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Biomedical subjects

E Murata

Publications and source records attributed to E Murata.

At least 37 records · Page 2Linked to original sources
PubMed1997

Formation of new capillary-like tubes in a three-dimensional in vitro model (aorta/collagen gel).

Direct sprouting (angiogenesis) does not occur during the formation of capillary-like tubes in an aorta/ collagen gel in the in vitro model. However, emigration of cells which stretch, arrange themselves side by side, form contacts (unspecific, tight and gap junctions), develop a lumen and show differentiation of endothelial cells (including the formation of a lamina densa and the appearance of pericytes) have been observed, i.e. vasculogenesis occurs. The origin of long, stretched cells is not known with certainty. They possibly represent smooth muscle cells. In addition, other cell types have been found, such as fibrocyte-like and fibroblast-like cells, elastoblasts, fat cells, monocytes and macrophages. All these cells are able to produce factors that promote the formation of new capillaries. Hence, a knowledge of these cells appears to be important for the analysis of in vitro systems. Moreover, the occurrence of these cell types must be considered when assessing possible effects.

Animals↗
PubMed1997

Immunolocalization of type I or type II activin receptors in the rat brain.

We have studied immunolocalization of activin receptors in the central nervous system using polyclonal antibodies (IgG) to type I (50-55 kDa, ActRI), type II (70-75 kDa, ActRII) or a subtype of type II known as type IIB (ActRIIB) receptors of activin. A total of 7 antisera to rat activin receptors was generated, i.e. 3 kinds of antisera to the extracellular domain (ActRI(81-89), ActRII(91-100), or ActRIIB(90-99)) and 4 antisera to the kinase domain (ActRI(323-333), ActRII(307-319), ActRII(407-420) or ActRIIB(306-319)). The region of aa 407-420 of ActRII is identical with that of ActRIIB. At first, we characterized these antibodies by Western blot analysis using ovarian proteins fractionated by preparative SDS-PAGE. All antibodies to ActRII and ActRIIB specifically reacted with 75 kDa-proteins which could also bind to activin-A. Anti-ActRII(91-100) antibody also reacted with 62 kDa-proteins which were capable of binding with activin-A. Although no positive reactions to anti-ActRI(81-89) antibody were seen in ovarian proteins, a positive reaction was detected at 52 kDa only when the proteins were deglycosylated. By use of these antibodies, immunolocalization of activin receptors was examined in the rat brain. The patterns of expression of activin type I and type II receptors were different. Positive reactions to anti-ActRII(91-100) antibody were detected in neurons of the cerebral cortex, hippocampus, medial amygdala and thalamus. In the hypothalamus, some neurons of the supraoptic nucleus were weakly stained, and widely scattered neurons of the lateral hypothalamic area were moderately stained. On the contrary, the most intense reactions to anti-ActRI(81-89) antibody were detected in neurons of the lateral hypothalamic area. In addition, many neurons of the cerebral cortex were also stained, but neurons of the hippocampus and the amygdala were not stained. These results suggest that activin may have physiological roles not only for hypothalamic neuroendocrinological and feeding-related systems as suggested previously but may also have functions in cortical and limbic pathways as a neuromodulator or for maintenance of neurons.

Activin Receptors↗
PubMed1997

Quantitative RT-PCR for inhibin/activin subunits: measurements of rat hypothalamic and ovarian inhibin/activin subunit mRNAs during the estrous cycle.

