Biomedical subjects

H Kawaguchi

Publications and source records attributed to H Kawaguchi.

At least 19 recordsLinked to original sources

PubMed1992

A new co-culture method utilizing a dual compartment cell-allocation apparatus for observation and identification of synapse formation in vitro.

A new co-culture method for observation and identification of synapse formation utilizing dissociated retinal and tectal neurons from 7-day-old chick embryos was developed using a glass apparatus which we named 'Dual Compartment Cell-Allocation Apparatus.' This apparatus enabled us to culture these two types of cells by forming two parallel lanes, each with a dimension of 200 microns by 15 mm and with an interval of 200 microns separating the two lanes, on a collagen-coated cover slip. Infiltration and/or migration of retinal cells into the tectal cell lane was minimal with only 0.15% identified and confirmed with the fluorescence-dye labeling method. Growth of neurites connecting the two lanes was observed within 48 h of culture in a serum-free medium. Double-immunostaining of the culture preparations with anti-neurofilament and anti-synaptophysin antibodies revealed the presence of synaptophysin-hot spots on neurofilament-positive processes connecting the retinal neurons with the tectal neurons, thus confirming the formation of synapses between the neurons of the retinal lane and the neurons of the tectal lane. The dual compartment cell-allocation apparatus has proven itself to be advantageous and useful for studying the formation of synapses between two types of neurons from different origin.

Animals↗

PubMed1992

Inhibition of IgM antibody-mediated aggregation of Trypanosoma gambiense in the presence of complement.

This paper deals with the immune reaction between Trypanosoma gambiense and monoclonal IgM mouse antibody at equivalence with or without rabbit complement. Antibody-mediated trypanosome clumps formed in the absence of complement, and were readily dissociated by complement to become free. In the presence of complement, on the other hand, T. gambiense were not aggregated by the antibody. Free parasites adhered readily to cultured peritoneal macrophages. Complement-mediated dissociation of the clumped trypanosomes in the equivalence area released a large number of previously bound surface antigens. These antigens were capable of binding again to fresh IgM antibody. Experimental results further indicated that the complement system caused a functional alteration, changing the multivalent nature of the IgM antibody in the immune complex into a univalent one. This phenomenon is of great advantage to the infected host in clearing pathogens in vivo, as it allows more antibodies to attach to trypanosomes and subsequently initiate complement activity.

Animals↗

PubMed1992

Dynamical behavior of psb gene transcripts in greening wheat seedlings. I. Time course of accumulation of the pshA through psbN gene transcripts during light-induced greening.

The time course of the accumulation of the transcripts from 13 psb genes encoding a major part of the proteins composing photosystem II during light-induced greening of dark-grown wheat seedlings was examined focusing on early stages of plastid development (0.5 h through 72 h). The 13 genes can be divided into three groups. (1) The psbA gene is transcribed as a single transcript of 1.3 kb in the dark-grown seedlings, but its level increases 5- to 7-fold in response to light due to selective increase in RNA stability as well as in transcription activity. (2) The psbE-F-L-J operon, psbM and psbN genes are transcribed as a single transcript of 1.1 kb, two transcripts of 0.5 and 0.7 kb and a single transcript of 0.3 kb, respectively, in the dark-grown seedlings. The levels of accumulation of every transcript remain unchanged or rather decrease during plastid development under illumination. (3) The psbK-I-D-C gene cluster and psbB-H operon exhibit fairly complicated northern hybridization patterns during the greening process. When a psbC or psbD gene probe was used for northern hybridization, five transcripts differing in length were detected in the etioplasts from 5-day old dark-grown seedlings. After 2 h illumination, two new transcripts of different length appeared. Light induction of new transcripts was also observed in the psbB-H operon.

Blotting, Northern↗

PubMed1992

C4 genes of the chimpanzee, gorilla, and orang-utan: evidence for extensive homogenization.

