Biomedical subjects

Y Takeuchi

Publications and source records attributed to Y Takeuchi.

At least 235 records · Page 13Linked to original sources

PubMed1999

Green Fluorescent Protein As a Cell-Labeling Tool and a Reporter of Gene Expression in Transgenic Rainbow Trout.

: Green fluorescent protein (GFP) has been used as an indicator of transgene expression in living cells and organisms. For testing the utility of GFP in rainbow trout, we microinjected fertilized eggs with four types of supercoiled constructs containing two variants of GFP complementary DNA (S65T and EGFP), driven by two ubiquitous regulatory elements, human cytomegalovirus immediate early enhancer-promoter (CMV) and Xenopus laevis elongation factor 1alpha enhancer-promoter (EF1). Green fluorescence was first observed at 3 days postfertilization, when the embryo was in the mid-blastula stage. Fluorescence could be detected mosaically in various types of embryonic cells and tissues of swim-up fry. Both the percentage of fluorescent cells and the fluorescence intensity of GFP-expressing cells on blastoderms, measured with a microscopic photometry system, were highest in CMV-EGFP-microinjected embryos. We conclude that GFP is capable of producing detectable fluorescence in rainbow trout, and can be a powerful tool as a cell marker and reporter gene for cold-water fish, and that analysis of GFP expression in living cells is useful for characterizing the activity of cis-elements in vivo.

Journal Article↗

PubMed1999

Increase in histidine decarboxylase activity in tissues of mice bearing Colon-26 tumor cells.

The changes in histidine decarboxylase (HDC) activity, histamine and tele-methylhistamine contents were examined in tissues of mice after the inoculation of Colon-26 tumor cells subcutaneously into the lower back. The HDC activity in the spleen of mice increased significantly 14 days after the inoculation of Colon-26 and the increase in HDC activity continued for up to 28 days. However, the histamine content in the spleen of tumor-bearing mice was not changed significantly during the observation period. In the following experiments, two subclones of the Colon-26 cell line, cachexia-inducing clone-20 and non cachexia-inducing clone-5, were used and the induction of HDC activity in mice was examined in four tissues, spleen, lung, liver and kidney. Both clone-20 and clone-5 induced the increase in HDC activity to the same extent in the spleen and lung, but not in the liver and kidney. As observed using the Colon-26 original cell line, the histamine contents in the four tissues of tumor-bearing mice were not different from those in the control mice. In contrast, the levels of tele-methylhistamine, one of the major catabolites of histamine, in the tumor-bearing mice increased significantly compared with the control mice in all four tissues examined. There was a correlation between the increase in tele-methylhistamine level and the increase in HDC activity in the tissues. A histological study indicated that the tissue mast cells were not increased in spleen and lung of tumor-bearing mice. These findings indicated that the increase in HDC activity in the spleen and lung occurred in parallel with the growth of inoculated tumor cells in mice and suggested that the cells other than mast cells may be involved in the increase in HDC activity. The tumor-bearing state produced histamine with a high turnover rate in the mouse tissues, especially in the spleen and lung.

Adenocarcinoma↗

PubMed1999

Changes in serotonergic neurons in the brain of pyrithiamine-induced acute thiamine-deficient mice.

We examined changes in 5-hydroxytriptamine (5-HT, serotonin) neurons in pyrithiamine-induced thiamine deficiency in mice immunohistochemically. Extensive decreases in the densities of 5-HT-immunoreactive fibers were detected in the lateral septal nucleus, the thalamus, the medial mammillary nucleus, the dorsal and the median raphe nuclei, the raphe obscurus nucleus, the tegmental area, the cerebellum and the vestibular nucleus, though only a small decrease was detected in the inferior colliculus. Most remarkably, degenerative winding fibers were detected between the deep mesencephalic nucleus and the ventral tegmental area. Increases in intensity of 5-HT immunoreactivity in the dorsal raphe nucleus and decreases in the number of 5-HT-immunoreactive cell bodies in the dorsal and the median raphe nuclei were detected. These results demonstrated the differential vulnerability of 5-HT neurons in thiamine-deficient mice. This is the first report to demonstrate changes in 5-HT neurons immunohistochemically throughout the brain of pyrithiamine-induced thiamine deficient mouse.

Animals↗

PubMed1999

Results of surgical treatment of thymomas with special reference to the involved organs.

