Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P30536 (PBS)
 9,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Potent and novel fibrinolytic enzymes (lumbrokinase [LK]) were extracted from the earthworm, Lumbricus rubellus. These enzymes were very stable and showed greater antithrombotic activity than other currently used fibrinolytic proteins. An LK fraction showing the most potent fibrinolytic activity was immobilized onto a polyurethane (PU) surface to investigate its enzymatic activity and antithrombotic activity. A methanol-extracted PU surface was coated with 3% (wt/vol) maleic anhydride methylvinyl ether copolymer (MAMEC)/tetrahydrofuran (THF) solution, and the surface was incubated in an LK solution/phosphate-buffered saline (PBS, pH 7.4). The surface properties were characterized by attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), electron spectroscopy for chemical analysis (ESCA), and dynamic contact angle. The stability of immobilized LK was determined by caseinolytic activity assay and the specificity of immobilized LK on fibrinogen/fibrin was observed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The antithrombotic activity of immobilized LK was evaluated using an ex vivo rabbit A-A shunt experiment. LK immobilization was confirmed by ATR-FTIR and ESCA. Immobilized LK demonstrated stable proteolytic activity during various incubation periods. Immobilized LK proteolyzed fibrinogen and fibrin almost specifically, while it hardly hydrolyzed other plasma proteins including plasminogen and albumin. In the ex vivo A-A shunt experiment, the LK-immobilized surface significantly prolonged occlusion time over control surfaces. This is primarily due to the high thrombolytic activity of immobilized LK. In this work, a highly efficient surface modification method on the PU surface was developed, and this LK immobilization technique will be very useful in improving the blood compatibility of blood-contacting devices.
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PMID:Surface characteristics and properties of lumbrokinase-immobilized polyurethane. 761 90

Angiostatin is a potent angiogenesis inhibitor that is composed of the first four kringles of plasminogen fragment. Angiostatin with one less kringle molecule (kringle 1 to 3) was recently demonstrated to be an effective angiogenic inhibitor. To determine whether recombinant plasminogen kringle 1-3 (rPK1-3) can inhibit the corneal neovascularization induced by potent angiogenic factors; angiogenin, bFGF, or VEGF, hydron polymer discs each containing 2.0 microg of angiogenin, 500 ng of bFGF, or 500 ng of VEGF respectively were implanted into the corneal stroma of 138 rabbit eyes, and then discs each containing 10 microg, 12.5 microg, 20 microg or 30 microg of rPK1-3 were implanted randomly. Discs containing phosphate buffered saline were also implanted as a control. The angiogenesis score on number and length of newly formed vessels on the each of the rabbit's cornea were recorded daily by two observers (blinded). The treated corneas were also examined histologically. Recombinant PK1-3 treated corneas showed less neovascularization induced by all angiogenic factors (p < 0.05). and the extent of inhibition of neovascularization was proportional to the concentration of rPK1-3 (p < 0.05). Histologic examination showed leukocyte infiltration into the corneal stroma on the PBS treated eyes whereas rPK1-3 treated eyes showed only traces of leukocytes. These results of the effective rPK1-3 inhibition of corneal neovascularization induced by angiogenin, bFGF, or VEGF suggest that this angiostatin related fragment, rPK1-3, may be useful in the treatment of various neovascular diseases.
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PMID:The inhibitory effects of recombinant plasminogen kringle 1-3 on the neovascularization of rabbit cornea induced by angiogenin, bFGF, and VEGF. 1063 Mar 75

To clarify the release properties of anti-cancer drugs from fibrin glue, a study was performed using several anti-cancer drugs with remarkably different physical properties. Concentrated fibrinogen, fibronectin, and coagulation factor XIII were prepared from healthy human plasma according to the cryoprecipitate method. Fibrin glue containing anti-cancer drugs was prepared as follows; the cryoprecipitate was mixed with each anti-cancer drug and aprotinin, then thrombin was added. These glues were incubated in PBS containing plasminogen and urokinase at 37 degrees C for seven days, and the medium was then sampled several times after centrifugation. The drug concentration in each sample was measured using HPLC. Fibrin glue without aprotinin was quickly hemolyzed and disappeared after 2--4 h. That with aprotinin was only slightly hemolyzed and more than half remained after 7 days. Mitomycin C and fluorouracil were quickly released from the glue regardless of the presence or absence of aprotinin. However, enocitabine was gradually released from glue with aprotinin although quickly released from that without. The rate of release of each drug from the glue with aprotinin correlated well with its hydrophobicity. Thus, to establish a sustained release system using fibrin glue, one should use the more lipophilic anti-cancer drugs and a fibrinolytic enzyme inhibitor.
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PMID:Novel drug delivery system using autologous fibrin glue--release properties of anti-cancer drugs. 1072

