymph node metastatic (MM-AN, MM-BP and MM-RU), and a visceral metastatic (MM-LH) melanoma cell lines were used. Reverse transcriptase-coupled polymerase chain reaction and Western blotting revealed that all expressed and produced TIMP-1 and TIMP-2 except for PM-WK, which neither expressed nor produced TIMP-1. TIMP-1 and TIMP-2 secretion levels were measured by enzyme-linked immunosorbent assay in supernatants of cells. We found that the TIMP-1 production level was correlated with the cell migration rate. Moreover, TIMP-1 enhanced the cell migration of PM-WK. The growth of the primary melanoma cell lines was stimulated by TIMP-1 and inhibited by TIMP-2. In contrast, the growth of the visceral metastatic melanoma cell line was stimulated by TIMP-2.
Maspin belongs to a tumor suppressing protein which is usually expressed highly in myoepithelial cells, and is significantly downregulated in breast cancer cells. We focused on identifying the correlation between maspin expression and the clinicopathological features of human breast cancer tissues using immunohistochemistry. There was a significant correlation in that maspin-positive tumor specimens showed a low pathological grade of malignancy, such as a lower infiltration of the tumor into the surrounding tissue and a downregulation of c-erbB2 expression. Moreover, maspin-positive cases showed a significant decrease in tumor vessels positively stained with anti-factor VIII-related antigen antibody. These results suggest that maspin production in myoepithelial cells could downregulate the malignant phenotype of breast cancer.
Here we isolated tautomycetin, TC, and examined its phosphatase inhibitory activity. Recently we have reported that the left-hand moiety of tautomycin, TM, and the right one containing the spiroketal are essentially required for inhibition of protein phosphatase, PP, and induction of apoptosis, respectively. TC is structurally almost identical to TM except that TC is lacking the spiroketal, which has the potential apoptosis-inducing activity. TC specifically inhibited PP1 activity, IC50 values for purified PP1 and PP2A enzymes being 1.6 and 62 nM, respectively, whereas the IC50 values of TM were 0.21 and 0.94 nM, respectively. These results demonstrate that TC is the most specific PP1 inhibitor out of over 40 species of natural phosphatase inhibitors reported, strongly suggesting that TC is a novel powerful tool to elucidate the physiological roles of PP1 in various biological events.
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Mitogen-activated protein kinase (MAPK) phosphatases (MKPs) negatively regulate MAPK activity. In the present study, we have identified a novel MKP, designated MKP-7, and mapped it to human chromosome 12p12. MKP-7 possesses a long C-terminal stretch containing both a nuclear export signal and a nuclear localization signal, in addition to the rhodanese-like domain and the dual specificity phosphatase catalytic domain, both of which are conserved among MKP family members. When expressed in mammalian cells MKP-7 protein was localized exclusively in the cytoplasm, but this localization became exclusively nuclear following leptomycin B treatment or introduction of a mutation in the nuclear export signal. These findings indicate that MKP-7 is the first identified leptomycin B-sensitive shuttle MKP. Forced expression of MKP-7 suppressed activation of MAPKs in COS-7 cells in the order of selectivity, JNK p38 > ERK. Furthermore, a mutant form MKP-7 functioned as a dominant negative particularly against the dephosphorylation of JNK, suggesting that MKP-7 works as a JNK-specific phosphatase in vivo. Co-immunoprecipitation experiments and histological analysis suggested that MKP-7 determines the localization of MAPKs in the cytoplasm.
BACKGROUND: Cytokines induce apoptosis in vascular disease lesions through enhancement of inducible nitric oxide (NO) synthase (iNOS) activation. The thiazolidinediones, novel insulin-sensitizing agents, have been demonstrated to modulate cytokine-induced NO production. We have investigated the role of pioglitazone in the apoptosis of vascular smooth muscle cells (VSMCs) in vitro and developed intimal hyperplasia in vivo. METHODS AND RESULTS: Pioglitazone (0.1 to 10 micromol/L) significantly enhanced cytokine-induced expression of iNOS and NO production in a dose-dependent manner in rat VSMCs, but 15-deoxy-Delta(12,14)-prostaglandin J2 (up to 10 micromol/L), a native peroxisome proliferator-activated receptor-gamma ligand, showed no effect. Pioglitazone also significantly enhanced reduction of cell viability, as evidenced by the increase in the number of TUNEL-positive cells. All of these effects of pioglitazone were blocked by treatment with N-monomethyl-L-arginine, an NO synthesis inhibitor. In an in vivo study with a balloon-injured rat carotid artery, neointimal thickness had reached maximum levels at 2 weeks after injury. Then, rats were fed with or without pioglitazone (3 mg. kg(-1). d(-1)) for an additional week. The ratio of intima to media area of carotid artery was significantly decreased by 30%, and the ratio of apoptotic cells in neointima was significantly increased in pioglitazone-treated rats compared with vehicle-treated control rats. CONCLUSIONS: Pioglitazone enhanced apoptosis in an NO-dependent manner in cytokine-activated VSMCs and induced significant regression of intimal hyperplasia in balloon-injured rat carotid artery. It appears that pioglitazone is a potent apoptosis inducer in vascular lesions, providing a novel pharmacological strategy to prevent restenosis after vascular intervention.