OBJECTIVE: The purpose of this study is to clarify the significance of the particular involved organ as a prognostic factor and its relation to other previously reported factors. METHODS: The prognoses of 194 consecutive patients with thymoma who had undergone complete or subtotal resection were reviewed retrospectively. Survival was evaluated as actuarial freedom from tumor death. Analysis of prognostic factors was performed by the Kaplan-Meier method with the log rank test and Cox's proportional hazards model. RESULTS: The Masaoka staging system and involvement of the great vessels were the independent prognostic factors in the entire study group; age, sex, histologic subtype, completeness of resection, association of myasthenia gravis, or involvement of other organs were not factors. The 10-year and 20-year survivals were 99% and 90% in stage I, 94% and 90% in stage II, 88% and 56% in stage III, 30% and 15% in stage IVa, 0% and 0% in stage IVb, 93% and 83% in the absence of involvement of the great vessels, and 54% and 20% in the presence of it. Involvement of the great vessels was also the single independent prognostic factor in the patients with stage III disease although completeness of resection or involvement of other organs were not. The 10-year and 20-year survivals in patients with stage III disease were 97% and 75% in the absence of involvement of the great vessels, and 70% and 29% in the presence of it. CONCLUSION: Although the Masaoka staging system is a valuable prognostic factor, the category of stage III is heterogeneous and consists of 2 groups with distinct prognoses depending on involvement of the great vessels.

Adolescent↗

PubMed1999

Limited uptake of foreign materials by resident macrophages in murine ovarian tissues.

In the present study, the distributions of exogenously administered horseradish peroxidase (HRP) and colloidal carbon in gonadal tissues were observed in both male and female mice using light microscopy. HRP was injected intravenously and colloidal carbon was directly injected into the gonadal parenchyma. Thereafter, the gonads were obtained for histological examination. The results showed that staining of HRP and carbon was detected in the ovaries at a low level. In contrast, much staining was observed in the interstitial cells of the testes for a long period. This suggests that ovarian tissues are less active in the uptake of exogenous materials than testicular tissues in vivo. However, immunohistochemical examination using anti-macrophage antibodies revealed that the ovaries contained a large number of macrophages, as did the testes, under normal conditions. Therefore, the results indicate that resident macrophages in the ovaries exhibit weak endocytic activity of foreign materials in vivo.

Animals↗

PubMed1999

Application of a computer based education system for aged persons and issues arising during the field test.

We plan to apply a computer-based multimedia text-book system for health education in the future. To do this, the computer system should be manageable to any person regardless of age or computer experience. We developed a multimedia text-book system and field-tested it to survey the reaction of aged persons. The system offers information about obesity prevention, such as a guide for eating and exercise. All contents are presented by text, image and voice. Also, a guide for exercise is shown by a movie. In this investigation we found that there were many considerations for aged persons using a computer. They seemed to have a strong aversion toward computers. They had the feeling that computers are difficult to use; actually 22 persons among 65 could not use it. Improvements in user interface seems to be the most important thing to make computers accessible to aged persons. On the other hand, question-and-answer programs for diagnosing one's health status were well accepted. Such a function is useful for attracting the interest of aged persons when we design a computer-based education system.

Aging↗

PubMed1999

Differential diagnosis of cervical lymph nodes in head and neck cancer by ultrasonography.

OBJECTIVE: Determination of whether an enlarged cervical lymph node is metastatic or not is clinically important in head and neck oncology. Differential diagnosis of the lymph node, however, is still a diagnostic problem. The purpose of this study is to clarify the ultrasonographic findings of the metastatic lymph nodes of head and neck squamous cell carcinoma and to establish the criteria. METHODS: We investigated 36 metastatic lymph nodes in head and neck squamous cell carcinoma and 24 non-metastatic nodes in benign disease with a 10-MHz transducer. We examined the size, shape, and internal echo (echo level, punctate bright echogenic spots, hilus echogenic line, cystic pattern) of these nodes. Based on this investigation, we evaluated 70 lymph nodes from 25 other patients by ultrasonography. RESULTS: The short axis diameter and shape of metastatic nodes were larger and rounder than those of non-metastatic ones. Of the metastatic nodes, 69% showed hypoechoic and 31% isoechoic levels, and 78% exhibited punctate bright echogenic spots. Of the non-metastatic nodes, 92% showed hypoechoic and 8% isoechoic levels, and none of them showed the spots. The hilus echogenic line was not present in any metastatic node, but it was seen in 58% of non-metastatic ones. Of the metastatic nodes, 19% exhibited a cystic pattern; none of the non-metastatic nodes showed the pattern. According to our criteria based on these results, the accuracy rate was 98.6% (69/70). The sensitivity and specificity were 97.2% (35/36) and 100% (34/34), respectively. The false positive rate and the false negative rate were 0% (0/70) and 1.4% (1/70), respectively. CONCLUSION: Internal echo findings and shape of lymph nodes can be an important diagnostic tool, and our ultrasonographical criteria of the lymph nodes are very useful for the differential diagnosis of the cervical lymph nodes.

