Larval bullfrog skin lacks amiloride-blockable epithelial transport because α-ENaC is located within intracellular vesicles in epidermal apical cells and not in the apical plasma membrane.

The epithelial Na channel (ENaC) plays an essential role in sodium transport across epithelia such as adult frog skin. Transport across the skin, measured as short-circuit current (SCC), is blocked by amiloride. Bullfrog alpha-ENaC (α-fENaC) is expressed in adult bullfrog skin, and the SCC across this skin is blocked by amiloride.

Localization of ENaC subunit mRNAs in adult bullfrog skin.

Adult amphibian skin has served as a model for the investigation of Na(+)-transporting epithelia, such as mammalian renal tubules. The amiloride-blockable epithelial Na(+) channel (ENaC), which is located in the apical membrane of the outer living cell layer, regulates Na(+) transport across the epithelium. ENaC is thought to develop during the terminal differentiation of epidermal cells, but the details are unclear.

Vasotocin- and mesotocin-induced increases in short-circuit current across tree frog skin.

In adult amphibian skin, Na(+) crosses from outside to inside. This Na(+) transport can be measured as the amiloride-blockable short-circuit current (SCC) across the skin. We investigated the effects of arginine vasotocin (AVT) and mesotocin (MT), and those of antagonists of the vasopressin and oxytocin receptors, on the SCC across Hyla japonica skin.

Effects of arginine vasotocin and mesotocin on the activation and development of amiloride-blockable short-circuit current across larval, adult, and cultured larval bullfrog skins.

Amphibian skin has osmoregulatory functions, with Na(+) crossing from outside to inside. Na(+) transport can be measured as the short-circuit current (SCC). We investigated the short-term and long-term effects of arginine vasotocin (AVT) and mesotocin (MT) (which modulate Na(+) transport) on the activation and development of an amiloride-blockable SCC (adult-type feature) in larval, adult, and corticoid-cultured larval bullfrog skins.

Lysophosphatidylcholine for efficient intestinal lipid absorption and lipoprotein secretion in caco-2 cells.

Phosphatidylcholine (PC) and its hydrolysates are considered to stimulate intestinal lipid absorption, however, their exact effects on lipoproteins and apolipoprotein (apo) metabolism remain ambiguous. This study aimed to further differentiate the effects of them using fully differentiated enterocyte-like Caco-2 cells. Lipid micelles (oleic acid 0.

A possible role of lysophospholipids produced by calcium-independent phospholipase A(2) in membrane-raft budding and fission.

Phospholipase A(2) (PLA(2)) not only plays a role in the membrane vesiculation system but also mediates membrane-raft budding and fission in artificial giant liposomes. This study aimed to demonstrate the same effects in living cells. Differentiated Caco-2 cells were cultured on filter membranes.

Vasotocin has the potential to inhibit basolateral Na(+)/K (+)-pump current across isolated skin of tree frog in vitro, via its V(2)-type receptor/cAMP pathway.

Adult frog skin transports Na(+) from the apical to the basolateral side across the skin. Antidiuretic hormone (ADH) is involved in the regulation of Na(+) transport in both mammals and amphibians. We investigated the effect of arginine vasotocin (AVT), the ADH of amphibians, on the short-circuit current (SCC) across intact skin and on the basolateral Na(+)/K(+)-pump current across apically nystatin-permeabilized skin of the tree frog, Hyla japonica, in which the V(2)-type ADH receptor is expressed in vitro.

Prolactin increases Na+ transport across adult bullfrog skin via stimulation of both ENaC and Na+/K+-pump.

PRL is involved in osmoregulation in lower vertebrates. Its serum concentration starts to increase during the metamorphosis of bullfrog tadpoles. Adult bullfrog skin transports Na(+) from the apical to the basolateral side across the skin.

Disruption of the murine intestinal alkaline phosphatase gene Akp3 impairs lipid transcytosis and induces visceral fat accumulation and hepatic steatosis.

Intestinal alkaline phosphatase (IAP) is involved in the process of fat absorption, a conclusion confirmed by an altered lipid transport and a faster body weight gain from 10 to 30 wk in both male and female mice with a homozygous null mutation of the IAP coding gene (Akp3(-/-) mice). This study was aimed to delineate morphologically and quantitatively the accelerated lipid absorption in male Akp3(-/-) mice. Feeding a corn oil bolus produced an earlier peak of triacylglycerol in serum (2 vs.

