Tohgo Nonaka

An anteroposterior axis of the tibia for total knee arthroplasty.

The objective of the current study was to identify a new extraarticular anatomic landmark indicating the anteroposterior orientation of the tibia using computed tomography. In 39 volunteers (20 males, 19 females), computed tomography scans for healthy right knees in extension were done perpendicular to the tibial shaft axis. The anteroposterior axis of the tibia was defined as a line perpendicular to the transepicondylar axis and passing through the middle of the posterior cruciate ligament. At the level of the tibial plateau and the patellar tendon attachment, the mean medial percentage width of intersecting point of the patellar tendon and the anteroposterior axis was 10.8% ± 9.8% (range, −9.3%–+30.0%) and −0.2% ± 10.4% (range, −23.6%–+23.0%), respectively. The mean angle between the anteroposterior axis and a line connecting the middle of the posterior cruciate ligament to the medial border of the patellar tendon attachment was 0.0° ± 2.8° (range, −6.3°–+5.2°). The medial border of this attachment therefore can serve as a reliable anterior anatomic landmark to determine the anteroposterior axis of the tibia, and the line connecting the middle of the posterior cruciate ligament and the medial border of the attachment may be useful as a reference axis indicating the anteroposterior orientation of the tibia.

SIGNIFICANCE OF SERINE PROTEINASE AND MATRIX METALLOPROTEINASE SYSTEMS IN THE DESTRUCTION OF HUMAN ARTICULAR CARTILAGE

1. During the destruction of articular cartilage, fibrinolytic enzymes and matrix metalloproteinases (MMP) may contribute to the related pathology. The activities, antigens and messenger RNA (mRNA) levels of urokinase‐type plasminogen activator (uPA) and plasminogen activator inhibitor‐1 (PAI‐1) in articular cartilage were measured in patients with no history of joint diseases (control), those with osteoarthritis (OA) classified into osteophyte‐formed site (OS) and weight‐bearing site (WS), and in patients with rheumatoid arthritis (RA).

Analysis of binding protein for tissue-type plasminogen activator in human endothelial cells

The specific binding sites for tissue-type plasminogen activator (t-PA) were investigated in human umbilical vein endothelial cells. After adding 125I-t-PA (M.W. 70 kDa) to endothelial cells in suspension culture, the ligand was recovered from the cell extract after disuccinimidyl suberate treatment as a high molecular complex with M.W. of 90 kDa on SDS-PAGE. The complex reacted to only anti-t-PA IgG but not to anti-PAI-1 IgG immunoblot analysis, indicating a t-PA specific binding protein. 125I-t-PA ligand blotting of the cell extract revealed that the binding protein had M.W. 20 kDa. The binding of 125I-t-PA to endothelial cells was reduced in the presence of an excess amount of t-PA, plasminogen and 6-aminohexanoic acid, indicating that the binding sites were also recognized by plasminogen, and that t-PA and plasminogen were bound via lysine binding sites in the molecule. These findings suggest that human endothelial cells have specific t-PA binding molecules which may be expressed on the cell surface as t-PA receptors.

Effects of sodium hyaluronic acid on fibrinolytic factors in the synovial fluid (in vivo)

Abstract Twelve patients with osteoarthritis of the knee were administrated hyaluronic acid (HA), and measured for fibrinolytic factors on synovial fluids. It was observed that urokinase-type plasminogen activator (u-PA) activity is enhanced 3 h after administration of HA, and that the activity gradually decreased from the 2nd to the 4th week on the patients whose clinical parameters showed improvement. Antigen of u-PA and PA inhibitor-1 (PAI-1) were increased 3 h after the administration of HA in the cases that improved. However, u-PA antigen gradually decreased from the 2nd to the 4th week similarly to u-PA activity. On the other hand PAI-1 antigen was increased from the 2nd to the 4th week in the improved cases. These results demonstrate that the decrease of fibrinolytic activity in synovial fluid is associated with the improvement of osteoarthritis by treatment with HA.