The human complement component 4 is encoded in two genes, C4A and C4B, residing between the class I and class II genes of the major histocompatibility complex. The C4A and C4B molecules differ in their biological activity, the former binding more efficiently to proteins than to carbohydrates while for the latter, the opposite holds true. To shed light on the origin of the C4 genes we isolated cosmid clones bearing the C4 genes of a chimpanzee, a gorilla, and an orang-utan. From the clones, we isolated the fragments coding for the C4d part of the gene (exons and introns) and sequenced them. Altogether we sequenced eight gene fragments: three chimpanzee (Patr-C4-1*01, Patr-C4-1*02, Patr-C4-2*01), two gorilla (Gogo-C4-1*01, Gogo-C4-2*01), and three orang-utan (Popy-C4-1*01, Popy-C4-2*01, Popy-C4-3*01). Comparison of the sequences with each other and with human C4 sequences revealed that in the region believed to be responsible for the functional difference between the C4A and C4B proteins the C4A genes of the different species fell into one group and the C4B genes fell into another. In the rest of the sequence, however, the C4A and C4B genes of each species resembled each other more than they did C4 genes of other species. These results are interpreted as suggesting extensive homogenization (concerted evolution) of the C4 genes in each species, most likely by repeated unequal, homologous, intragenic crossing-over.

Animals↗

PubMed1992

Polyphosphoinositide metabolism in hypertrophic rat heart.

The accumulations of inositol-1,4,5-trisphosphate (IP3) and inositol-1,3,4,5-tetrakisphosphate (IP4) after hormonal stimulation may have a physiological role, possibly by alteration of Ca2+ levels in cardiac tissue. But the accumulation of inositol polyphosphate in a pathophysiological condition has not been studied. We investigated phosphatidylinositol-4,5-bisphosphate (PIP2) metabolism in hypertrophic cardiac myocytes, and clarified that the accumulations of IP3, IP4 and diacylglyceride after stimulation with norepinephrine were significantly enhanced in isolated myocytes from spontaneously hypertensive rat heart. Phospholipase C activity increased with age in SHRSP heart cells. These data suggest that PI turnover pathways, which can be mediated by both phosphatidylinositol-4,5-bisphosphate and diacylglyceride, may play an important role in development of hypertrophy in the hearts of rats with spontaneous hypertension.

Animals↗

PubMed1992

Organization of C4 and CYP21 loci in gorilla and orangutan.

The standard human haplotype contains two C4 and two CYP21 loci arranged in the order C4A ... CYP21P ... C4B ... CYP21 and intercalated between the class I and class II loci of the HLA complex. The C4A gene is 22 kilobases (kb) long; the C4B gene is either 22 kb or 16 kb long. The CYP21P is a pseudogene characterized by an eight base pair (bp) deletion in exon 3 and other defects; the CYP21 is a functional gene. The standard chimpanzee haplotype is arranged in the same way as the standard human haplotype, except that both C4 genes are of the short variety; like the human gene, the chimpanzee CYP21P gene contains the 8 bp deletion. In the present study we demonstrate that a representative gorilla haplotype also consists of two short C4 genes and two CYP21 genes, neither of which, however, has the characteristic 8 bp deletion. On the other hand, the single characterized orangutan haplotype is organized in the following way: C4A ... CYP21 ... C4A ... CYP21 ... C4B ... CYP21. The first two C4 genes are of the long variety, the third gene is short. None of the defects characterizing the human CYP21P gene is present in any of the three orangutan genes. These conclusions are based on the analysis of overlapping clones isolated from cosmid libraries of the indicated species. The observed haplotype organization of the four primate species can be explained by expansion and contraction of the C4-CYP21 region through unequal homologous crossing-over, which preserves the differentiation of the C4 genes into the A and B categories but otherwise homogenizes these genes, as well as the CYP21 genes, within a given species. The 8 bp deletion in the CYP21P gene is postulated to have occurred before the separation of the lineages that led to modern humans and chimpanzees, but after the separation of these two lineages from the lineage that led to modern gorillas. The 6 kb insertion generating the long C4 gene is postulated to have occurred before the separation of the orangutan, gorilla, chimpanzee, and human lineages.

Animals↗

PubMed1992

Direct purification of multiple ATF/E4TF3 polypeptides from HeLa cell crude nuclear extracts using DNA affinity latex particles.