Inhibins (alpha-beta(A) and alpha-beta(B)) and activins (beta(A)-beta(A), beta(A)-beta(B) and beta(B)-beta(B)) were originally isolated from ovarian follicular fluids as FSH secretion modifiers. Inhibin/activin subunits, alpha, beta(A) and beta(B), are widely distributed in several tissues, including gonads and brain, and inhibins and activins have been reported to be involved in ovarian or hypothalamic functions. In this study, we established and employed a competitive RT-PCR assay system for rat inhibin/activin subunits by capillary electrophoresis to determine rat hypothalamic and ovarian inhibin/activin subunit mRNA levels during the estrous cycle. Linearity of standards for alpha, beta(A), and beta(B) subunit assays were between 0.01-0.3 amol, 0.003-0.09 amol and 0.002-0.02 amol of each fragment DNA as a standard, respectively. Hypothalamic beta(A) subunit mRNA during the estrous morning (1000 h) tended to be increased compared with that of the proestrous evening (1700 h), although they were not significantly different. Ovarian alpha subunit mRNA levels tended to be increased during the proestrous morning (1000 h) and were significantly increased in the proestrous evening (1700 h), compared with diestrus and estrus (P < 0.05). Ovarian beta(A) subunit mRNA was also significantly higher in the proestrous evening, compared with diestrus and estrus (P < 0.05), but in the case of beta(B) subunit mRNA there was no difference among diestrus, proestrus and estrus. We thus established a sensitive competitive RT-PCR system for the measurement of inhibin/activin alpha, beta(A) and beta(B) subunits, and this assay system would be helpful for the study of inhibin/activin action in brain and other tissues where these factors are expressed at low levels.

Animals↗
PubMed1996

Unilateral anomalous left common carotid artery; a case report.

An anomaly of the left common carotid artery was observed in a Japanese male cadaver during an anatomy class at the Saitama Medical School in 1995. The superior thyroid, lingual and facial arteries arose from the common carotid artery, and the posterior auricular, maxillary and superficial temporal arteries arose from the common carotid artery by a common trunk. The occipital and ascending pharyngeal arteries arose from the internal carotid artery. The left carotid body (glomus caroticum) was observed to be slightly below the lingual artery, behind the common carotid artery, and it was located at the level of the intervertebral disk between C2 and C3 or at the same level as the right carotid body. The carotid body was richly innervated by a branch of the glossopharyngeal nerve and by a plexus of sympathetic fibers from the vagus and glossopharyngeal nerves. We assumed that the artery above the level of the carotid body was the internal carotid artery; there was no specific external carotid artery and all branches of the external carotid artery arose from the internal carotid artery.

Aged↗
PubMed1996

Unique recognition of activin and inhibin by polyclonal antibodies to inhibin subunits.

Inhibin-A is a glycoprotein composed of an alpha subunit containing a glycosylation site and a beta A subunit, whereas activin-A is a homodimer of two inhibin beta A subunits. We examined the recognition of activin-A and inhibin-A by several antisera to the alpha or beta A subunit, and factors affecting the recognition. A total of six polyclonal antibodies to inhibin subunits, i.e., two antisera to a peptide fragment of the alpha subunit [alpha (1-19) and alpha (1-26)], and four antisera to the beta A subunit [beta A (1-10), beta A (70-79), beta A (87-99), and beta A (94-105)], was generated. On Western blot analysis, the anti-beta A (87-99) and beta A (94-105) sera recognized recombinant human activin-A but not inhibin-A under non-reducing conditions. When inhibin-A was deglycosylated with N-glycosidase-F, inhibin-A could be recognized by the anti-beta A (87-99) and beta A (94-105) sera. In addition, when activin-A bound to a nitrocellulose membrane was pre-incubated with recombinant human follistatin, the recognition of activin-A by the anti- beta A (87-99) and beta A (94-105) sera was decreased. These results suggested that the lower affinity of follistatin to inhibin-A than to activin-A might be likely explained as reflecting a site associated with the glycosylation of inhibin-A. However, the exposure of amino acids 87-105 of the inhibin beta A subunit on the molecular surface through deglycosylation did not increase the affinity of inhibin-A for follistatin but rather resulted in poor binding with follistatin. The present data suggest that (1) amino acids 87-105 of the inhibin/activin beta A subunit are located on the molecular surface, although this region of inhibin-A is concealed by the carbohydrate chain of the alpha subunit, (2) the region responsible for follistatin binding within the activin beta A subunit is spanned by amino acids 87-105, and (3) the mode of binding of inhibin-A to follistatin is quite different from that of activin-A to follistatin, and the former may be influenced by glycosylation.

Activins↗
PubMed1996

Follistatin and activin in bone: expression and localization during endochondral bone development.

