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Biomedical subjects

M Imawari

Publications and source records attributed to M Imawari.

At least 91 records · Page 5Linked to original sources
PubMed1982

A simple and sensitive assay for 25-hydroxyvitamin D, 24,25-dihydroxyvitamin D and 1,25-dihydroxyvitamin D in human serum.

An improved method is described which permits the simultaneous determination of 25-hydroxyvitamin D [25-(OH)D], 24,25-dihydroxyvitamin D [24,25-(OH)2D] and 1,25-dihydroxyvitamin D [1,25-(OH)2D] in milliliters of human serum. Methodological improvements enabled a rapid and almost complete extraction of the three metabolites from serum and omission of adding labeled internal standards to each serum sample for the calculation of individual recoveries. Commercially available stable chick embryo intestinal mucosa cytosol preparation made the troublesome preparation of cytosol receptor for 1,25-(OH)2D unnecessary. The procedure involves saturation of serum with ammonium carbonate and extraction with methanol/ethyl acetate, followed by separation of 25-(OH) D from the dihydroxy metabolites of vitamin D by Sephadex LH-20 column chromatography and further separation of the dihydroxy metabolites into 24,25-(OH)2D and 1,25-(OH)2D by high-pressure liquid chromatography. This is followed by individual determination of each metabolite by competitive protein-binding assay or radioreceptor assay.

24,25-Dihydroxyvitamin D 3↗
PubMed1982

Synthesis of serum and cytosol vitamin D-binding proteins by rat liver and kidney.

The synthesis of vitamin D-binding proteins in rat was examined using liver and kidney slices, isolated hepatocytes, and isolated renal tubules. Rat liver synthesized both serum vitamin D-binding protein (DBP) and a tissue DBP-binding component and secreted serum DBP. Rat kidney also synthesized a tissue DBP-binding component and cytosol vitamin D-binding protein (CDBP) which was immunologically related to serum DBP, but secretion of renal CDBP was not observed. Renal CDBP had a higher molecular weight (approximately 65,000) than that of serum DBP (54,000) and was heat-labile, while serum DBP was heat-stable (60 degrees C, 60 min). Renal CDBP was considered to exist in cytosol, forming a complex with a tissue DBP-binding component. These results indicate that liver is a site of synthesis of serum DBP and that CDBP, which is immunologically related to but physicochemically different from serum DBP, is synthesized in situ in kidney.

Animals↗
PubMed1980

Isolation and partial characterization of two immunologically similar vitamin D-binding proteins in rat serum.

Two immunologically similar, probably identical, binding proteins for vitamin D and its metabolites (DBP1 and DBP2) were isolated separately from rat serum after approximately 180-fold purification by novel procedures using Blue Sepharose CL-6B chromatography. The freshly purified DBP1 and DBP2 each showed a single band of protein on polyacrylamide gel electrophoresis, and had alpha-mobility, although DBP1 moved slightly faster than DBP2. DBP1 and DBP2 had the same molecular weight, which was estimated as approximately 54,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric points of DBP1 and DBP2 were estimated as 4.9 and 5.0, respectively, from the results of isoelectric focusing experiments. DBP1 and DBP2 both appeared to have one binding site for 25-hydroxyvitamin D3 per molecule of protein, with apparently similar association constants at 4 degrees C of 5--7 x 10(9) M-1. The amino acid compositions of DBP1 and DBP2 were also determined and compared. A monospecific antiserum against rat DBP2 was prepared in a rabbit and was used for immunological studies of rat DBP. On double immunodiffusion, anti-DBP2 antiserum produced precipitin lines of complete reaction-of-identity against the purified DBP1, the purified DBP2, and rat whole serum. There was no immunological cross-reactivity between rat DBP and sera from man, dog, and rabbit, but mouse serum showed a pattern of partial identity with rat DBP. When rat serum samples were analyzed by immunoelectrophoresis using anti-DBP antiserum, three patterns of precipitin line were observed: a pattern showing the existence of only DBP1, designated as DBP 1-1; a pattern showing the existence of only DBP2, designated as DBP 2-2; and a pattern showing the existence of both DBP1 and DBP2, designated as DBP 2-1. Using single radial immunodiffusion assay for rat serum DBP, the mean (+/- S.D.) serum DBP concentrations were found to be 461 +/- 59 microgram/ml in adult male rats and 328 +/- 16 microgram/ml in adult female rats, and the difference was significant (p < 0.001). In molar terms, DBPs are present in normal rat serum in large excess relative to vitamin D and its metabolites, and most of the serum DBP, therefore, circulates as apo-DBP, not containing a bound molecule of vitamin D or of its metabolites. The immunoprecipitation studies of DBP in rat serum showed that DBPs were common main transport proteins for naturally occurring vitamin D and its metabolites, and that DBP played some, but not a principal, role in the transport of synthetic 1 alpha-hydroxyvitamin D3.