We developed a method using affinity latex particles to rapidly and efficiently purify DNA-binding proteins directly from crude cell extracts. The particles are composed of a styrene core and a polyglycidyl methacrylate surface, to which DNA oligomers were immobilized by means of epoxy groups. Multiple polypeptides were copurified, which bound to the ATF/E4TF3-binding site from crude nuclear extracts of HeLa cells, within a few hours. Affinity-purified polypeptides stimulated transcription in vitro from a promoter in which ATF/E4TF3-binding sites were present. At least eight polypeptides with molecular masses of 116, 80, 65, 60, 55, 47, 45, and 43 kDa were copurified. About 2 micrograms of the 43-kDa protein was purified directly from 8 mg of crude nuclear extracts. All the polypeptides directly bound to the same DNA sequence and were thought to form a family. The results indicated that the particles are useful for quickly purifying various DNA-binding proteins directly from crude cell extracts.

Activating Transcription Factors↗

PubMed1992

Estimation of relative fecundity in Eimeria tenella strains by a mixed infection method.

The relative fecundity of populations of Eimeria tenella was estimated by means of mixed infection using electrophoretic variation of glucose-phosphate isomerases (GPIs) as a genetic marker. A decoquinate-resistant strain with GPI-9 isozyme (DR-NIAH), a decoquinate-sensitive one with GPI-1 (DS-Iwate), and three decoquinate-resistant lines (No. 3, 4, and 5) derived from cross-fertilization between DR-NIAH and DS-Iwate, were used. The GPI phenotypes of the No. 3 and No. 4 lines are GPI-9, and that of No. 5 is GPI-1. Mixed infection experiments were performed between DR-NIAH and DS-Iwate, No. 3 and No. 5, and No. 4 and No. 5. DR-NIAH was predominant over DS-Iwate in the mixed infection, whereas, in single infections, the total oocyst output of DR-NIAH was similar to or less than that of DS-Iwate. Among three lines, No. 4 was predominant over No. 5, and No. 5 was predominant over No. 3 in the mixed infection. Relative fecundity between No. 3 and No. 5 and their patterns of oocyst output in single infections were similar to those in the mixed infection. In contrast, No. 5 produced more oocysts than No. 4 in single infections, suggesting that the oocyst production in the mixed infection may be influenced by the mutual interference or competition between the populations of E. tenella in the chicken caeca.

Animals↗

PubMed1992

Immunohistochemical demonstration of bone morphogenetic protein-2 and transforming growth factor-beta in the ossification of the posterior longitudinal ligament of the cervical spine.

To clarify the mechanism of ossification of the posterior longitudinal ligament, immunohistochemical localization of bone morphogenetic protein-2 and transforming growth factor-beta was examined using surgical specimens of ligament tissues from an affected patient. Two polyclonal antibone morphogenetic protein-2 antibodies and an anti-human transforming growth factor-beta antibody were used as primary antibodies. Bone morphogenetic protein-2 and transforming growth factor-beta were present in ossified matrix and chondrocytes of adjacent cartilaginous areas of ossification of the posterior longitudinal ligament. Although immunostaining with antibone morphogenetic protein-2 antibodies also was observed in mesenchymal cells with fibroblastic features in the immediate vicinity of the cartilaginous areas, no staining could be detected with anti-human transforming growth factor-beta antibody in these cells. The presence of these factors were specific for the ossified ligament because no immunostaining was observed after using antibodies in the posterior longitudinal ligament at unossified levels from the same patient. It is suggested that bone morphogenetic protein-2 and transforming growth factor-beta play important roles in the development of ossification of the posterior longitudinal ligament and that bone morphogenetic protein-2 may act as an initiating factor in the development of ossification of the posterior longitudinal ligament by stimulating differentiation of mesenchymal progenitor cells. Transforming growth factor-beta may stimulate bone formation at a later stage of the process of ectopic ossification.

Aged↗

PubMed1992

A case of congenital trichofolliculoma.

