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K Nokihara

Publications and source records attributed to K Nokihara.

53 records · Page 3Linked to original sources

Effect and distribution of intravenously injected 125I-endothelin-1 in rat kidney and lung examined by electron microscopic radioautography.

The morphological effect of endothelin-1 (ET-1) and the distribution of endothelin-binding sites on the kidney and lung was investigated ultrastructually by intravenous injection of [125I]-ET-1 into rats. About 10% decrease of the diameter of glomeruli was observed at 10 min after the injections of ET-1 or [125I]-ET-1 (1.3-2.4 nmole/kg). When localization of [125I]-ET-1 in the kidney was examined by light and electron microscopic radioautography, silver grains were preferably localized on the fenestrated endothelial cells of glomeruli and peritubular capillary endothelial cells. Some grains were also localized on the interdigitating processes of urinary tubules. Quantitative analysis of silver grains in the glomeruli showed that 83% of grains were located on the fenestrated endothelial cells, 12% on the podocytes of visceral cells, and 5% on mesangial cells at 10 min. After 60 min, 50% of silver grains were incorporated into the cytoplasm of fenestrated endothelial cells. In contrast to glomeruli, silver grains were rare on the arteries and large arterioles. However, a few silver grains were often observed on the smooth muscle cells of small arterioles (8-20 microns in diameter). In the lung, 70% of silver grains were located on the alveolar capillary endothelial cells. These results indicate the abundance of ET receptors on the glomerular fenestrated endothelium, peritubular fenestrated endothelium and alveolar capillary endothelium.

Animals↗

Binding of 125I-endothelin-1 to fat-storing cells in rat liver revealed by electron microscopic radioautography.

Localization of intravenously injected [125I]-endothelin-1 was examined in rat liver by light and electron microscopic radioautography. At 10 min after injection, silver grains were localized along the sinusoidal wall, i.e., mostly on the thin processes of fat-storing cells and sinusoidal endothelial cells, and also on the Kupffer cells and the microvilli of hepatocytes. About 35% of the total silver grains were located on the processes of fat-storing cells at 10 min. The grain density (number of silver grains/cell area) of fat-storing cells was three-fold that of Kupffer cells, and 18-fold that of hepatocytes. At 60 min, 60% of the total grains were observed on the fat-storing cells, though the value of grain density was not changed. Silver grains were internalized into the cytoplasm of fat-storing cells and often associated with multivesicular bodies. In contrast, the grain density of endothelial cells and Kupffer cells decreased with time. These results indicate that hepatic fat-storing cells have a considerable number of endothelin-binding sites, and incorporate bound endothelin into cytoplasm.

Animals↗

Localization of [125I]endothelin-1 in rat tissues observed by electron-microscopic radioautography.

[125I]Endothelin-1 (ET-1) was administered intravenously into rats, and binding sites in various tissues were examined by electron-microscopic radioautography. Labeling was observed predominantly on the glomerular fenestrated endothelial cells of kidney, alveolar capillary endothelial cells of lung, fat-storing cells in the sinusoids of liver, and subepithelial myofibroblasts of the small intestine. At first, silver grains were observed mostly on the plasmalemma of these cells and with time translocated into the cytoplasm of these cells. Myofibroblasts of the small intestine were cultured to study the cellular effects of ET. By fura-2 fluorescence method, transient increase and oscillation of intracellular Ca2+ level were observed in the cultured myofibroblasts following the application of ET-1 or endothelin-3 (ET-3). These results suggest that ET induces diverse physiological reactions in specific cells of various tissues.

Animals↗

125I-endothelin binds to fibroblasts beneath the epithelium of rat small intestine.

Rats were intravenously injected with synthetic 125I-endothelin, and then were fixed by intracardiac perfusion at 10 min, 30 min and 1 hr after the injection. Localization of 125I-endothelin was examined in the small intestine by light and electron microscopic radioautography. Through the time course examined, more than seventy percent of total silver grains were localized in the fibroblasts, especially beneath the basal lamina of epithelium of the intestinal villi. Silver grains were observed mostly on the thin processes, and also on the vacuoles. At 10 min, 80% of silver grains on the fibroblasts were located on the plasmalemma. By the chase, grains on the plasmalemma decreased, and silver grains in the cytoplasm increased gradually.

Animals↗

The heart is the center of a new endocrine, paracrine, and neuroendocrine system.

