ABSTRACT
BACKGROUND: Fibrinogen gamma', a fibrinogen gamma-chain variant generated via alternative mRNA processing, has been associated with susceptibility to thrombotic disease. OBJECTIVE: The present case-control study searched for potential determinants of the plasma fibrinogen gamma' concentration and examined the relationship between this variant and risk of myocardial infarction (MI). PATIENTS AND METHODS: The Stockholm Coronary Artery Risk Factor study, comprising 387 postinfarction patients and 387 healthy individuals, was employed. The fibrinogen gamma (FGG) 9340T > C [rs1049636], fibrinogen alpha (FGA) 2224G > A [rs2070011] and fibrinogen beta (FGB) 1038G > A [rs1800791] polymorphisms were determined. The plasma fibrinogen gamma' concentration was measured by enzyme-linked immunosorbent assay. The multifactor dimensionality reduction method was used for interaction analyses on risk of MI. RESULTS: The FGG 9340T > C and FGA 2224G > A polymorphisms, total plasma concentrations of fibrinogen, insulin and high-density lipoprotein, and gender appeared to be independent determinants of plasma fibrinogen gamma' concentration in patients, and the corresponding determinants in controls included FGG 9340T > C and FGA 2224G > A polymorphisms and plasma fibrinogen concentration. An elevated plasma fibrinogen gamma' concentration proved to be an independent predictor of MI [adjusted odds ratio (OR) (95% CI): 1.24 (1.01, 1.52)]. The plasma fibrinogen gamma' concentration was involved in a high-order interaction with total plasma fibrinogen and the FGG 9340T > C and FGA 2224G > A polymorphisms, associated with a further increased risk of MI [OR (95% CI): 3.22 (2.35, 4.39)]. CONCLUSIONS: Plasma fibrinogen gamma' concentration influences the risk of MI, and this relationship seems to be strengthened by the presence of an elevated total plasma fibrinogen concentration and the FGG 9340T and FGA 2224G alleles.
Subject(s)
Fibrinogen/genetics , Myocardial Infarction/blood , Peptide Fragments/blood , Peptide Fragments/genetics , Female , Fibrin/chemistry , Fibrinogen/chemistry , Genetic Predisposition to Disease , Humans , Male , Models, Genetic , Polymorphism, Genetic , Protein Isoforms , RNA, Messenger/metabolism , Risk , Risk FactorsABSTRACT
A high-affinity thrombin-binding site in an alternately processed fibrinogen variant, the gammaA/gamma' isoform, is characterized in this report. The binding site has been shown to be situated between gamma' 414 and 427, and Tyr418 and 422 in this part of the gamma' chain are known to be sulfated. A synthetic peptide corresponding to the gamma' chain carboxyl terminus is shown to bind thrombin with a Kd = 0.63 +/- 0.16 micro mol L-1. Maximum binding of this peptide requires negative charges on Tyr418 and 422. Competitive binding studies with hirudin peptides, heparin and DNA aptamers specific for thrombin exosites I or II indicate thrombin binds to the gamma' peptide via exosite II. Thus, thrombin binding to the gamma' chain leaves exosite I and the active site accessible to substrates. This may explain why fibrin-bound thrombin can retain enzymatic activity, and why fibrin-bound thrombin is heparin-resistant.
Subject(s)
Fibrinogen/genetics , Fibrinogen/metabolism , Thrombin/metabolism , Amino Acid Sequence , Animals , Binding Sites , Binding, Competitive , Cell Line , DNA/chemistry , DNA/metabolism , Fibrinogen/chemistry , Heparin/chemistry , Heparin/metabolism , Hirudins/chemistry , Hirudins/metabolism , Humans , Kinetics , Mice , Molecular Sequence Data , Protein Binding , Protein Isoforms/genetics , Protein Isoforms/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Substrate Specificity , Thrombin/chemistry , Transfection , Tyrosine/chemistry , Tyrosine/metabolismABSTRACT
FF6 tumor cells are derived from a spontaneous rat squamous cell carcinoma (SCC) which originally arose in the facial skin of a DA rat. In this study, FF6 tumor cells were implanted into rat oral mucosa to establish an ex vivo metastatic model. We analyzed the expression of intercellular cell adhesion molecule-1 (ICAM-1) in the implanted primary and metastatic FF6 tumors by immuno-staining with a monoclonal antibody (mAb) against ICAM-1. The implanted primary FF6 cells showed strong expression of ICAM-1, whereas the tumor cells of metastatic lesions showed weak or negative expression of ICAM-1. By immunostaining with mAb OX6, a number of MHC class II-positive macrophages were detected in tumor mesenchyme and surrounding the metastatic foci. These results suggested that the local immune reaction in the lymph node influenced the expression of ICAM-1 on tumor cells, and that MHC class II-positive macrophages may play a role in transplanted tumor growth and metastases.
