Biomedical subjects

T Akino

Publications and source records attributed to T Akino.

At least 73 records · Page 4Linked to original sources

PubMed1993

Beta-arrestin and arrestin are recognized by autoantibodies in sera from multiple sclerosis patients.

Multiple sclerosis (MS), one of the most common chronic neurologic diseases, is characterized by the presence of multiple plaques of demyelination throughout the central nervous system. Although the etiology of the disease has not been established, it is believed to involve autoimmune mechanisms. We have examined sera from patients with MS for the presence of antibodies to antigens from brain and retina. Immunoblot analysis of soluble fraction of proteins from bovine brain revealed a prominent band at 45 kDa stained with sera of 8-14 patients with MS. In two patients with MS, serum antibody titers during relapse were higher compared with those when the patients were in remission. These antibodies were undetectable in cerebrospinal fluid of our MS patients and additionally were absent in sera of patients with other neurological diseases and normal control subjects. Furthermore, immunoblot analysis of the soluble fraction from bovine retinal rod outer segments revealed a prominent protein band at 48 kDa stained with MS sera. This antigen was purified to homogeneity from bovine retinal outer segments and identified as arrestin. Additionally, sera from MS patients reacted with purified beta-arrestin 1, a 45-kDa protein homologous to arrestin that is found in various tissues. Using limited proteolysis of arrestin and a competitive ELISA test with a synthetic peptide, we identified the recognition site(s) for antibodies in sera of MS patients at a dominant immunogenic site on arrestin located at the C-terminal region of the molecule. We suggest that the presence of circulating antibodies reactive with beta-arrestin or arrestin may be related to the course of MS progression.

Amino Acid Sequence↗

PubMed1993

Role of the C-terminal domain of pulmonary surfactant protein A in binding to alveolar type II cells and regulation of phospholipid secretion.

Surfactant protein A (SP-A), with a reduced denatured molecular mass of 26-38 kDa, is characterized by a collagen-like sequence in the N-terminal half of the protein. This protein forms an oligomeric structure which is dependent upon this collagenous domain. SP-A has been demonstrated to function as an inhibitor of phospholipid secretion by primary cultures of alveolar type II cells via a cell surface receptor for the protein. However, the receptor-binding domain of SP-A has not been identified. The purpose of the present study was to investigate the role of the C-terminal domain of SP-A in binding to type II cells and regulation of phospholipid secretion. A monoclonal antibody to human SP-A, whose epitope was localized at the C-terminal domain of the protein, abolished the inhibitory activity of human SP-A on lipid secretion by type II cells, and attenuated the ability of human SP-A to compete with 125I-(rat SP-A) for receptor binding. SP-A was then digested with collagenase and the collagenase-resistant fragment (CRF), which is the C-terminal domain of SP-A (thus lacking the N-terminal domain), was isolated. Gel filtration chromatography revealed that CRF exists as a monomer in solution containing Ca2+. CRF had the ability to inhibit phospholipid secretion, although at a higher concentration than for SP-A, and was also able to compete with 125I-(rat SP-A) for binding to type II cells. A direct binding study showed that CRF bound to type II cells in a concentration-dependent manner. The present study demonstrates that the non-collagenous, C-terminal, domain of SP-A is responsible for the protein's inhibitory effect on lipid secretion and its binding to type II cells.

Amino Acid Sequence↗

PubMed1993

Purification and characterization of three MEKA-like proteins in liver: association of a 94 kDa protein with beta gamma subunits of G-proteins.

Retinal 32 kDa MEKA protein (rMEKA) exists in the photoreceptor cells and forms a complex with beta gamma subunit of transducin. Bovine liver contained three MEKA-like proteins (94 kDa, 35 kDa-a, 35 kDa-b) which reacted with a rMEKA antibody. Each protein was purified as a single band on a SDS-PAGE and used for a reconstitution experiment with alpha and beta gamma subunits of cerebral G-proteins (Go/i). The 94 kDa protein inhibited GTP-binding ability of G alpha by forming a complex with beta gamma subunit.

Animals↗

PubMed1993

Identification of a novel gamma-subunit from bovine brain GTP binding regulatory proteins (Gi/o).