A case of congenital trichofolliculoma on the right cheek of a 3-month-old boy is reported. Trichofolliculoma usually appears in middle age; it is unusual in childhood. This is the first reported case of congenital trichofolliculoma.

Cheek↗

PubMed1992

Ultrastructural and ultracytochemical characteristics of multinucleated cells after hydroxyapatite implantation into rat periodontal tissue.

Multinucleated cells (MNCs) that appeared after hydroxyapatite (HAP) implantation into experimentally-produced bone defects in rat periodontal tissues were investigated both ultrastructurally and ultracytochemically. At day 5 after implantation, MNCs first appeared along the HAP surface. They had no features of typical osteoclasts such as ruffled border and clear zone. By d 14, these cells acquired features similar to osteoclasts, including ruffled border and clear zone. With the appearance of ruffled borders in MNCs, new bone deposited around the implanted HAP. MNCs appeared to excavate both newly-formed bone and implanted HAP simultaneously. Ingested HAP particles were observed not only in MNCs but also in macrophages. MNCs contained both tartrate-resistant acid phosphatase (ACPase) and carbonic anhydrase (CAase). ACPase activity was detected along all the biosynthesizing pathways in MNCs. Extracellular ACPase activity around the ruffled border region was also demonstrable. CAase activity could be detected only in the cytosol, vesicles and mitochondrial cristae of the MNCs. These cytochemical characteristics were almost the same regardless of the time elapsed after implantation.

Acid Phosphatase↗

PubMed1992

Expression of the angiotensinogen gene and localization of its protein in the human heart.

BACKGROUND: There have been no reports on the presence of the tissue renin-angiotensin system in the human heart, although the presence of angiotensinogen has been described in the animal heart. METHODS AND RESULTS: To determine whether angiotensinogen is synthesized in the human heart, we examined angiotensinogen messenger RNA (mRNA) synthesis in autopsy hearts by using ribonuclease protection assay. As a result, angiotensinogen mRNA was detected in the atrial muscle, muscles of the conduction system, and the left ventricular wall. In the left ventricular wall, mRNA expression was more prominent in the subendocardial muscles than in the midcardial or epicardial muscles. Using a monoclonal antibody to human angiotensinogen in immunoblotting experiments, we detected two closely spaced bands at approximately 70 kd in the heart, which was quite consistent with the human angiotensinogen molecule. Immunohistochemical studies with this monoclonal antibody demonstrated intense immunoreactivity in the atrial muscles, the muscles of the conduction system, and those of the subendocardial layers. CONCLUSIONS: We conclude that angiotensinogen was synthesized in the human heart. It was evident that the localization of angiotensinogen was not ubiquitous in the cardiac muscles, showing its predilection for the atrial muscles, muscles of the conduction system, and subendocardial layer of the left ventricle.

Angiotensin II↗

PubMed1992

The studies of cell damaging and cell growth factors which induce cardiomyopathy.

We demonstrated that phosphatidylinositide-specific phospholipase C (PLC) activity was greater in cardiomyopathic hamster hearts (BIO 14.6 and BIO 53.58) then in hamster controls (F1b). Inositol trisphosphate (IP3) production was markedly greater in both of the cardiomyopathic hamsters, BIO 14.6 and BIO 53.58. We have also determined the sarcoplasmic reticulum (SR) function of heart. Calcium uptake into SR markedly increased in BIO 14.6. On the other hand, it significantly decreased in BIO 53.58 compared with F1b. It is well known that IP3 stimulates calcium release from SR. In BIO 14.6, calcium release from SR stimulated by IP3 increased, but its effect decreased in BIO 53.58 compared with F1b. These results suggest that PI response may produce high intracellular calcium levels in both BIO 14.6 and BIO 53.58 myocytes. In addition, in the BIO 53.58 hamster the sarcoplasmic reticulum deteriorate in function. It was concluded from these results that a prolonged high intracellular calcium level may lead to the death of BIO 53.58 myocytes. The expression of angiotensinogen mRNA was observed in the hamster heart. There was no differences in its expression level between F1b, BIO 14.6 and BIO 53.58. There was no effect of ages on its expression in these hamster hearts. We have also determined the distribution of angiotensinogen in these hamsters. At 4 weeks of age, the immunohistochemical study revealed that angiotensinogen was widely distributed in subendocardium in these hamsters. There was no difference in its distribution between F1b, BIO 14.6 and BIO 53.58. But at 20 weeks old of age its immunoreactivity decreased in BIO 53.58.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensinogen↗

PubMed1992

Phosphatidylinositol and inositolphosphatide metabolism in hypertrophied rat heart.