Biopsy↗

PubMed1999

Outbreak of severe neurologic involvement associated with Enterovirus 71 infection.

Enterovirus 71 has been associated with several outbreaks, as well as sporadic cases, of central nervous system infection and has a worldwide distribution. Seven children with encephalitis and five with aseptic meningitis caused by Enterovirus 71 were seen at Otsu Municipal Hospital during the summer of 1997. The infections were confirmed serologically, although detection of the viral genome in cerebrospinal fluid was unsuccessful. Seven children were diagnosed as having hand-foot-and-mouth syndrome, two were diagnosed as having herpangina, and three patients younger than 12 months old developed no eruptions. The skin or mucosal manifestations of this outbreak demonstrated considerable variation. The Enterovirus 71 strain that caused the outbreak had a strong neurovirulent tendency. Among the patients with encephalitis, symptoms originating from the impairment of diencephalon were seen in four patients, and those originating from cerebellar impairment were seen in two patients. Brain magnetic resonance imaging in one patient revealed an abnormality in the pons. The neurologic manifestations associated with Enterovirus 71 infection may be characterized by involvement of the cerebellum, brainstem, and diencephalon. Enterovirus 71 is one of the pathogenic viruses that cause hand-foot-and-mouth syndrome, as well as a variety of other clinical manifestations. The most important of these is neurologic disease, especially in infants and young children.

Antibodies, Viral↗

PubMed1999

Influences of keratinocyte-fibroblast interaction on the expression of epimorphin by fibroblasts in vitro.

Epimorphin was demonstrated to be a mesenchymal signal factor modulating epithelial morphogenesis of skin, lung and liver in vitro. Most of the previous studies were performed biochemically and functionally. In the present study, expression of epimorphin was immunohistochemically compared between cultured fibroblasts and cocultured fibroblasts with keratinocytes obtained from normal skin. Cultured fibroblasts revealed a low level of epimorphin expression. In contrast, the expression by fibroblasts was greatly enhanced in skin explant culture where both fibroblasts and keratinocytes were present. In three-dimensional coculture of fibroblasts and keratinocytes, the expression of epimorphin was enhanced. The staining pattern of epimorphin in three-dimensional coculture was similar to that in human skin. These results suggest that dermal fibroblasts are manufacturers of epimorphin, and keratinocyte-fibroblast interaction may play important roles in the expression of epimorphin in vitro.

Adult↗

PubMed1999

Diagnostic utility of magnetic resonance imaging skeletal survey in a patient with oncogenic osteomalacia.

Oncogenic osteomalacia is a rare paraneoplastic syndrome characterized by hypophosphatemic osteomalacia due to renal phosphate wasting. The same biochemical features are found in patients with X-linked hypophosphatemic rickets/osteomalacia and sporadic hypophosphatemic osteomalacia with unknown etiology. Oncogenic osteomalacia is cured by resection of the responsible tumor. In contrast, patients with other types of hypophosphatemic rickets/osteomalacia need long-term treatment with large doses of active vitamin D3. Therefore, detection of the responsible tumor for oncogenic osteomalacia has great clinical importance. However, there is no standard method for detecting the tumor for oncogenic osteomalacia, and the responsible tumor is often very difficult to be found. We describe a patient with adult-onset osteomalacia due to renal phosphate wasting. Although oncogenic osteomalacia was suspected, cranial, chest, and abdominal computed tomography scanning, urological and otolaryngological examinations, and detailed palpation for soft tissue mass failed to detect the responsible tumor. However, magnetic resonance imaging skeletal survey revealed a tumor in the right femoral bone. Resection of the tumor resulted in normalization of serum phosphate and renal phosphate handling. Because the most frequent causes for oncogenic osteomalacia are tumors in bone or soft tissue, magnetic resonance imaging skeletal survey is a very powerful method for detecting the responsible tumor. Vigorous search for tumors with this method in patients with hypophosphatemic osteomalacia would be helpful not only for proper management of patients, but also for clarifying the identity of sporadic hypophosphatemic osteomalacia.