Heterogeneity of the gamma-subunit of G proteins has been demonstrated by cDNA cloning and by partial sequence analyses. We have isolated two intact beta gamma-subunit isoforms from bovine brain Gi/o mixture, in which only gamma subunits are distinct (Sohma, H., et al. (1992) Biochem. Biophys. Res. Commun. 184, 175-182). In this study, we isolated the gamma-subunit isoforms, gamma-I and gamma-II, and examined their amino acid sequences. Both gamma-I and gamma-II had blocked N-terminal amino acid residues, and the terminal amino acids of both were able to be truncated by an acylamino-acid-releasing enzyme. Gamma-I seemed to be identical with the gamma-subunit reported elsewhere, while the gamma-II appeared to be a novel protein. Antibodies to synthetic peptides based on the part of the amino acid sequences of gamma-I and gamma-II reacted specifically to gamma-I and gamma-II, respectively.

Amino Acid Sequence↗

PubMed1993

Results of vascularized joint allograft under immunosuppression with cyclosporine.

The effects of cyclosporine (CsA), a strong immunosuppressive drug, on vascularized allogeneic joint transplantations were examined. An orthotopical transplant model of a vascularized knee joint allograft was developed using inbred DA and Lewis rats to investigate the fate of grafts following withdrawal of short-term immunosuppression compared to continuous immunosuppression with CsA. Five isograft controls acquired solid bone union at both femur and tibia sites within 4 weeks, and joint function as skeletal support was maintained until 25 weeks. Without immunosuppression, ten allografts were severely rejected within the first week, and joint destruction occurred immediately. Twenty-five short-term immunosuppressed rats acquired solid union, but, after withdrawal of immunosuppression, grafted joints showed gradual rejection and were destroyed due to pathological fractures or joint instability, although partial revascularization from the recipient occurred. Ten allografts under continuous immunosuppression at a dose of 10 mg/kg/day showed no rejection and remained viable for 12 weeks postoperatively, but thereafter all rats died. Death was considered to be a side effect of CsA. Fifteen animals, under continuous immunosuppression at a dose of 5 mg/kg/day, showed no rejection except in the bone marrow; the grafted joint function was not effected until 25 weeks. Continuous treatment with low and nontoxic doses (5 mg/kg/day) of CsA was necessary to maintain the functions of the grafted joint.

Animals↗

PubMed1993

Pulmonary surfactant protein A-mediated uptake of phosphatidylcholine by alveolar type II cells.

Pulmonary surfactant protein A (SP-A)-mediated uptake of phosphatidylcholine (PC) by alveolar type II cells was investigated. SP-A enhanced the uptake of liposomes containing dipalmitoylphosphatidylcholine (DPPC), 1-palmitoyl-2-linoleoyl phosphatidylcholine (PLPC), or 1,2-dihexadecyl-sn-glycero-3-phosphocholine (DPPC-ether), a diether analogue of DPPC, but about twice as much DPPC was taken up by type II cells as PLPC or DPPC-ether. When subcellular distribution was analyzed, 51.3 +/- 2.9% (mean +/- SD, n = 3) of cell-associated radiolabeled DPPC was recovered in the lamellar body-rich fraction in the presence of SP-A, whereas only 19.3 +/- 1.9% (mean +/- SD, n = 3) was found to this fraction in the absence of SP-A. When type II cells were incubated either with DPPC at 0 degree C or with DPPC-ether at 37 degrees C, or no cells were included, low proportions of the cell-associated lipids were present in the fractions corresponding to lamellar bodies even in the presence of SP-A. Anti-SP-A antibody significantly reduced the radioactivity incorporated into the lamellar body fraction. Phosphatidylcholine that had been incorporated into lamellar bodies remained largely intact when SP-A was present. Subcellular fractionations of type II cells with radiolabeled SP-A and DPPC revealed that the sedimentation characteristics of cell-associated SP-A are different from those of DPPC, although a small broad peak of radiolabeled SP-A was found in the lamellar body fraction.(ABSTRACT TRUNCATED AT 250 WORDS)

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1993

Elevated levels of lung surfactant protein A in sera from patients with idiopathic pulmonary fibrosis and pulmonary alveolar proteinosis.