Larval bullfrog skin expresses ENaC despite having no amiloride-blockable transepithelial Na+ transport.

Amiloride-blockable Na(+) transport, measured as an amiloride-blockable short-circuit current (Am-SCC), is mediated by the epithelial Na(+) channel (ENaC). Am-SCC is not normally present in bullfrog tadpole skin, but when such skin is cultured with corticoids an amiloride-blockable Na transport appears. Prolactin (PRL) inhibits its corticoid-induced development.

Changes in receptor activator of nuclear factor-kappaB, and its ligand, osteoprotegerin, bone-type alkaline phosphatase, and tartrate-resistant acid phosphatase in ovariectomized rats.

We investigated time-course changes in the expression of receptor activator of nuclear factor-kappaB (RANK), its ligand (RANKL), osteoprotegerin (OPG), bone-type alkaline phosphatase (BAP), and tartrate-resistant acid phosphatase (TRAP) in ovariectomized (OVX) rats. Samples of sera and coccyges were used for analysis of the enzyme activities and expression levels of proteins and mRNAs, and an immunohistochemical analysis was also performed. Serum BAP activity increased to 158.

Ambroxol reduces LPS toxicity mediated by induction of alkaline phosphatases in rat lung.

Alkaline phosphatases (APs) have been suggested to detoxify lipopolysaccharide (LPS) by dephosphorylation. Ambroxol, a bronchial expectorant, is known to accelerate the secretion of pulmonary surfactant particles including AP molecules as a pharmacological action. In the present study, some beneficial effects of ambroxol on LPS toxicity in the rat lung were investigated.

Characterization of the epitopes specific for the monoclonal antibody 9F5-3a and quantification of oxidized HDL in human plasma.

Background: We previously isolated a monoclonal antibody, 9F5-3a, that is specific for HDL oxidized by CuSO(4).

Methods: We examined the characteristics of the 9F5-3a epitope by Western blot and measured the concentration of oxidized HDL in human plasma by enzyme-linked immunosorbent assay using this antibody.

Results: The monoclonal antibody specifically reacted with oxidized HDL in a mixture of HDL, LDL and modified lipoproteins.

Expression of alpha-amylase isozymes in rat tissues.

Gene expressions of alpha-amylase isozymes in rat tissues were analyzed by a reverse transcription-polymerase chain reaction (RT-PCR), followed by EcoRI digestion. This procedure is based on evidence that an RT-PCR product from mouse pancreas RNA is sensitive to EcoRI, but not the product from the salivary gland or liver RNAs. The method was applied to the analysis of alpha-amylase expression in rat liver after partial hepatectomy, in which a potent expression of pancreas type isozyme was observed.

NF-kappa B activation in endothelial cells treated with oxidized high-density lipoprotein.

We first determined whether oxidized high-density lipoprotein (ox-HDL) activates transcription factor nuclear factor-kappa B (NF-kappa B) in cultured human umbilical vein endothelial cells (HUVECs). Treatment for 7h with 100 microg/ml ox-HDL elicited a marked downregulation of I kappa B alpha and upregulation of the phosphorylated form of I kappa B alpha in HUVECs in a manner dependent on the dose of ox-HDL. Electrophoretic mobility shift assay in nuclear fraction from HUVECs showed translocation of NF-kappa B to the nucleus and binding of NF-kappa B to NF-kappa B consensus oligonucleotides during ox-HDL exposure for 7h, suggesting that ox-HDL brings about NF-kappa B activation in endothelial cells.

Rabbit liver dCMP phosphohydrolase: a pyrimidine 5'-nucleotidase I-like enzyme in non-erythrocytic cells.

A nucleotide phosphomonoesterase activity that preferably hydrolyzed dCMP was detected in rabbit liver and purified approximately 20-fold. The enzyme was similar in the catalytic and molecular properties to pyrimidine 5'-nucleotidase subclass I (P5N-I), which distributed specifically in vertebrate erythrocytes. In addition to liver, the activity was found in rabbit kidney, spleen, heart, intestine, but was not detected in any rat or chicken tissues tested.