Amino Acids↗
PubMed1980

The effect of vitamin D2 on hypocalcemia in patients under chronic hemodialysis.

The effects of vitamin D2 (VD2) on hypocalcemia were studied in 54 hypocalcemic patients (29 males and 25 females) on chronic hemodialysis. Calcium lactate (3 g/day) or VD2 (10,000 IU/day, 50,000 IU/day and 80,000 IU/day) were administered for 4 months in order to correct the hypocalcemia. Serum calcium, phosphate and alkaline phosphatase levels were measured and the effects of VD2 on these parameters of calcium metabolism were followed. 1) Calcium lactate or 10,000 IU/day of VD2 were not effective for the correction of hypocalcemia, while 50,000-80,000 IU/day of VD2 were effective. The effects of VD2 on serum calcium concentrations were dose-dependent, and the normalization of serum calcium concentrations was achieved more rapidly with higher doses of VD2. However, in the group treated with 80,000 IU/day of VD2, many patients developed hypercalcemia, but in the group treated with 50,000 IU/day of VD2, only a few patients did it. From these results, suitable dose (initial and maintenance doses) of VD2 in dialysed patients would be 50,000 IU/day. 2) When the responder group (normal serum calcium levels after 4 months of treatment with 50,000 IU/day of VD2) and the non-responder group serum calcium levels lower than 4.2 mEq/liter on the same condition) were compared, the durations of dialysis were significantly shorter in the former than those in the latter. This fact may suggest that the effects of VD2 administration on hypocalcemia in dialysed patients are partly dependent on the residual renal function concerning the conversion of 25-OH-D3 into 1,25 (OH)2D3.

Adolescent↗
PubMed1980

Serum 25-hydroxyvitamin D and vitamin D-binding protein levels and mineral metabolism after partial and total gastrectomy.

The effects of gastrectomy, especially total gastrectomy, on the serum levels of 25-hydroxyvitamin D and vitamin D-binding protein and on mineral metabolism were examined. The serum 25-hydroxy-vitamin D levels were markedly decreased in patients with total gastrectomy and Billroth II gastrectomy. Decreased levels of serum vitamin D-binding protein and serum calcium, and increased levels of serum alkaline phosphatase were observed in both patients with partial gastrectomy and patients with total gastrectomy. The results show that vitamin D deficiency could develop in high frequency patients with total gastrectomy and Billroth II gastrectomy, and that deranged mineral metabolism could develop in patients with any type of gastrectomy with or without vitamin D deficiency. The decreased levels of serum vitamin D-binding protein in postgastrectomy patients may be a sensitive reflection of the failure of hepatic protein synthesis.

Aged↗
PubMed1979

Quantitative studies of the interaction of cholecalciferol ((vitamin D3) and its metabolites with different genetic variants of the serum binding protein for these sterols.