This review indicates that the heart is a polypeptide-producing organ which should be classified among the traditional endocrine tissues. Cardiac hormones have only been known for a few years, the discovery of their endocrine functions, however, occurred in the 1950ies when Gauer, Henry and Kisch observed specific physiological and morphological features of the heart atria indicative of an endocrine activity. Because of their basic effects many target organs involved in the regulation of body fluid pressure and composition are related to this endocrine organ located in the atrial appendages of the heart. The compact endocrine portion of the heart is built up by myoendocrine cells which form the functional endocrine units and produce a variety of polypeptide hormones called cardiodilatin (CDD) or atrial natriuretic polypeptide (ANP), which belong to one family. Also, co-storage of a partially homologous regulatory polypeptide called brain natriuretic polypeptide (BNP) occurs, as has been determined by immunohistochemistry and radioimmunoassay. CDD and/or BNP are found in numerous organs where they exert paracrine and neurocrine functions, e.g., in the brain, peripheral nervous system, kidney, and adrenal medulla. In these organs, a differential post-translational processing of cardiac polypeptides is observed, resulting in different functional activities according to discriminating receptor interactions and degrading metabolism. Some of the extra-auricular sites of synthesis and storage of CDD-like peptides are briefly mentioned. In summary the heart constitutes the center of a multilocal and multifunctional system of specific cardiac polypeptides of endocrine, paraneuronal, and neuronal character.

Animals↗

Immunocytochemical evidence for CDD/ANP-like peptides in strands of myoendocrine cells associated with the ventricular conduction system of the rat heart.

Immunohistochemical investigations with different antisera against cardiodilatin 99-126 or alpha atrial natriuretic polypeptide revealed the presence of cardiac hormones not only in the atria of rats but also in strands of myoendocrine cells located in subendocardial regions of the ventricular septum. The localization of CDD-IR (cardiodilatin immunoreactivity) in the ventricle is associated with the location of the conduction system in the rat. The significance of the morphological relationship between cardiodilatin and the conduction system of the rat heart is discussed.

Aging↗

PHI structural requirements for potentiation of glucose-induced insulin release.

Immunoreactive PHI was detected in rat pancreas. The potentiating effect of 10(-9) M PHI upon insulin release from the isolated perfused rat pancreas was significant and most consistent when 250 mg% glucose was present in the perfusion medium. PHI(1-15) retained a substantial potentiating effect on insulin release, while PHI(14-27) was essentially inactive. Replacement of amino-terminal decapeptide portion of the PHI molecule with the corresponding part of VIP resulted in a drastic decrease of the potentiating effect of PHI on insulin release. 10(-8) M PHI(14-27) substantially diminished the potentiation by 10(-9) M PHI while PHI(1-15) was without an inhibitory effect. The present results indicate that the PHI active site for potentiation of glucose-induced insulin release resides in the amino-terminal segment of the molecule but requires the carboxyl terminal segment primarily for binding to exhibit full biological activity.

Animals↗

Absence of effect of the peptide PHI-27 on LH release in vitro.

The effects of PHI-27, a peptide of the glucagon-secretion family, on luteinizing hormone (LH) release and on LH-releasing hormone (LH-RH)- or estradiol-induced LH release were examined in a sequential double chamber perifusion system by perifusing the pituitary alone or in sequence with the mediobasal hypothalamus (MBH) from normal female rats in diestrus. PHI at 10(-7) M had no significant effect on LH release from the pituitary in series with the MBH. Moreover, on perifusion of the pituitary alone with medium containing 10(-7) M PHI, LH release induced by 20 ng/ml LH-RH from the pituitary was not significantly different from that without PHI. Furthermore, PHI had no effect on estradiol-induced LH release from the pituitary in sequence with the MBH. These data indicate that PHI has no effect on LH release in vitro.

Animals↗

Immunocytochemical demonstration of PHI and its co-existence with VIP in intestinal nerves of the rat and pig.

Immunocytochemical studies of the rat and pig indicate that immunoreactivity for PHI (heptacosapeptide recently isolated by TATEMOTO and MUTT, 1981, from the porcine intestine) is located in numerous nerve fibers in the ileum and colon. Myenteric and submucosal plexuses contain abundant PHI-immunoreactive fibers. Alternate staining of adjacent sections with anti-PHI and anti-VIP sera indicates that PHI-immunoreactive nerves coincide with VIP nerves. This co-existence of both peptides in the same neurons may be accounted for by their probable sharing of a common precursor.

Animals↗

Applications of an automated apparatus for two-dimensional electrophoresis, Model TEP-1, for microsequence analyses of proteins.

Two-dimensional electrophoresis has become a rapid and powerful separation method for the preparation of samples for sequence analysis depending on the development of automated microsequencers accompanied by a blotting technique. We have constructed an automated apparatus for two-dimensional electrophoresis, Model TEP-1, which allows precise analytical runs under computer-control with a fully automated transfer from the first- to the second-dimensional gel. In the present paper we describe several applications of TEP-1 followed by blotting and sequencing to reveal the primary structure of proteins in a complex mixture. Several known and unknown proteins were analyzed. Enzymatic digests of proteins could be directly applied onto the TEP-1, in which a gel of lower molecular weight proteins was used as the second dimension. Subsequent blotting and sequencing provided internal sequence information. The use of TEP-1, followed by blotting, provides spots containing ca. 10-100 pmol protein. These figures permitted the identification of 10-40 amino acid residues from the N-terminus. The TEP-1 provides a simple and rapid separation of complex mixtures obtained from natural sources and chemical, biological or genetic products.