The involvement of activin and follistatin, an activin-binding protein, in endochondral bone development was examined by sc implantation of demineralized bone matrix in rats. Immunoreactive follistatin was localized in proliferating chondrocytes and round osteoblasts, whereas it was not detected in hypertrophic chondrocytes and osteoblasts surrounding bone marrow. Western blot analysis also revealed that immunoreactive follistatin was higher during the initial stages of chondrogenesis (day 5) and osteogenesis (days 11 and 14) and lower during the conversion from cartilage to bone (day 9). These results suggest that follistatin is produced by proliferating cells, and the expression decreases with differentiation of the cells. Implants injected with follistatin on days 9 and 10 contained lower calcium levels on day 14 than those injected with rat albumin. Furthermore, the follistatin-injected implants were still mainly composed of cartilage, suggesting that the disappearance of follistatin is necessary for the conversion of cartilage to bone. In contrast, immunoreactive activin beta A (55-60 kDa) was continuously detected in implants on days 7-14. The content of C propeptide of type II procollagen was increased and cartilageous area was enlarged on day 7 by activin A injections on days 5 and 6, suggesting a chondrogenic effect of activin in the initial stage of cartilage formation. These results indicate that proliferating chondrocytes and round osteoblasts produce follistatin, and that the activity of activin is regulated by changes in the expression of follistatin at the stages of chondrogenesis and transition from cartilage to bone.

Activins↗
PubMed1995

Basement membrane formation of fetal mouse intestinal epithelial cells in organoid cultures.

Basement membrane formation of fetal mouse intestinal epithelial cells was investigated in organoid cultures. Intestinal cells were dissociated with a commercial collagenase/dispase preparation, and the cells were grown at high density on a membrane filter at the interface between the medium and air. This type of culture allows the histotypical reorganization of cells. After 2 days in culture, epithelial cells began to accumulate on the surface, in particular the periphery of the culture. These cells were usually cuboid, and small vesicles were formed in the center of the culture. Laminin-positive material was observed at peripheral sites. However, no basement membrane could be identified beneath the epithelial cells at the electron-microscopic level. After 3 days, epithelial cells that had gathered at the periphery became columnar in shape. Laminin-positive material extended across the surface of the culture. However, the vesicles formed in the center of the culture were not associated with laminin-positive material. Basement membrane was observed by electron microscopy at some sites beneath groups of epithelial cells, but did not extend continuously beneath these cells. Some epithelial cells made contact with the underlying mesenchymal cells through the discontinuous basement membrane via intercellular contacts. After 5-6 days, the surface of the culture was almost completely covered with epithelial cells and, at some sites, villus-like structures were visible. Laminin-positive material was clearly detectable under epithelial cells, as well as around epithelial vesicles located in the center of the culture. By electron microscopy, basement membrane was clearly visible between the epithelial and mesenchymal cells. After 9 days, villus-like structures were rarely observed. After 3 weeks, the cell mass had become smaller and villi had disappeared. Basement membrane was extensively folded and no basement membrane was visible at some sites. Formation of basement membrane by epithelial cells in monolayer culture occurs in an incomplete and irregular manner. It occurs rapidly in organoid cultures that include mesenchyme and epithelium. The organoid culture used here should be a useful tool for studies of the formation and degeneration of the basement membrane as well as interactions between the epithelium and mesenchyme.

Animals↗
PubMed1994

Crystallization and a preliminary X-ray diffraction study of macrophage migration inhibitory factor from human lymphocytes.

Two crystal forms of the macrophage migration inhibitory factor (MIF) from human lymphocytes have been obtained by the hanging drop method of vapor diffusion from ammonium sulfate solution. A trigonal crystal form belongs to the space group P3(1)21 (P3(2)21), with unit cell dimensions of a = b = 96.4 A and c = 105.5 A. Assuming that the asymmetric unit contains two or three dimers, the Vm value is calculated as 2.9 or 1.9, respectively. A second crystal form is orthorhombic, space group P2(1)2(1)2(1) with unit cell dimensions a = 68.4 A, b = 68.8 A and c = 86.8 A. The Vm value is 2.1 for two dimers in the asymmetric unit. Both crystals diffract to at least 1.9 A and are suitable for X-ray crystallographic structure determination at a high resolution.