Cholecalciferol (vitamin D3) and its 25-hydroxy metabolite are transported in plasma bound to a specific protein, the binding protein for cholecalciferol and its metabolites (DBP). DBP is identical with the group-specific component (Gc) proteins, which are known to display genetic polymorphism. Studies were conducted to explore whether or not major differences in the transport of cholecalciferol and its biological metabolites might exist among persons with different Gc phenotypes. Detailed quantitative studies were first carried out on the interaction of 25(OH)D3 with DBP in 21 different samples of serum, representing eight different Gc phenotypes. The studies used a filter disc assay method that provided highly reproducible quantitative results with cholecalciferol-related sterols. The Gc phenotypes studied included the three common types (Gc 1-1, 2-1, and 2-2) and several uncommon genetic variants (Gc Ab-Ab, Ab-1, Ab-2, Chip-1, and Chip-2). The binding affinities for 25(OH)D3 observed with these different sera were all fairly similar to each other. More extensive studies were then conducted to compare the binding of four cholecalciferol-related sterols to each of three genetic variants of DBP, by using sera from homozygous persons with the Gc 1-1, Gc 2-2 and Gc Ab-Ab phenotypes. The ligands tested included cholecalciferol, 25(OH)D3, 1,25(OH)2D3, and 24(R) 25(OH)2D3. The affinities of the three genetic types of DBP/Gc protein were found to be similar for each of the four cholecalciferol-related sterols. The apparent association constants for 25(OH)D3 and 24,25(OH)2D3 were similar (approx. 1--2 x 10(8) M-1); lesser affinities were observed for 1,25(OH)2D3 (kA approx. 1 x 10(7) M-1) and for cholecalciferol (kA approx. 3--4 x 10(5) M-1). Thus the common genetic variants of DBP/Gc protein, and the uncommon genetic variants studied here, all appear to have similar binding properties for cholecalciferol and its several metabolites.

Alpha-Globulins↗
PubMed1977

Immunological and immunoassay studies of the binding protein for vitamin D and its metabolites in human serum.

This study reports the development of a specific and sensitive radioimmunoassay and a simple and accurate radial immunodiffusion (RID) assay for the human serum-binding protein for vitamin D and its metabolites (DBP). These immunoassays employed a monospecific antiserum that was prepared in rabbits against human DBP. The radioimmunoassay effectively measured DBP in amounts of 1-10 ng, whereas the RID assay measured DBP accurately in amounts of 0.2-0.8 mug. The results obtained with the two immunoassays on the same samples of serum agreed well with each other. Using the RID assay, the mean (+/- SD) serum DBP concentration observed in 35 normal persons was 422 +/- 27 micrograms/ml. Generally similar levels were observed in 66 hyperlipidemic subjects. In molar terms, the mean DBP concentration (approximately 8 microgramsM) was of the order of 50 times the usual serum level of 25-hydroxyvitamin D (25-OH-D) plus vitamin D. Thus, most of plasma DBP circulates as apo-DBP, not containing a bound molecule of 25-OH-D or of vitamin D. DBP and 25-OH-D concentrations were measured in a limited number of patients with hypercalcemia, mild hypocalcemia, and markedly elevated serum 25-OH-D levels due to oral vitamin D supplementation. It was found that major changes can occur in the serum levels of 25-OH-D and of calcium with very little or no associated changes occurring in the serum concentration of DBP, The results suggest that neither serum 25-OH-D nor serum calcium plays an important role in the regulation of the metabolism of DBP. Data were obtained that confirmed and extended an earlier report on the identity of the group-specific component (Gc) protein in plasma with the plasma vitamin D-binding protein. On immunodiffusion against whole serum, the line formed with the anti-DBP antiserum showed a complete reaction-of-identity with the line formed with commercial antiserum against Gc protein. Furthermore, serum that had been depleted of DBP by treatment with Sepharose containing covalently coupled antibodies against DBP was found to be depleted also of immunoreactivity against anti-GC protein antiserum. In addition, the properties of the purified DBP preparation agreed closely with those previously reported by others for Gc protein. Finally, a comparative immunology study showed that sera from several different mammalian orders showed some immunoreactivity against the antihuman DBP antiserum. Thus, proteins immunologically similar to human DBP are present in sera from a number of mammalian species and orders.

Calcium↗
PubMed1976

The transport of vitamin D and its 25-hydroxy metabolite in human plasma. Isolation and partial characterization of vitamin D and 25-hydroxyvitamin D binding protein.