An enzyme-linked immunosorbent assay using monoclonal antibodies to human lung surfactant protein A (SP-A) was applied to sera from patients with lung diseases. We examined whether SP-A appears in the sera of patients with diseases that are known to cause alterations in surfactant composition in bronchoalveolar lavage fluids, and we characterized the SP-A that was found. The level of SP-A in sera from 57 healthy volunteers was 45 +/- 3 ng/ml (mean +/- SEM). The levels in patients with idiopathic pulmonary fibrosis (IPF) (205 +/- 23 ng/ml, n = 32) and pulmonary alveolar proteinosis (PAP) (285 +/- 23 ng/ml, n = 6) were significantly higher than those in healthy control subjects (p < 0.01), whereas those of sarcoidosis (n = 16), pneumonia (n = 14), and tuberculosis (n = 14) were 52 +/- 27 ng/ml, 65 +/- 11 ng/ml, and 49 +/- 23 ng/ml, respectively. Electrophoresis and immunoblotting analysis demonstrated that the fraction isolated from serum of a patient with PAP or IPF by anti-SP-A immunoaffinity column chromatography consisted chiefly of human IgG and IgM, and that it also contained SP-A. Furthermore, IgG was found in preparation of purified human SP-A. SP-A was demonstrated to bind to nonimmune IgG coated onto microtiter wells. Gel filtration analysis revealed that serum SP-A was eluted at fractions of larger molecular size than was the purified SP-A. These findings suggest that SP-A appears in the bloodstream as a complex with immunoglobulin in IPF and in PAP.

Aged↗

PubMed1993

A study of the binding of immunoglobulin G and immunoglobulin E from children with bronchial asthma to peptides derived from group II antigen of Dermatophagoides pteronyssinus.

The peptides that were produced by the treatment of the purified group II antigen of Dermatophagoides pteronyssinus with cyanogen bromide, lysylendopeptidase, Staphylococcus aureus V8 protease, or N-tosyl-L-phenylalanylchloromethyl ketone-treated trypsin were examined for their binding with IgG and IgE from children with bronchial asthma by immunoblotting analysis. Both IgG and IgE bound to the peptides from the amino terminus to the 76th residue and the carboxyl terminal side from the 13th or 15th residue; on the other hand, both Ig failed to bind to the carboxyl terminal side from the 39th, 56th, or 77th residue. The data demonstrated that peptides lacking residues 1 to 15 possessed the ability to bind Ig, whereas those lacking residues 1 to 39 had lost it. The present study indicates that the amino terminal half of the group II antigen of Dermatophagoides pteronyssinus is more important for the binding of IgG and IgE than the carboxyl terminal half and suggests that the residues 15 to 39 may be directly related to the binding of both IgG and IgE.

Adolescent↗

PubMed1993

Surfactant protein-A concentration in bronchoalveolar lavage fluids of patients with pulmonary alveolar proteinosis.

Pulmonary alveolar proteinosis (PAP) is characterized by accumulation of large quantities of lipoproteinaceous materials in alveoli. Surfactant protein A (SP-A) is the predominant phospholipid-associated glycoprotein in pulmonary surfactant and is specific to the lung. The contents of SP-A in bronchoalveolar lavage (BAL) fluids of patients with PAP were measured with an enzyme-linked immunosorbent assay using two monoclonal antibodies to human SP-A to evaluate its usefulness for diagnosis. Concentration of SP-A in BAL fluid in PAP was significantly increased in comparison with that of normal volunteers. The ratio of SP-A to protein in BAL fluid of PAP was at almost the same level as in normal subjects, while the ratio of SP-A to phospholipid in PAP was significantly higher. These results indicate that measurement of BAL fluid SP-A is of clinical value for diagnosis of PAP and should be used as a biochemical diagnostic tool in the clinical laboratory.

Adult↗

PubMed1993

Calcium and dithiothreitol dependent conformational changes in beta-sheet structure of collagenase resistant fragment of human surfactant protein A.