Bone Neoplasms↗

PubMed1999

Induction of complement attack on human cells by Gal(alpha1,3)Gal xenoantigen expression as a gene therapy approach to cancer.

Galactose(alpha1,3)galactose on the surface of cells of non-primate organs is the major xenoantigen responsible for hyperacute rejection in xenotransplantation. The antigen is synthesised by (alpha1, 3)galactosyl transferase. Humans lack this enzyme and their serum contains high levels of pre-existing natural antibody which recognises the structure and activates complement. We have evaluated in vitro the potential for delivery of this enzyme to sensitise human cells to complement attack as a gene therapy approach to cancer. Retrovirus-mediated delivery of (alpha1,3)galactosyl transferase resulted in high level expression which led to serum-mediated lysis of five human cell targets, including endothelial and primary melanoma cells. Lysis was specific for those cells expressing the antigen in a mixed cell population. The mechanism of cell lysis mimicked that involved in hyperacute rejection: activation of the classical complement pathway by natural antibody specific for galactose(alpha1,3)galactose. The degree of lysis was determined by both the level of specific antibody and the expression of glycophosphatidylinositol-linked complement regulatory proteins. We conclude that expression of (alpha1,3)galactosyl transferase is a promising new therapeutic approach for cancer gene therapy, avoiding toxicity problems associated with application of prodrugs and with the potential to elicit further immunological responses.

Antigens, Heterophile↗

PubMed1999

An adult patient with severe hypercalcaemia and hypocalciuria due to a novel homozygous inactivating mutation of calcium-sensing receptor.

Inactivating mutations in the calcium-sensing receptor (CaSR) cause familial hypocalciuric hypercalcaemia (FHH) and neonatal severe hyperparathyroidism (NSHPT). Earlier investigations showed patients with FHH are heterozygous, and NSHPT are homozygous for inactivating mutations. However, one adult patient with severe hypercalcaemia and hypocalciuria has been reported to have a homozygous inactivating mutation in CaSR (Pro39Ala). This suggested that mutant CaSR in this patient had some residual activity and hypercalcaemia was not so severe as to be fatal. However, the function of this mutant CaSR was not evaluated. In the present study, we describe a novel homozygous mutation in an adult patient with severe hypercalcaemia and hypocalciuria, and evaluate the function of the mutant CaSRs. The DNA sequence of CaSR gene was determined by direct sequencing of the polymerase chain reaction product. The function of mutant CaSR was analysed by creating mutant cDNAs by in vitro mutagenesis, transfection of mutant cDNAs into HEK293 cells and measuring intracellular ionized Ca in response to changes in extracellular Ca. A 26-year-old Japanese woman showed marked hypercalcaemia with an elevated parathyroid hormone (PTH) level. Her consanguineous parents had asymptomatic hypercalcaemia with relative hypocalciuria. The proband had a homozygous mutation at codon 27 of CaSR gene (CAA-->CGA, Gln27Arg). Her parents were heterozygous for this mutation. EC50 for Ca of this mutant CaSR (GIn27Arg) was 4.9 mM. EC50 of another mutant CaSR (Pro39Ala) whose homozygous mutation was discovered in an adult patient was 4.4 mM. These EC50s were significantly higher than that of wild-type CaSR (3.7} 0.1 mM), but were the lowest among the reported EC50s for inactivating mutations of CaSR. These results indicate that serum Ca and PTH levels are determined by residual function of mutant CaSR in patients with homozygous mutation in CaSR, and that patients having homozygous mutant CaSRs with mild dysfunction do not suffer from fatal hypercalcaemia in infancy and can survive into adulthood.

Adult↗

PubMed1999

Degeneration of the seminiferous epithelium with ageing is a cause of spermatoceles?