The accumulation of both Inositol-(1,4,5)-trisphosphate (IP3) and Inositol-(1,3,4,5)-tetrakisphosphate (IP4) after hormonal stimulation has a physiological role, possibly in altering Ca2+ levels in cardiac tissue. However, the accumulation of inositol polyphosphate under pathophysiological conditions has not been studied. In our experiments the metabolism of phatidylinositol and IP3 in cardiac myocytes as investigated. It was shown that basal levels of cytosolic phosphatidylinositol specific phospholipase C (PI-PLC), phosphatidylinositol-(4,5)-bisphosphate specific phospholipase C (PIP2-PLC) activities markedly increased in stroke-prone spontaneously hypertensive rats (SHRSP) with age compared with age matched Wistar Kyoto rats (WKY). IP3 kinase and IP3 phosphatase activities also increased in SHRSP hearts with age. Their activities increased in WKY, but to a lesser extent than in SHRSPs. These data suggest that a PI turnover pathway such as the phosphatidylinositol 4,5-bisphosphate-IP3-Ca2+ pathway or the diacylglyceride-protein kinase C pathway may have an important role in the development of hypertrophy in SHRSP heart.

Animals↗

PubMed1992

A mechanism of catecholamine tolerance in congestive heart failure--alterations in the hormone sensitive adenylyl cyclase system of the heart.

It is well known that failing hearts show diminished responsiveness (desensitization) to catecholamines. In this study, 2 different animal models were used to investigate the alterations in the hormone sensitive cardiac adenylyl cyclase system in a congestive heart failure. In the first model, cardiomyopathic Syrian hamsters (BIO53.58), we found reduced activity of the catalytic protein of adenylyl cyclase; this reduction was more prominent in an older animals (28-week-old vs 16-week-old). At both ages, the amount of inhibitory GTP-binding protein (Gi) was markedly increased in BIO53.58 compared to control healthy hamsters. Moreover the increased Gi was shown to be fully functional in the inhibitory pathway of the adenylyl cyclase system. In the second model, chronically norepinephrine-infused rats, we found a decrease in beta-adrenergic receptor density at an early stage of injection (3 days), while the activity of catalytic protein decreased beyond 14 days of injection, and the amount of Gi increased after 7 days of injection. These results suggest that increased plasma catecholamine concentrations in the setting of congestive heart failure might be a major trigger for qualitative and quantitative alterations observed in various components of cardiac adenylyl cyclase system, and that GTP-binding proteins and catalytic protein of adenylyl cyclase are involved in the mechanism of desensitization especially after chronic in vivo stimulation of adrenergic receptors.

Adenylyl Cyclases↗

PubMed1992

Detection of serum antibodies in Eimeria tenella-infected chickens by enzyme linked immunosorbent assay (ELISA) with merozoite and oocyst antigens.

Soluble antigens prepared from sporulated oocytes and second generation merozoites of E. tenella were used for enzyme linked immunosorbent assay (ELISA) to investigate antibody in sera of two breeds of chickens, i.e. commercial broilers and SPF single comb white leghorn layers, which were experimentally infected with E. tenella. In broilers inoculated with oocysts at 15 days of age, ELISA values increased rapidly after day 19 post inoculation (PI) and reached the maximum lebel on days 29 and 32 PI against both merozoite and oocyst antigen. The values against merozoite antigen were significantly higher than those against oocyst antigen. In SPF layers infected at 15 days of age, the values increased gradually after 7 days PI. There were no significant differences between values against two antigens. Generally, the values in broilers tended to be higher than those in SPF layers, especially against merozoite antigen. In broilers inoculated with oocysts at 1 and 15 days of age, ELISA values increased rapidly and reached the maximum level on days 11 and 20 post second inoculation (PSI) against merozoite and oocyst antigens respectively and then the values against merozoite antigen decreased. The values against merozoite antigen were markedly higher than those against oocyst antigen. In SPF layers inoculated twice, the values reached the highest on day 11 PSI as in the case of broiler; however, after that day, the values against both antigens decreased. The sera reacted similarly against both antigens. The values against merozoite antigen were significantly higher in broilers than in SPF layers.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