Ca2+ dependent conformational change of collagenase resistant fragment (CRF) of human surfactant protein A (SP-A) was studied by measurements of the far UV circular dichroism spectrum. The spectrum was altered by Ca2+ and DTT. The beta-sheet content was decreased by the addition of Ca2+ from 28.1 to 26.6%. On the other hand, the beta-sheet content was increased in the presence of dithiothreitol from 28.1 to 36.0%, and decreased by the addition of Ca2+ from 36.0 to 30.5%. The total Ca2+ concentration required for half maximal change of the ellipticity at 220 nm was estimated to be 30 microM both in the presence and absence of dithiothreitol. One of the functions of SP-A, enhancement of phospholipid uptake by alveolar type II cells, was abolished by the addition of 2-mercaptoethanol. These results strongly indicate a relationship between the conformation of CRF and SP-A functions.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1993

Lung adenocarcinoma with type II pneumocyte characteristics.

We report a case of primary lung adenocarcinoma with type II pneumocyte characteristics. Electron microscopic examination demonstrated that the tumour cells had well-developed microvilli and cytoplasmic lamellar inclusion bodies. These ultrastructural features are similar to those seen in type II pneumocytes of normal lung tissue. Western blot analysis, using monoclonal antibody against human surfactant protein A (SP-A), clearly demonstrated that the tumour cells expressed human SP-A, which is a major pulmonary surfactant protein produced by type II pneumocytes. These observations suggest that the tumour was derived from a type II pneumocyte.

Adenocarcinoma↗

PubMed1993

[Light-induced dephosphorylation of phosphoproteins in rod outer segments in bovine and frog retina].

Several lines of evidence have suggested that protein phosphorylation and dephosphorylation may play an important role in the regulation of metabolism and signal transduction processes. In our present study, bovine and frog retinas were incubated in Krebs' solution containing [32P] H3PO4 for the labelling of all phosphoproteins. Then, photoreceptor outer segments were isolated from each retina, and further incubated under dark or light conditions. In such conditions, several phosphoproteins were dark- or light-dependently dephosphorylated. Interestingly, the light-dependent dephosphorylated 39 kDa protein as well as the 35-36 kDa protein was commonly observed in both bovine and frog retinas. The 35-36 kDa protein is considered to be the same as the 33 kDa protein that has previously been shown to be phosphorylated light-dependently, whereas 39 kDa protein is thought to be a novel protein that undergoes light-dependent dephosphorylation in retinal photoreceptor outer segments. Thus, these proteins were thought to have significant roles in the visual transduction processes.

Animals↗

PubMed1992

The presence of free D-serine, D-alanine and D-proline in human plasma.

Twelve neutral free amino acids, i.e. serine, threonine, glutamine, asparagine, alanine, proline, methionine, tyrosine, valine, leucine, isoleucine and phenylalanine, were surveyed for the presence of D-enantiomers in plasma samples from patients with renal diseases and from normal subjects. D-serine, D-alanine and D-proline were found in the patient's plasma. The highest concentrations (D/L ratio) of D-serine, D-alanine and D-proline were 0.2362, 0.2087 and 0.0986, respectively. The sum of the contents of the three D-amino acid was shown to be present in the plasma proteins.

Adult↗

PubMed1992

Pulmonary surfactant protein D specifically binds to phosphatidylinositol.

Alveolar type II cells produce and secrete a complex mixture of lipids and proteins called pulmonary surfactant of which phospholipids are the major components. Surfactant proteins (SP) A, B, and C interact with phospholipids and are believed to play important roles in alveolar spaces. However, whether surfactant protein D (SP-D) interacts with phospholipids is unknown. In the present study, we examined whether SP-D binds to phospholipids and investigated phospholipid specificities of SP-D binding and the structural requirements of phospholipids for that binding using 125I-SP-D as a probe. 125I-SP-D bound exclusively to phosphatidylinositol (PI) in various phospholipids or a fraction containing phospholipids extracted from surfactant, which were developed on thin layer chromatography. 125I-SP-D also bound to PI coated on microtiter wells in a manner dependent upon the SP-D concentration. Unlabeled SP-D competed well with 125I-SP-D for PI binding and the antibody against SP-D abolished 125I-SP-D binding to PI. PI liposome also attenuated 125I-SP-D binding to the solid phase PI. Ca2+ is absolutely required for the binding of SP-D to PI. SP-D failed to bind to lyso-PI, fatty acids derived from PI digested with phospholipase A2, or diacylglycerol obtained after phospholipase C treatment of PI. SP-D bound to neither phosphatidylinositol 4-monophosphate nor phosphatidylinositol 4,5-diphosphate. We conclude that SP-D specifically binds to PI. This is the first report that demonstrates that SP-D interacts with surfactant phospholipids.