Glycated high-density lipoprotein regulates reactive oxygen species and reactive nitrogen species in endothelial cells.

Nonenzymatic glycosylation of plasma proteins may contribute to the excess risk of developing atherosclerosis in patients with diabetes mellitus. Although it is believed that high-density lipoprotein (HDL) is glycosylated at an increased level in diabetic individuals, little is known about a possible linkage between glycated HDL and endothelial dysfunction in diabetes. To clarify whether glucose-modified HDL affects the function of endothelial cells, we first examined herein the level of H(2)O(2) generation from cultured human aortic endothelial cells (HAECs) exposed to a glycated oxidized HDL (gly-ox-HDL) prepared in vitro.

Detection of oxidized high-density lipoprotein.

This paper reviews working procedures for the separation and detection of oxidized high-density lipoproteins (ox-HDL) and their constituents. It begins with an introductory overview of structural alterations of the HDL particle and its constituents generated during oxidation. The main body of the review delineates various procedures for the isolation and detection of ox-HDL as well as the purification and separation of phosphatidylcholine metabolites and denatured apolipoproteins in the particle.

Alkaline phosphatases reduce toxicity of lipopolysaccharides in vivo and in vitro through dephosphorylation.

Intestinal alkaline phosphatase (AP), as a host defense factor, was first investigated in vivo using rats orally exposed to lipopolysaccharide (LPS). After the oral administration of LPS to rats, serum LPS content was increased within 2 hr and then decreased to 6 hr. In contrast, when L-phenylalanine (L-Phe), an inhibitor of intestinal-type AP isozymes, was simultaneously administered with LPS, serum LPS content significantly increased from 1 hr and the area under the concentration-time curve of serum LPS was augmented approximately 2-fold, suggesting that APs in the gastrointestinal tract reduced serum LPS content.

Electrophoretically unique amylases in rat livers: phylogenic and ontogenic study on the mammalian liver.

Liver amylase activity in rodents was assayed with Blue Starch as substrate, and found to be higher than in humans or pigs. Based on the result of concanavalin A affinity chromatography, we found that the sugar moieties of amylase molecules increased in parallel with amylase activity in the tested mammals. However, the amounts of amylase proteins determined by Western bloting with anti-human salivary-type antibody as the probe, were similar to the levels in mammalian livers.

Characterization of four monoclonal antibodies to recombinant human tartrate-resistant acid phosphatase.

In this study we produced a recombinant human Tartrate-resistant acid phosphatase (TRAP) enzyme from baculovirus-infected insect cells, generated four monoclonal antibodies (MAbs) 15A4, 13B9, 1C6 and 3G7, to the enzyme, and characterized these antibodies. In the human serum and lung specimen, all four antibodies appeared to have a high specificity for native TRAP enzyme in western blot analysis, immunohistochemical analysis and enzyme immunoassay. These antibodies may react with respective conformational determinants, therefore, they may be useful for detection of active TRAP.

A restriction endonuclease assay for expression of human alpha-amylase isozymes.

Background: The alpha-amylase isozymes can be detected separately by electrophoresis; however, sometimes the identification is difficult because of their microheterogeneity. In the present study, we tried to establish a convenient method for the detection of alpha-amylase isozyme expression.

Methods: The procedure is based on three different restriction sites presented in those genes; a PstI site in both AMY 2A and 2B genes, a HaeII site in both AMY 1 and 2A genes, and a BamHI site in AMY 2B gene.

Altered bone turnover in chlorpromazine-challenged rats and its effect on 1alpha-hydroxyvitamin D3 administration in vivo.

We reported previously that Ca and Pi levels are elevated and alkaline phosphatase (ALP) activity and 1alpha,25(OH)2D3 levels are reduced in chlorpromazine (CPZ)-challenged rats. In the present study, we determined the serum levels of interleukin (IL)-6 and acid phosphatase (ACP) in CPZ-challenged rats, in addition to levels of ALP protein. ALP mRNA and coccyx morphology were examined in CPZ-challenged rats as well as the effect of CPZ on 1alpha(OH)D3 production in vivo.