The role of calcium activated neutral protease on myocardial cell injury in hypoxia.

The aim of this study was to investigate the correlation between hypoxic myocardial cell injury and intracellular protease activity. Cardiac myocytes were isolated from neonatal rat hearts and cultured in Eagle's modified minimum essential medium. Myocytes were incubated in hypoxic conditions for 6 hours. The cell death rate during hypoxia rose to 80% after 6 hours. Extracellular protease activity was elevated to 4 units during hypoxia, much higher than the 0.7 units in aerobic states at 6 hours. This extracellular protease activity in hypoxic conditions was markedly inhibited by leupeptin and EDTA, and weakly inhibited by the cysteine protease inhibitor, NCO-700, but phenylmethyl sulfonyl fluoride did not inhibit the protease activity. To identify the protease activated during hypoxia, calpain-specific inhibitors were added to the incubation mixture. Calpain inhibitor 1 and calpastatin, an endogenous selective calpain inhibitor, markedly inhibited extracellular protease activity during hypoxia. NCO-700 also inhibited intracellular protease activity. NCO-700 reduced hypoxic cell death to 30% after 6 hours of hypoxygenation. These observations indicate that calpain is activated during hypoxia and leads to irreversible cell membrane degradation after 6 hours of hypoxygenation.

Animals↗

PubMed1992

[A fifty two-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T) in dogs].

A 52-week oral repeated dose toxicity study of suplatast tosilate (IPD-1151T), a newly developed anti-allergic agent, was carried out in beagles by oral administration of 30, 90, 270 and 810 mg/kg/day for 52 weeks. The recovery study was carried out by the withdrawal for 5 weeks using control and the 810 mg/kg groups. The results are as follows: 1. Observation of general conditions revealed soft feces, mucous feces, and diarrhea in both sexes of the 270 and 810 mg/kg groups during the administration period, and these findings disappeared during the withdrawal period. One female of the 810 mg/kg group exhibited tremors in the legs and neck, staggering, a decrease of spontaneous motor activity, and clonic convulsions in Week 17 of administration and died on Day 118. One male of the same group exhibited whole body tremors and staggering from Week 32 to Week 52. 2. Body weight gain tended to be inhibited in males of the 810 mg/kg group during the administration period. The body weight of the female that died decreased rapidly after the appearance of neurological symptoms. The body weight of the male that exhibited neurological symptoms decreased after their appearance but later increased. 3. There were no abnormal changes in food consumption in all of the sacrificed dogs. The female that died did not eat at all after the appearance of neurological symptoms. The male that exhibited neurological symptoms did not eat at all for 1 week after their appearance, but the food consumption returned to normal thereafter. 4. Prothrombin times were prolonged in males of the 270 and 810 mg/kg groups at Week 26, and activated partial thromboplastin times were prolonged in males of the 810 mg/kg group at Week 52. 5. Plasma levels of alkaline phosphatase, GPT and LDH were elevated in some males and females of the 810 mg/kg groups. 6. No abnormalities due to IPD-1151T administration were found in urinalysis, opthalmological examination, electrocardiography, and fecal occult blood examination, or organ weights. 7. Autopsies including histopathological and electron microscopic examinations on the sacrificed dogs revealed no abnormalities. Subserosal hemorrhage in the base of the heart, congestion in the lungs, congestion and vacuolation in the liver and slight cell infiltration around vessels of the brain were found in the female that died.(ABSTRACT TRUNCATED AT 400 WORDS)

Administration, Oral↗