Animals↗

PubMed1992

Calcium dependent conformational changes of surfactant protein A (SP-A) and its collagenase resistant fragment with or without dithiothreitol.

Calcium-dependent conformational changes of surfactant protein A (SP-A) and the collagenase resistant fragment (CRF) of SP-A were studied by measuring fluorescence spectra. The emission peaks of both SP-A and CRF in the absence of Ca2+ appeared at 343 nm when they were excited at 280 nm. In the presence of Ca2+, the peaks appeared at 340 nm and were accompanied by an increase in the fluorescence intensity. The magnitude of the fluorescence intensity change induced by Ca2+ was amplified by the addition of dithiothreitol (DTT) in both SP-A and CRF. The Ca2+ binding of CRF was measured by a flow dialysis method with 45CaCl2 in the Ca2+ concentration range where the Ca(2+)-induced fluorescence changes occurred. The maximum binding number of Ca2+ to CRF was about 2 mol per mol of CRF, and the value was independent of the presence of DTT.

Calcium↗

PubMed1992

Binding specificity of lung surfactant protein SP-D for glucosylceramide.

The specificities of the binding of lung surfactant protein SP-D to glycolipids were examined using 125I-labeled SP-D as a probe. When the binding study was performed on TLC plates, SP-D bound exclusively to GlcCer, whereas it failed to bind to GalCer, GM1, GM2, asialo-GM1, asialo-GM2, sulfatide, Forssman antigen, ceramide dihexoside, ceramide trihexoside, globoside, paragloboside or ceramide. Excess native SP-D competed with 125I-SP-D for the binding to GlcCer. Antibody to rat SP-D inhibited 125I-SP-D binding to GlcCer. Ca2+ was absolutely required for the binding of SP-D to GlcCer; Mg2+ failed to substitute for Ca2+. SP-D bound to ceramide monohexoside in glycolipids isolated from rat lung and bronchoalveolar lavage fluids of rats.

Animals↗

PubMed1992

Pulmonary surfactant protein A in pleural effusions.

Pulmonary surfactant protein A (SP-A) is known to be a major phospholipid-associated glycoprotein in pulmonary surfactant, which is specific to the lung. Immunohistochemically, expression of SP-A in tumor tissues is found in approximately 50% of patients with lung adenocarcinoma but not in the other histologic types of lung cancer of metastatic lung tumors. In this study, the SP-A content of pleural effusions was determined using an enzyme-linked immunosorbent assay. These results showed that approximately 40% of patients with lung adenocarcinomas (27 of 67) had high levels of SP-A (greater than 500 ng/ml) in their pleural effusions. By contrast, patients with other histologic types of lung cancers, adenocarcinomas of different primary sites, and tuberculosis had low levels of SP-A in their pleural effusions. The determination of SP-A in malignant effusions will contribute to distinguishing primary lung adenocarcinoma from adenocarcinomas of miscellaneous origin.

Adenocarcinoma↗

PubMed1992

Two gamma-subunits, gamma-I and gamma-II, complex with the same beta-subunits in bovine brain G-proteins (Gi/o).

When a mixture of bovine brain G-proteins (Gi/o) was loaded onto an octyl sepharose column in the presence of AlF4-, alpha-subunits of molecular weights 39 kDa and the 41 kDa were eluted separately, followed by the appearance of two distinct peaks containing beta gamma-subunits (beta gamma-I, beta gamma-II). Both beta gamma-I and beta gamma-II possessed identical beta-subunits but different gamma-subunits. The molecular weights of the two gamma-subunits determined by SDS-polyacrylamide gel electrophoresis both in the presence and absence of urea were 4.5 kDa (gamma-I) and 5.0 kDa (gamma-II). Tests indicated that the two isolated gamma-subunits are intact and have not undergone proteolysis. The amino acid composition of gamma-I appeared to be distinct from that of gamma-II. Therefore, this method is a simple procedure for isolating beta gamma-I and beta gamma-II.

Amino Acids↗