Subject(s)
Carcinoma, Squamous Cell/pathology , Intercellular Adhesion Molecule-1/analysis , Animals , Antibodies, Monoclonal , CD4 Antigens/analysis , CD4-Positive T-Lymphocytes/pathology , CD8 Antigens/analysis , CD8-Positive T-Lymphocytes/pathology , Carcinoma, Squamous Cell/secondary , Coloring Agents , Eosine Yellowish-(YS) , Facial Neoplasms/pathology , Fluorescent Dyes , Gene Expression Regulation, Neoplastic , Hematoxylin , Histocompatibility Antigens Class II/analysis , Immunoenzyme Techniques , Intercellular Adhesion Molecule-1/genetics , Lip Neoplasms/pathology , Lymph Nodes/immunology , Lymph Nodes/pathology , Lymphatic Metastasis/pathology , Macrophages/immunology , Macrophages/pathology , Mesoderm/pathology , Mouth Neoplasms/pathology , Mouth Neoplasms/secondary , Neoplasm Transplantation , Random Allocation , Rats , Rats, Inbred Strains , Tongue Neoplasms/pathology , Tumor Cells, CulturedABSTRACT
This article describes the recognition of a special membrane antigen of the rat squamous cell carcinoma (SCC) by a monoclonal antibody (mAb), UB23, and the characterization of the UB23 antigen expression in the implanted primary and metastatic SCC in rat models. The mAb UB23 was raised against the FF6 tumor, a well-differentiated rat SCC, and it recognized the 120- to 130-kD cell surface antigen in FF6 tumor cells. The UB23 antigen was found in frequently observed small 'basal' cells but not in keratinocytes, and an increased expression was seen in the cells at the interface with peritumoral stroma in both the implanted primary FF6 tumors and metastases. These results indicated that the UB23 antigen is closely related with the cell differentiation and invasion of FF6 cells, and could be useful for analyzing the mechanism of differentiation, invasion and metastasis of SCC in animal models.
Subject(s)
Antibodies, Monoclonal/immunology , Antigens, Neoplasm/immunology , Carcinoma, Squamous Cell/immunology , Membrane Proteins/immunology , Neoplasm Proteins/immunology , Skin Neoplasms/immunology , Animals , Antibodies, Monoclonal/biosynthesis , Antibodies, Neoplasm/immunology , Antigen-Antibody Reactions/immunology , Antigens, Neoplasm/isolation & purification , Blotting, Western , Carcinoma, Squamous Cell/pathology , Carcinoma, Squamous Cell/secondary , Electrophoresis, Polyacrylamide Gel , Immunoenzyme Techniques , Male , Membrane Proteins/isolation & purification , Mice , Mice, Inbred BALB C , Neoplasm Proteins/isolation & purification , Neoplasm Transplantation , Rats , Skin Neoplasms/pathologyABSTRACT
BACKGROUND: In previous studies, we developed several monoclonal antibodies (mAbs) against fetal and adult rat liver to analyze the hemopoietic microenvironment of the fetal liver during the gestational period. In this study, we have developed two new mAbs against fetal rat liver cells and have examined the characteristics at various gestational ages of fetal liver and of adult liver. METHODS: The characteristics of these monoclonal antibodies were demonstrated by examining several tissues using immunohistochemical staining and flow cytofluorometry. RESULTS: Monoclonal antibodies HAM10 and HAM11 were developed against fetal rat liver cells. These reacted with the cytoplasm of fetal and adult hepatocytes. HAM10 antigen expression was strong at approximately day 18 of gestation in the active period of hemopoiesis in fetal rat liver but was much lower in adult liver. HAM10 antigen expression also increased in liver after partial hepatectomy and was reduced abruptly to a normal level thereafter. HAM11 antigen expression in fetal liver was weaker than that of HAM10 antigen expression. The degree of HAM11 antigen expression increased as gestation proceeded, reaching a maximum in adult liver. CONCLUSIONS: Both HAM10 and HAM11 antigens may play a role in the morphogenesis of hepatocytes and in the hemopoietic microenvironment for hemopoietic cells. Moreover, HAM10 antigen is may play a role in hepatocyte proliferation in the fetal liver, whereas HAM11 antigen may contribute to the maturation of fetal- to the adult-type hepatocytes.