A spermatocele refers to the cystic accumulation of semen in the male reproductive tract. Although it is thought to be caused by narrowing of the lumen of the excurrent duct with resultant cystic dilatation of the duct, the pathogenesis of the narrowing remains unknown. In the present study, we histologically examined spontaneous spermatoceles in C3H/He mice to elucidate the pathogenesis of the lesions. Testes, efferent ducts, epididymides and vas deferens obtained from young and aged C3H/He mice were embedded in plastic for histological observation at the light microscopic level. It was found that spontaneous spermatoceles were localized in the rete testis and efferent ducts of aged mice, as seen in man. The dilated rete testis and efferent ducts contained many degenerated and aggregated germ cells derived from the exfoliated seminiferous epithelium in the aged testis. In particular, it was noted that the agglutinated germ cells obstructed the narrow lumen of the efferent ducts, resulting in the failure of transport of germ cells to the caput epididymis, and spermatoceles were consistently found in the region between the rete testis and the obstructed site in the efferent ducts. However, no inflammatory cell infiltration, traumatic injury or spermatic granulomas were found in the occluded region. These results suggest that agglutinated germ cells may occupy the narrow lumen of the efferent ducts, resulting in the formation of a spermatocele. It may be that a senile change to the seminiferous epithelium, which releases immature germ cells into the lumen of the seminiferous tubules, is the cause of this type of spermatocele.

Aging↗

PubMed1999

Major differences between the testis and epididymis in the induction of granulomas in response to extravasated germ cells. I. A light microscopical study in mice.

A spermatic granuloma is a chronic inflammatory lesion which surrounds extravasated spermatozoa. Clinically, the lesion develops in the interstitial spaces of the epididymis and vas deferens, and only exceptionally in the testis itself. In the present study, murine testes and epididymides were injured using a needle and the histological appearances of these organs was then compared. Traumatic injury induced extravasation of germ cells in both testes and epididymides. A few days later, spermatic granulomas consistently formed in the epididymides, however, such lesions were not induced in the testes. To examine the possibility that epididymal spermatozoa have inherently greater ability to form spermatic granulomas than do testicular germ cells, isolated epididymal spermatozoa or testicular germ cells were locally injected into the testes and epididymides of recipient mice. Spermatic granulomas readily formed in the epididymides after local injection of either epididymal spermatozoa or testicular germ cells. In contrast, such lesions did not form in the testes even when epididymal spermatozoa were injected. Therefore, this study suggests that the microenvironment of the testicular interstitium, rather than the extravasated components from the ruptured seminiferous tubules, is the main factor determining the limited formation of spermatic granulomas in the testis.

Animals↗

PubMed1999

A 42-kDa glycoprotein from chicken egg-envelope, an avian homolog of the ZPC family glycoproteins in mammalian Zona pellucida. Its first identification, cDNA cloning and granulosa cell-specific expression.

A glycoprotein with molecular mass of 42 kDa was identified as the major component of the chicken egg-envelope, the filamentous, extracellular matrix known as the perivitelline layer. By using a DNA probe amplified with degenerative primers derived from the protein's partial amino acid sequences, a cDNA clone encoding the egg-envelope 42-kDa glycoprotein (gp42) was isolated from a hen's ovary cDNA library. The gp42 open reading frame encoded 435 amino acid residues, including a putative signal peptide of 20 amino acids. The deduced amino acid sequence of gp42 showed significant similarity to egg-envelope glycoproteins of the ZPC family of several other vertebrate species, including human ZP3, mouse ZP3, Xenopus laevis gp43 and medaka (Oryzias latipes) ZI3 (LS-F), which play important roles for sperm-egg interaction. A single N-glycosylation site present in chicken gp42 is conserved among all five of these proteins: carbohydrate analysis of gp42 revealed the presence of a complex type glycan chain at this site. N-terminal sequence analysis of the mature polypeptide suggests that C-terminal processing of the pro-protein occurs during synthesis and secretion. The 1.4-kb gp42 transcript was detected only in follicles, and was found to be accumulated in granulosa cells in a manner dependent on ovarian follicular development. Furthermore, a metabolically radio-labeled gp42 was immunopreciptated from both cell lysate and culture supernatant of the granulosa cells with specific anti-gp42 antibody, suggesting granulosa cell-specific synthesis and secretion of the glycoprotein.