Base Sequence↗
PubMed1994

Changes associated with the basal lamina during metamorphosis of Xenopus laevis.

We examined the changes in the morphology of the basal lamina of the colon, abdominal wall and tail muscles of tadpoles during metamorphosis and compared our results to those previously obtained for the small intestine. At the early stage of metamorphic climax (stages 60-62), we observed a curving of the basal lamina along the processes of the epithelial cells. At a later stage of metamorphic climax (stages 63-66), more extensive curving and folding of the basal lamina were observed. The basal lamina of the mesothelial cells in the abdominal wall did not change extensively as compared to the colon. Folded basal lamina was observed among degenerated muscle cells. Our results suggest that the folding of the basal lamina depends on the degree of deformation of the particular organ, which occurs mainly by physical factors such as the shortening or shrinking of the organ due to cellular degeneration. Cells that are newly differentiated produce an additional basal lamina, which results in transient doubling or layering. The combination of physical factors and cell differentiation results in a basal lamina that is more complex and contains folds and layers.

Animals↗
PubMed1993

Cell shape and arrangement of cultured aortic smooth muscle cells grown on collagen gels.

Aortic smooth muscle cells (SMC) grown on conventional plastic culture dishes have morphological and functional properties of dedifferentiated cells in subculture. We examined the influence of collagen gels on the cell shape and arrangement. The cells grown on collagen gels showed a multilayered growth with formation of nodules. When the edge of the collagen gels was detached from the culture dish, the shape and arrangement of cells on the edge differed from that of the central, still attached region. The cells grown on floating collagen gels exhibited a spindle-like shape and were arranged in concentric circles. These findings suggest that the physical property of the substrate influences the cell shape and arrangement.

Animals↗
PubMed1993

Type I collagen synthesis in cultured aortic smooth muscle cells from spontaneously hypertensive rats and normotensive control, Wistar-Kyoto rats.

An inhibition ELISA was used to quantify the amount of type I collagen synthesized in culture media and cell layers from aortic smooth muscle cells (SMCs) of spontaneously hypertensive rats (SHR) and normotensive control, Wistar-Kyoto rats (WKY). Cultured cells were also observed by electron microscopy. Collagen content in the culture media was strongly increased after 6 days in both cultures. Collagen and protein contents in the medium and cell layer from SHR were significantly higher than those in WKY at day 14. However, cell density in SHR-derived cells was also higher than that of WKY. No significant differences were detected in the rates of collagen content between SHR and WKY on a per cell basis. The main differences between SHR and WKY in collagen and protein levels may be due to the greater number of SHR cells and increased amounts of extracellular matrix components. The assay system outlined here should be useful for studying the control of extracellular-matrix synthesis.

Animals↗
PubMed1993

Phenotypic modulation and elastin formation of cultured aortic smooth muscle cells grown on and within collagen gels.

Aortic smooth muscle cells (SMC) were grown on and within collagen gels and their phenotype and elastin formation were examined. Cells grown on the gels showed multilayer growth with nodule formation. They contained many ribosomes and microfilament bundles with dense bodies. In contrast, cells grown within collagen gels did not form nodules. When collagen gels containing SMC were manually detached from the culture dish, the cells were found to be of the synthetic phenotype with well-developed rough endoplasmic reticulum (ER) and Golgi complexes, but few microfilament bundles. There were numerous elastin deposits in the intercellular spaces. Our findings suggest that the physical property of the substrate affects the phenotype and function of the SMC.

Animals↗
PubMed1991

Morphologic changes of the basal lamina in the small intestine of Xenopus laevis during metamorphosis.

