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2001 Fiscal Year Final Research Report Summary

Characterization and measurement of high-density lipoprotein (HDL)-binding proteins present in human peripheral monocytes

Research Project

Project/Area Number 12672241
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Laboratory medicine
Research InstitutionShinshu University

Principal Investigator

KATSUYAMA Tsutomu  Department of Laboratory Medicine, Shinshu University School of Medicine, Professor, 医学部・医学科臨床検査医学, 教授 (90020809)

Co-Investigator(Kenkyū-buntansha) OTA Hiroyoshi  Department of Biomedicai Laboratory Sciences, Shinshu University School of Medicine, Professor, 医学部・保健学科生体情報検査学, 教授 (50273107)
NAKAYAMA Jun  Department of Pathology, Shinshu University School of Medicine, Professor, 医学部・医学科病理学, 教授 (10221459)
Project Period (FY) 2000 – 2001
Keywordsmonocytes / HDL-binding proteins / high-density lipoprotein (HDL) / mass spectrometry / flow cytometry / affinity chromatography / reverse-phase high performance liquid chromatography
Research Abstract

Plasma levels of high-density lipoprotein (HDL) are inversely correlated with the risk of atherosclerotic coronary artery disease. HDL is believed to play a role in the process of reverse cholesterol transport (RCT). Although the interaction of HDL with binding sites on the surface of peripheral cells appears to be an important first step in RCT, the nature of this association is not fully understood. We previously identified a binding site for HDL3 on the surface of human peripheral blood monocytes.
In this study, we found that pre-β-HDL purified by ion-exchange chromatography from very high density lipoprotein (VHDL, 1.21 < d < 1.25 g/ml) was susceptible to degradation if isolated in the absence anti-proteases, resulting in the smaller lyso-pre-β-HDL The mass of the larger fragment from lyso-pre-β-HDL was confirmed using MALDI-TOF mass spectrometrical analysis, which showed a fragment of approximately 22625.3Da. The ability of pre-β-HDL and lyso-pre-β-HDL to compete for HDL binding to … More human monocytes was determined using a flow cytometry-based assay. Pre-β-HDL competed efficiently for binding whereas lyso-pre-β-HDL was significantly less effective. We propose that in vivo degradation of pre-β-HDL would affect the ability of these particles to bind to human monocytes.
The purification of HDL-binding proteins was achieved by chromatography using DEAE ion-exchange, wheat germ lectin and apoHDL3 affinity columns. Analysis of purified HDL-binding proteins by reverse-phase high performance liquid chromatography showed the presence of two main proteins at 100k and 120kDa. In order to measure the amount of HDL-binding protein present in CHAPS-solubilized human mononuclear cells a 96-well plate assay was developed that utilized similar principles as the purification method. The present study detailing the purification and measurement of HDL-binding proteins in mononuclear cells provides significant information likely to further our understanding of the physiological function and fate of HDL. Less

  • Research Products

    (36 results)

All Other

All Publications (36 results)