Effect of cyclic AMP on urokinase-type plasminogen activator receptor and fibrinolytic factors in a human osteoblast-like cell line

We investigated the effect of cyclic AMP (cAMP) on the pericellular fibrinolytic system in NY cells. Dibutyryl cAMP (dbcAMP) or forskolin increased the level of urokinase-type plasminogen activator (u-PA) mRNA and enhanced the secretion of u-PA antigen into the conditioned medium. These agents also increased u-PA antigen on the cell surface. PA inhibitor-1 (PAI-1) antigen was inhibited by dbcAMP or forskolin. Butyrate had no effect on the production and secretion of u-PA and PAI-1. A binding assay of 125I-DFP-u-PA to NY cells revealed a single class of binding sites with a Kd of 3.85 nM and Bmax of 0.89.10(5) binding sites/cell. The Bmax was increased by dbcAMP (1 mM or 10 mM), forskolin (2 microM or 20 microM) of 1.0-, 1.4-, 1.2- and 1.8-fold, respectively. However, the Kd value was not changed. Furthermore, the level of mRNA for the u-PA receptor (u-PAR) was increased by these agents 1.2-, 1.7-, 1.8- and 2.5-fold, respectively. However, butyrate did not alter either the Bmax or the u-PAR mRNA level. These results indicated that the pericellular fibrinolytic activity induced by u-PA/u-PAR is modulated by cAMP in osteoblast-like cells.

Effects of α2-plasmin inhibitor on plasminogen activation by staphylokinase/plasminogen complex

Abstract Using a stable cross-linked SAK/plg complex, the effects of α2-plasmin inhibitor on plasminogen activation by SAK were investigated. α2-Plasmin inhibitor inhibited dose-dependently plasminogen activation by the SAK/plg complex. When FCB-2 or EACA was added to the reaction mixture of SAK/plg complex and α2-plasmin inhibitor, the inhibitory activity of α2-plasmin inhibitor was abolished and the enzymatic activity of the complexes was restored. α2-Plasmin inhibitor inhibited the activity of the SK/plg complex, but neither FCB-2 nor EACA restored the plasminogen activator activity in the mixture of SK/plg complex and α2-plasmin inhibitor. Using 125I-labeled SAK/plg complex or SK/plg complex, the reaction of the complex with α2-plasmin inhibitor was analyzed. The SAK/plg complex produced a new complex with α2-plasmin inhibitor. The formation of a new high molecular weight complex with α2-plasmin inhibitor was abolished by both EACA or FCB2. With regard to the SK/plg complex, neither EACA nor FCB-2 suppressed the complex formation with α2-plasmin inhibitor. These findings indicate that the SAK/plg complex binds to fibrin, and that this complex expresses plasminogen activator activity without being affected by α2-plasmin inhibitor.

Effect of bone resorbing factors on u-PA and its specific receptor in osteosarcoma cell line

This study investigated the effect of bone resorbing factors on the pericellular fibrinolytic system of osteosarcoma NY cells. Parathyroid hormone (PTH), prostaglandin E2, (PGE2) or tumor necrosis factor alpha (TNF-alpha) enhanced the secretion of urokinase-type plasminogen activator (u-PA) antigen and suppressed the secretion of plasminogen activator inhibitor-1 (PAI-1) antigen to the conditioned medium. The former two factors also increased u-PA antigen in the cell surface. Transforming growth factor beta (TGF-beta) enhanced u-PA antigen, but its activity was suppressed due to the increased secretion of PAI-1. The binding assay of [125I]DFP-u-PA to NY cells revealed the presence of a single class of binding sites with a Kd of 5.51 nM and Bmax of 0.92 x 10(5) binding sites/cell. PTH or PGE2 increased Bmax 1.4-fold and enhanced the u-PA receptor (u-PAR) mRNA level 1.4-fold or 2.4-fold, respectively. However, TGF-beta did not alter either the Kd or u-PAR mRNA level. Thus, pericellular fibrinolytic activity by u-PA/u-PAR and PAI-1 is modulated by bone resorbing factors.