Amino Acid Sequence↗

Immunochemical study on PHI/PHM with use of synthetic peptides.

We have synthesized PHI and PHM (human PHI) as well as their fragments, PHI (1-6), PHI (1-15), PHI (14-19), PHI (14-27), PHI (20-27), PHM (1-15) and PHM (13-27), by the solution or solid-phase method for peptide synthesis. Using the highly purified synthetic peptides as immunogens or haptenic immunogens, five kinds of PHI/PHM specific antisera were produced. The major antibody-recognition sites of the five antisera were located respectively in the PHI C-terminal (R8201), in the PHI N-terminal (R8403), in the PHM C-terminal (R8502), and in the PHM whole molecule (R8702 and R8703). Radioimmunoassays (RIAs) with antisera R8201, R8403 and R8502, respectively, showed a wide distribution of immunoreactive (IR) PHI/PHM in porcine and human gastrointestinal and brain tissues. The concentrations of IR-PHI in the porcine gastrointestinal tissues, however, differed between the R8201 and R8403 RIAs employed for measurement. By using these two different PHI RIAs, the IR-PHI in the porcine brain tissue extract was shown to be almost a single component coeluting with synthetic PHI in gel filtration. The IR-PHI in the extract of porcine lower intestine on the other hand, contained, besides a PHI-like component, unidentified component(s) eluting immediately after synthetic PHI in gel filtration; this crossreacted with the PHI C-terminal specific R8201 antiserum but not with the N-terminal specific R8403 antiserum, suggesting the presence of the C-terminal-related fragment(s) of PHI in the tissues.

Animals↗

Rapid isolation and chemical synthesis of two porcine cardiodilatins, the cardiac hormone precursor and related fragment.

To evaluate the chemically synthesized materials, two cardiodilatins, CDD-126 and CDD-88/CDD(39-126), a precursor of atrial hormone and a related fragment, were isolated from porcine atria by immunoaffinity chromatography or alginic acid adsorption followed by an ion exchange high performance liquid chromatography. The chemical synthesis was carried out using an automated peptide synthesizer. After cleavage and refolding, the crude CDDs were directly characterized by the novel method of primary structure determination using electroblotting and microsequencing. The purified synthetic CDDs were identical with the natural ones in physicochemical and immunochemical properties, as well as their biological actions both in vivo and in vitro.

Amino Acid Sequence↗

Vasodilator effect of pituitary adenylate cyclase activating polypeptide (PACAP) on femoral blood flow in dogs.

The effect of PACAP, a new VIP-like neuropeptide, on femoral arterial blood flow was studied in anesthetized beagle dogs. Intra-arterial infusion of PACAP38, PACAP27, or VIP (0.25-25 pmol/kg) induced a dose-related increase in femoral blood flow. The dose-response curve for PACAP27 or PACAP38 was similar to that for VIP, but the response to PACAP38 at 25 pmol/kg was significantly less than the others. The vasodilator effect of PACAP38, however, lasted three to four times longer than that of PACAP27 or VIP. Vascular effects of PACAP38 and PACAP38(COOH) did not differ. PACAP(1-15), PACAP(10-20), PACAP(27-38), and PACAP(27-38)NH2 had little effect on blood flow. Thus, PACAP38 and PACAP27 are potent VIP-like vasodilators of the femoral arterial bed in dogs; PACAP38, but not PACAP27, differs from VIP in its prolonged vasodilator effect.

Animals↗

Development of pituitary adenylate cyclase activating polypeptides (PACAPs) specific radioimmunoassay systems and distribution of PACAP-like immunoreactivity in guinea pig tissues.

Pituitary adenylate cyclase activating polypeptide (PACAP) specific radioimmunoassay systems have been developed and the distribution of PACAPs in guinea pig tissues has been studied. The antibody against PACAP38 was characterized, using synthetic peptide fragments. It was shown to recognize the C-terminal portion with the C-terminal amide group and no cross-reaction was observed with vasoactive intestinal polypeptide (VIP), which has a high homology with PACAP27 and the N-terminal 28 amino acid residues of PACAP38. The antibody against PACAP27 was specific to PACAP27. With the two PACAPs (PACAP38 and PACAP27) specific RIA systems, high concentrations of PACAP38- and PACAP27-like immunoreactivity (LI) were observed in the brain of guinea pigs, especially in the diencephalon (mostly hypothalamus). In all tissues PACAP38-LI was higher than that of PACAP27-LI. Reverse-phase HPLC showed that PACAP38- and PACAP27-LI of tissue extracts were superimposed at the elution position of those of the synthetic peptides, respectively.

Adenylyl Cyclases↗