Further investigations of the epithelial and mesothelial basal lamina of the duodenum of Xenopus laevis during metamorphosis were performed by means of scanning electron microscopy (SEM) and histochemical techniques using polyethyleneimine (PEI) to demonstrate anionic sites as well as light- and transmission-electron-microscopic methods involving morphometric analysis. The basal lamina of the duodenal epithelial cells was smooth, and it was occasionally curved along the processes of the epithelial cells (stages 56-59). The basal lamina became thicker by folding, and the thickness of the folded basal lamina exceeded 1 micron (stages 60-62). Subsequently, the folded basal lamina disappeared gradually and became almost smooth again and consisted of only one layer (stages 63-66). After removing the epithelium by boric acid, SEM revealed that the small ridges of the basal lamina protruded like a mesh-work into the luminal side, and the luminal surface of the basal lamina became smooth at later stages of the metamorphic climax. The electron-dense granules of PEI-positive material were localized at both sides of the lamina densa at regular intervals (80-100 nm). The basal lamina of the mesothelial cells was almost smooth at stages 56-59 and started to show occasional slight folding. This folding became continuous and deeper (stages 60-62). The folded mesothelial basal lamina disappeared except for the cell-associated basal lamina and became smooth again at later stages of the metamorphic climax (stages 63-66). These morphologic changes of the basal lamina observed in the epithelium and mesothelium may be induced by common factors. We suggest that physical changes in the small intestine involving the shortening and narrowing should be a main factor to cause these changes in the basal lamina. Furthermore, morphometric analysis proposed that the basal lamina becomes more complex by adding newly synthesized basal lamina material, especially in the epithelium.

Animals↗
PubMed1989

Observation on the basal lamina of duodenal mesothelial cells during metamorphosis of Xenopus laevis.

Morphologic changes in the basal lamina of duodenal mesothelial cells during metamorphosis of Xenopus laevis were observed. In the prometamorphosis stage (Stage 56-59), the basal lamina was almost completely flat; the lamina densa of the basal lamina was a 50 nm layer of high electron density. In the early stages of metamorphic climax (Stage 60-62), the basal lamina showed occasional slight folding (stage 60), with the lapse of time, the folding became continuous and deeper. The development of an additional thin basal lamina was observed in areas where the folded basal lamina was separated from mesothelial cells, viz. on the side adjacent to the mesothelium. The lamina densa in this stage was approximately twice the thickness of the prometamorphosis stage and exhibited high electron density. In the later stages of metamorphic climax (stage 63-66), the basal lamina just under mesothelium became more apparent. The folded basal lamina shifted from the mesothelium into the subserosa and gradually disappeared, and the basal lamina became a single layer. The thickness of the lamina densa was almost the same as in the prometamorphosis stage. Since the timing of the folding of the basal lamina coincides with the shortening of the digestive tract and the marked narrowing of the lumen, we suggest that physical changes in the digestive tract during metamorphosis may play an important role in these morphologic changes of the basal lamina.

Animals↗
PubMed1987

New technique for pancreaticojejunostomy using a biological adhesive.

A new technique of pancreaticojejunostomy using a fibrin glue biological adhesive system (BAS) after pancreaticoduodenectomy is reported. Pancreaticojejunostomy was performed on 112 patients between 1971 and 1985. Forty-two of the patients received BAS application (Group I), but the remaining seventy did not (Group II). To evaluate more precisely the effectiveness of BAS, 22 patients in Group I (Group I-ST) and 26 patients in Group II (Group II-ST) who were operated upon by the same surgeon during the same period were reviewed retrospectively. The selection of patients for the use of BAS was randomized. In Group I-ST, one patient had a minor leak; no major leaks or operative deaths occurred. Two of the twenty-six patients in Group II-ST had minor leaks and one patient had a major leak causing death. This new technique for pancreaticojejunostomy using a biological adhesive system may be useful for preventing anastomotic leaks, especially in a patient with a normal pancreas.

Bandages↗
PubMed1987

Immuno-electron-microscopic application of antiserum against elastin.

The antiserum against insoluble elastin from human aorta was applied to immuno-electron microscopy. In the preembedding method, only the outer surface of the amorphous component (elastin) of elastic fibers showed a positive reaction. A major problem encountered with the preembedding method is associated with the penetration of either the primary antiserum or the secondary antibody into the tissue and, in particular, into elastin. On the contrary, a positive reaction was observed inner zones of elastin in the postembedding method. While the postembedding method employed here has limitations with nonspecific binding to the embedding medium, the postembedding method offers a decided advantage over the preembedding method.

Animals↗