Activation of histamine H1 receptor results in enhanced proteoglycan synthesis by human articular chondrocyte: involvement of protein kinase C and intracellular Ca2+

In earlier work, we obtained evidence for the presence of histamine H1 and H2 receptors on chondrocytes. Activation of the H1 receptor enhanced keratan sulfate synthesis, and protein kinase C (PKC) inhibitors antagonized histamine-stimulated keratan sulfate (KS) synthesis. These data do indicate the involvement of PKC in activation of H1 receptor, but precise mechanisms remained to be clarified. Human articular chondrocytes were treated with different concentrations of histamine and its antagonist. Intracellular Ca(2+) and proteoglycan synthesis was measured, using the fluorescent indicator dye Fura-2 AM and [35S]-sulfate incorporation, respectively. Activation of the H1 receptor led to stimulation of proteoglycan synthesis and evoked increases in levels of intracellular Ca(2+). Activity of PKC was also enhanced with activation of the H1 receptor. Intracellular Ca(2+) and activation of PKC are involved in the signal transduction pathway of H1 receptor-mediated stimulation of proteoglycan synthesis.

Comparison of intravenous and subcutaneous erythropoietin therapy for preoperative acquisition of blood for autologous transfusion in patients undergoing total arthroplasty

Before undergoing total arthroplasty, 120 patients agreed to provide their own blood for autologous transfusion. Recombinant human erythropoietin (EPO) was given intravenously to 30 patients (group I), and subcutaneously to 40 (group S); 50 patients (group N) were not given EPO. Fifteen of the 120 patients (group I,n=3; group S,n=3; group N,n=9) required homologous blood transfusion because their hemoglobin levels fell below 7.0g/dl or their vital signs changed after surgery. The hemoglobin recovery rates (HRRs) of groups I, S, and N increased by 22.8%, 47.2%, and 11.8% respectively, compared to estimated hemoglobin rates. The HRRs per 10000 IU EPO increased by 4.7% and 3.6%, respectively, in groups S and I. Groups I and S lost less blood, in total, during and after surgery than group N. Five of the 70 patients in groups I and S combined did not respond to EPO, but no patients treated with EPO experienced any side effects prior to the surgery. During the trial, one woman, who underwent total hip arthroplasty, developed a pulmonary embolism on the 10th postoperative day. Platelet counts and plasminogen activator inhibitor-1 antigen levels in all groups increased after blood donation, with values in the EPO-treated groups being higher than those in group N. Treatment with EPO makes more feasible the donation of their own blood for autologous transfusion in patients about to undergo total arthroplasty.

Comparison of the inhibitory effects of two types (90 kDa and 190 kDa) of hyaluronic acid on the expression of fibrinolytic factors in human synovial fibroblasts.

Abstract  Hyaluronic acid (HA) has been shown to be clinically effective, and is currently used for the treatment of arthropathy. We previously reported that HA of molecular weight 90 kDa (90-HA) inhibits the fibrinolytic factors in human synovial fibroblasts. In the present study, we investigated the effect of high molecular weight (190 kDa) HA (190-HA) compared with 90-HA on the pericellular fibrinolytic system of human synovial fibroblasts in osteoarthritis (OA) and rheumatoid arthritis (RA). Human synovial fibroblasts were obtained from synovial tissues of OA and RA, and were cultured in the presence and absence of 90-HA and 190-HA. Antigens of urokinase-type plasminogen activator (u-PA) and PA inhibitor-1 (PAI-1) were measured by ELISA, and the u-PA activity of the cell surface fraction was evaluated by electrophoretic enzymography. The binding assay of u-PA and the immunocytochemical analysis of u-PA were performed to detect u-PA receptor (u-PAR). HA inhibited the secretion of both u-PA and PAI-1 antigens from the synovial fibroblasts of OA to their conditioned medium, and the degree of inhibition was more effective in OA than in RA. The u-RA binding assay to these cells showed that both 90-HA and 190-HA slightly decreased the maximal number of binding sites (Bmax) in OA. However, in RA, stimulation with 90-HA and 190-HA decreased Bmax by a half and a quarter, respectively. Immunohistochemical analysis showed that u-PAR was constitutively expressed in both synovial fibroblasts, but if these cells were treated with HA, the decrease in the staining of u-PAR was more pronounced in RA than in OA. Furthermore, the degree was more effective with 190-HA than with 90-HA. HA inhibited the pericellular fibrinolytic activity mediated by the u-PA/u-PAR system in synovial fibroblasts of OA and RA, and 190-HA inhibited it more effectively than 90-HA.