  • [Publications] Sugano M: "Analysis of hemoglobin and globin chain variants by a commonly used capillary isoelectric focusing method"Electrophoresis. 21. 3016-3019 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yamauchi K: "Effect of apolipoprotein AII on the interaction of apolipoprotein E with beta-amyloid : some apo(E-AII) complexes inhibit the internalization of beta-amyloid in cultures of neuroblastoma cells"J Neurosci Res. 62. 608-614 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Terasawa F: "Identification of a dysfibrinogen, the substitution of gamma308Asn(AAT) to Lys(AAG), using coagulation tests, immunoblot analysis, and allele-specific polymerase chain reaction."Clin Chim Acta. 295. 77-85 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kato M: "Clinical usefulness of urine-based enzyme-linked immunosorbent assay for detection of antibody to Helicobacter pylori : a collaborative study in nine, medical institutions in Japan"Helicobacter. 5. 109-119 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Okumura N: "Difference in electrophoretic mobility and plasmic digestion profile between four recombinant fibrinogens, gamma 308K, gamma 308I, gamma 308A, and wild type"Electrophoresis. 21. 2309-2315 (2000)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Hidaka H: "Characterization of an apolipoprotein E3 variant (Arg 145→His) associated with mild hypertriglyceridemia"Ann Clin Lab Sci. 31. 163-170 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Oshiro M: "Immunohistochemical localization of heparan sulfate proteoglycan in human gastrointestinal tract"Histochem Cell Biol.. 115. 373-380 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Yamauchi K: "Isoform-specific effect of apolipoprotein E on endocytosis of beta-amyloid in cultures of neuroblastoma cells"Ann Clin Lab Sci.. 32. 65-74 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Suzuki T: "Predominant apolipoprotein J exists as lipid-poor mixtures in cerebrospinal fluid"Ann Clin Lab Sci. 32. 369-376 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Okumura N: "Analysis of fibrinogen gamma-chain truncations shows the C-terminus particularly gammalle387, is essential for assembly and secretion of this multichain protein"Blood. 99. 3654-3660 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Hidaka H: "Identification and characterization of triglyceride rich lipoproteins with very light density (d.<1.000g/ml) by ultracentrifugation and agarose gel electrophoresis using triglyceride-and cholesterol-staining"Ann Clin Lab Sci. (in press). (2003)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Hidaka H: "Purification and measurement of HDL3-binding proteins present in human peripheral mononuclear cells"Ann Clin Lab Sci. (in press). (2003)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Taketomi T: "Confirmation of minor components of less polar neutral and acidic glycolipids in monkey brain tissue"J Lipid Res. 42. 873-885 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ota H: "Cell lineage specificity of newly raised monoclonal antibodies against gastric mucins in normal, metaplastic, and neoplastic human tissues their application to pathology diagnosis"Am J Clin Pathol.. 115. 69-79 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Nakabayashi T: "Evaluation of the automatic fluorescent image analyzer, Image Titer, for quantitative analysis of antinuclear antibodies"Am J Clin Pathol.. 115. 424-429 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Kumagai T: "Serum Immunoglobulin G Immune Response to Helicobacter pylori Antigens in Mongolian Gerbils"J Clin Microbiol.. 39. 1283-1288 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Fu L: "Extrahepatic Expression of Apolipoprotein A-II in Mouse Tissues : Possible Contribution to Mouse Senile Amyloidosis"J Histochem Cytochem.. 49. 739-748 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Ishii K: "A new diagnostic method for adenoma malignum and related lesions : latex agglutination test with a new monoclonal antibody, HIK1083"Clin Chim Acta. 312. 231-233 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Sugano M: "Analysis of hemoglobin and globin chain variants by a commonly used capillary isoelectric focusing method."Electrophoresis. 21. 3016-3019 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yamauchi K: "Effect of apolipoprotein All on the interaction of apolipoprotein E with beta-amyloid: some apo(E-AII) complexes inhibit the internalization of beta-amyloid in cultures of neuroblastoma cells."J Neurosci Res. 62. 608-614 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Terasawa F: "Identification of a dysfibrinogen, the substitution of gamma308Asn(AAT) to Lys(AAG), using coagulation tests, immunoblot analysis, and allele-specific polymerase chain reaction."Clin Chim Acta. 295. 77-85 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kato M: "Clinical usefulness of urine-based enzyme-linked immunosorbent assay for detection of antibody to Helicobacter pylori: a collaborative study in nine medical institutions in Japan."Helicobacter. 5(2). 109-119 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Okumura N: "Difference in electrophoretic mobility and plasmic digestion profile between four recombinant fibrinogens, gamma 308K, gamma 308I, gamma 308A, and wild type."Electrophoresis. 21. 2309-2315 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Hidaka H: "Characterization of an apolipoprotein E3 variant (Arg 145→His) associated with mild hypertriglyceridemia"Ann Clin Lab Sci. 31. 163-170 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Taketomi T: "Confirmation of minor components of less polar neutral and acidic glycolipids in monkey brain tissue."J Lipid Res. 42. 873-885 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ota H: "Cell lineage specificity of newly raised monoclonal antibodies against gastric mucins in normal, metaplastic, and neoplastic human tissues and their application to pathology diagnosis."Am J Clin Pathol. 115. 69-79 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Nakabayashi T: "Evaluation of the automatic fluorescent image analyzer, Image Titer, for quantitative analysis of antinuclear antibodies."Am J Clin Pathol. 115. 424-429 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Kumagai T: "Serum Immunoglobulin G Immune Response to Helicobacter pylori Antigens in Mongolian Gerbils."J Clin Microbiol. 39. 1283-1288 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Fu L J: "Extrahepatic Expression of Apolipoprotein A-II in Mouse Tissues: Possible Contribution to Mouse Senile Amyloidosis."Histochem Cytochem. 49. 739-748 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Ishii K: "A new diagnostic method for adenoma malignum and related lesions: latex agglutination test with a new monoclonal antibody, HIK1083."Clin Chim Acta. 312. 231-233 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Oshiro M: "Immunohistochemical localization of heparan sulfate proteoglycan in human gastrointestinal tract."Histochem Cell Biol.. 115. 373-380 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Yamauchi K: "Isoform-specific effect of apolipoprotein E on endocytosis of beta-amyloid in cultures of neuroblastoma cells."Ann Clin Lab Sci.. 32. 65-74 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Suzuki T: "Predominant apolipoprotein J exists as lipid-poor mixtures in cerebrospinal fluid."Ann Clin Lab Sci.. 32. 369-376 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Okumura N: "Analysis of fibrinogen gamma-chain truncations shows the C-terminus, particularly gammaIle387, is essential for assembly and secretion of this multichain protein."Blood. 99. 3654-3660 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Hidaka H: "Identification and characterization of triglyceride rich lipoproteins with very light density (d. < 1.000 g/ml) by ultracentrifugation and agarose gel electrophoresis using triglyceride- and cholesterol-staining."Ann Clin Lab Sci.. (in press). (2003)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Hidaka H: "Purification and measurement of HDL3-binding proteins present in human peripheral mononuclear cells."Ann Clin Lab Sci.. (in press). (2003)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 2003-09-17  

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