Amino Acid Sequence↗

PubMed1999

The effect of the timing of ethanol exposure during early postnatal life on total number of Purkinje cells in rat cerebellum.

We have previously shown that exposing rats to a high dose of ethanol on postnatal d 5 can affect Purkinje cell numbers in the cerebellum whilst similar exposure on d 10 had no such effect. The question arose whether a longer period of ethanol exposure after d 10 could produce loss of Purkinje cells. We have examined this question by exposing young rats to a relatively high dose (approximately 420-430 mg/dl) of ethanol for 6 d periods between the ages of either 4 and 9 d or 10 and 15 d of age. Exposure was carried out by placing the rats in an ethanol vapour chamber for 3 h per day during the exposure period. Groups of ethanol-treated (ET), separation controls (SC) and mother-reared controls (MRC) were anaesthetised and killed when aged 30 d by perfusion with buffered 2.5% glutaraldehyde. Stereological methods were used to determine the numbers of Purkinje cells in the cerebellum of each rat. MRC, SC and rats treated with ethanol between 10-15 d of age each had, on average, about 254-258 thousand cerebellar Purkinje cells; the differences between these various groups were not statistically significant. However, the rats treated with ethanol vapour between 4-9 d of age had an average of only about 128000+/-20000 Purkinje cells per cerebellum. This value was significantly different from both the MRC and group-matched SC animals. It is concluded that the period between 4 and 9 d of age is an extremely vulnerable period during which the rat cerebellar Purkinje cells are particularly susceptible to the effects of a high dose of ethanol. However, a similar level and duration of ethanol exposure commencing after 10 d of age has no significant effect on Purkinje cell numbers.

Animals↗

PubMed1999

Nuclear localization of the delta subunit of Ca2+/calmodulin-dependent protein kinase II in rat cerebellar granule cells.

To examine the physiological roles of the delta subunit of Ca2+/calmodulin-dependent protein kinase II (CaM kinase IIdelta) in brain, we examined the localization of CaM kinase IIdelta in the rat brain. A specific antibody to CaM kinase IIdelta1-delta4 isoforms was prepared by immunizing rabbits with a synthesized peptide corresponding to the unique carboxyl-terminal end of these isoforms. The prepared antibody did not recognize the alpha, beta, and gamma subunits, which were each overexpressed in NG108-15 cells. Immunoblot analysis on various regions and the nuclear fractions from rat brains suggested that some isoforms of CaM kinase IIdelta1-delta4 were abundant in the nucleus in the cerebellum. Total RNA from the cerebellum was analyzed by RT-PCR with a primer pair from variable domain 1 to variable domain 2. We detected the three PCR products delta3.1, delta3.4, and delta3 that contained the nuclear localization signal. These CaM kinase IIdelta3 isoforms were localized in the nuclei in transfected NG108-15 cells. Immunohistochemical study suggested the existence of these isoforms in the nuclei in cerebellar granule cells. These results suggest that CaM kinase IIdelta3 isoforms are involved in nuclear Ca2+ signaling in cerebellar granule cells.

Amino Acid Sequence↗

PubMed1999

The prolactin gene is expressed in the mouse kidney.

BACKGROUND: Prolactin (PRL), originally identified as an anterior pituitary hormone exhibiting lactogenic activity, is now recognized as a versatile hormone expressed in a wide variety of tissues. METHODS: In this study, the expression of PRL in the mouse kidney was investigated by solution-phase and in situ reverse transcription-polymerase chain reaction (RT-PCR) methods and immunohistochemistry. RESULTS: Mouse PRL (mPRL) transcript and protein are localized in the parietal epithelial cells of Bowman's capsule. Pit-1 is a positive transcription factor for the expression of the PRL gene. The presence of Pit-1 transcript in the kidney was also assessed by RT-PCR methods. The localization of Pit-1 mRNA coincided well with that of PRL. Immunoreactivity to mouse PRL receptor (mPRL-R) is distributed on the luminal membrane of the proximal tubule cells and the parietal epithelial cells of Bowman's capsule. CONCLUSION: These data indicate that the parietal epithelial cells of Bowman's capsule synthesize PRL de novo and suggest that Pit-1 contributes to the transcriptional regulation of PRL gene expression in the kidney, and PRL expressed in this tissue functions in an autocrine/paracrine fashion.

Animals↗