This study reports the isolation and partial characterization of vitamin D and 25-hydroxyvitamin D binding protein (DBP), the specific transport protein for vitamin D and its 25-hydroxy metabolite in human plasma. DBP was labeled by the addition of a tracer amount of 3H-labeled 25-OH-D3 to the original plasma used for protein fractionation. Previous experiments have shown that such 25-OH-D3 added in vitro binds to the same protein normally responsible for the transport of endogenous 25-OH-D and of vitamin D. The isolation of human DBP was achieved by an extensive sequence of procedures which resulted in a final yield of only approximately 4 mg of purified DBP from a starting volume of 34 liters of plasma. Purified DBP was homogeneous in the analytical ultracentrifuge and showed a single band of protein on analytical polyacrylamide gel electrophoresis. DBP had a sedimentation constant of 3.49s and a mol wt of approximately 52,000. The molecular weight was assessed by sedimentation equilibrium analysis and also by sodium dodecyl sulfate-disc-gel electrophoresis and by gel filtration on a standardized column of Sephadex G-150. The amino acid composition of DBP was determined and was generally consistent with the estimated extinction coefficient (E1cm1% at 280 nm) of about 9.1. The isoelectric point of DBP was estimated as 4.8 from isoelectric focusing experiments. Direct study of the binding capacity of the purified DBP for added 25-OH-D3 showed that the isolated DBP had a high affinity for 25-OH-D3, with an apparent maximum binding capacity of one molecule of 25-OH-D3 per molecule of protein.

Amino Acids↗
PubMedDate not supplied

Androgen and estrogen receptors in hepatocellular carcinoma and in the surrounding noncancerous liver tissue.

Both androgen and estrogen receptors were studied in human hepatocellular carcinoma and noncancerous liver tissue surrounding it. Androgen receptor was detected in the cytosol and/or nucleosol of 4 of 8 cancerous tissues and 1 of 6 noncancerous tissues. The levels of androgen receptor in hepatocellular carcinomas ranged from 3.4 to 37.6 fmoles per mg protein with dissociation constants (Kd) of 0.226 - 51.3 X 10(-9) M. That in the surrounding noncancerous tissue was 2.1 fmoles per mg protein with Kd of 0.941 X 10(-9) M. Estrogen receptor was detected in the cytosol of 1 of 7 cancerous tissues, while it was detected in the cytosol and/or nucleosol of 3 of 7 noncancerous tissues. The level of estrogen receptor in hepatocellular carcinoma was 4.9 fmoles per mg protein with Kd of 1.20 X 10(-9) M, and those in the surrounding noncancerous tissues ranged from 2.6 to 1,073 fmoles per mg protein with Kd of 0.223 - 3.15 X 10(-9) M. The results suggest that the expression of androgen receptor may be augmented in association with malignant transformation of hepatocytes while the expression of estrogen receptor may be rather suppressed and that some of hepatocellular carcinomas may be androgen-dependent.

Adult↗
PubMedDate not supplied

Protective effects of fibronectin in galactosamine-induced liver failure in rats.

The effects of supplementation with fibronectin on liver damage and survival in rats with galactosamine-induced liver failure were studied. In rats with acute liver failure induced by a low dose of galactosamine, supplementation with purified plasma fibronectin at 3 hr after the administration of galactosamine provided significant increase of plasma fibronectin levels and augmentation of reticuloendothelial system function at 4 hr, significantly higher plasma fibronectin levels and significant protection of liver damage with shorter prothrombin times, lower AST and less histological damage at 48 hr as compared to control animals. Plasma fibronectin levels were inversely correlated with both plasma prothrombin times and AST. Fibronectin supplementation at 6 hr also resulted in the significant decrease of liver damage at 48 hr as evaluated histologically. When rats with liver failure, induced by a high dose of galactosamine, were supplemented with fibronectin at 3 hr, the survival rate was significantly higher than that of control rats. The results indicate that fibronectin supplementation in the early stages of acute liver failure could reduce liver damage and improve the survival of rats with galactosamine-induced liver failure.

Acute Disease↗