Angiostatin, an internal fragment of plasminogen, has been shown to inhibit the process of angiogenesis or neovascularization. In this study, we have expressed the cDNA for murine angiostatin under the control of the human cytomegalovirus promoter from a human type-5 adenovirus and shown that this vector produces a protein which retains biological activity. Angiostatin expression was determined by Northern blot analysis and Western immunoblotting. Ad-angiostatin, but not a control vector Ad-dl70, significantly reduced the viability of infected human umbilical cord vein endothelial cells (HUVEC) in vitro. In an in vivo model of basic fibroblast growth factor-induced angiogenesis, Ad-angiostatin (1 x 10(9) pfu) could inhibit endothelial cell migration and the formation of capillaries within a Matrigel plug which had been implanted for one week subcutaneously into C57BL/6 mice. Endothelial cells in these plugs had an altered, rounded, phenotype with dark picnotic nuclei indicative of apoptosis, which was confirmed using transmission electron microscopy. In contrast, endothelial cells from bFGF alone or in combination with the control vector-treated plugs retained the long spindle shape characteristic of endothelial cells. Intranasal delivery of Ad-angiostatin into the lungs of FVB/n mice demonstrated comparable cellular infiltration in the recovered bronchoalveolar lavage fluid with no signs of abnormal pathology as compared to PBS or control vector-treated animals. In a pulmonary metastatic breast cancer model, the delivery of Ad-angiostatin (1 x 10(9) pfu) to the lung significantly delayed tumor growth as measured by the number of visible surface tumor nodules. This study has demonstrated that the specific targeting of tumors to inhibit angiogenesis using an adenovirus expressing angiostatin, may deliver localized concentrations of protein having a greater impact on inhibition of tumor growth.
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PMID:Adenoviral vector expressing murine angiostatin inhibits a model of breast cancer metastatic growth in the lungs of mice. 1154 7

Reliable data on plasmin activities in blood of patients during fibrinolytic treatment are lacking. This is due to continuing plasminogen activation by plasminogen activators after blood withdrawal. The purpose of this study was to establish a new method for stabilization of blood and to detect plasmin activity in stabilized plasma. For optimization of plasma stabilization by arginine, 50 microL pooled normal citrated plasma was incubated with 50 microL of 0 to 1500 mM arginine, pH 8.7, and 25 microL 100 IU/mL u-PA, 1250 IU/mL t-PA, 10000 U/mL reteplase, 400 U/mL plasminogen-streptokinase-activator complex, 10 microg/mL tenecteplase in 6% BSA-PBS or 25 microL 25 microg/mL plasmin in 20% glycerol. Twenty-five microliters 3 mM HD-Val-Leu-Lys-pNA were added immediately (1 step) or after 90 minutes (room temperature [RT]). The same experiment was performed with pooled normal citrated plasma supplemented with 3.2 mg/mL EDTA, preoxidized with 0 mM or 20 mM chloramine-T for 10 minutes (37 degrees C). For optimization of plasmin activity, the oxidation time of the arginine-stabilized plasma sample containing 0.5 U/mL active plasmin and the chloramine-T amount was varied. Citrated plasma is stabilized against the in vitro action of all six plasminogen activators tested if the final arginine concentration is greater than 500 mM. Neither the addition of EDTA nor the addition of chloramine-T changes this plasma-stabilizing power of arginine. The optimized functional plasmin assay consists of incubation of 10 microL arginine-stabilized plasma with 10 microL 1.5 M arginine, pH 8.7, and 10 microL 100 mM CT in PBS. After 30 minutes (37 degrees C), 75 microL 1.2 M KCl, 1.6 M Arg, 0.75 mM Val-Leu-Lys-pNA (Stop-CS Reagent), and 175 microL 6% BSA-PBS are added and the absorbance increase (DeltaA) at 405 nm is determined. With the present arginine stabilization procedure of plasma and the determination of plasmin activity in arginine-stabilized plasma as described, it is feasible to determine the activity of plasmin in blood of patients receiving fibrinolytic treatment without artefactual in vitro changes in the samples.
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PMID:Functional determination of plasmin in arginine-stabilized plasma. 1601 16

Reliable data on plasminogen activator (PA) activities in blood of patients receiving fibrinolytic treatment are lacking. This is due to the continuing in vitro action of PA after blood withdrawal. We have elaborated a new simple stabilization technique for plasma involving the addition of arginine in final concentrations greater than 500 mM. In this study, new assays for PA in stabilized plasma are developed. The assay was performed with substrate plasma, that is, pooled normal plasma, preoxidized with chloramine-T; oxidant amount and oxidation time were optimized. The chloramine consumption by plasma was assayed with a KJ-assay (absorbance increase at 405 nm by addition of 200 microL 4 M KJ to 25 microL oxidized plasma). The substrate plasma concentration in the PA assay and the PA acting time was optimized. The inhibition of PA by the cations Na(+), K(+), Mg(2+), and Ca(2+) was evaluated. The optimized PA assay consists of incubation of 10 microL arginine-stabilized plasma with 10 microL 1.5 M arginine, pH 8.7 and 10 microL 100 mM CT in PBS. After 30 minutes (37 degrees C), 175 microL 15 mM CT oxidized EDTA plasma are added. After 40 minutes (37 degrees C), 75 microL Stop-CS Reagent is added and DeltaA at 405 nm was determined, giving PA + plasmin activity in plasma. A control value (basal plasmin activity) consists of the addition of Stop-CS Reagent before 175 microL oxidized EDTA plasma. To obtain plasmatic PA activity, the control value has to be subtracted from the PA main value. The assay is matrix-independent and linear up to 1250 IU/mL t-PA, 790 U/mL reteplase, or 199 IU/mL u-PA (37 nM). With arginine stabilization of plasma and the described determination of plasminogen activator activity in arginine-stabilized plasma, it is feasible to determine the activity of plasminogen activators in blood of patients receiving fibrinolytic treatment without artefactual in vitro changes of the samples.
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PMID:Functional determination of plasminogen activator in arginine-stabilized plasma. 1601 17

Kringle 5 (K5), a proteolytic fragment of plasminogen, has been proved to be an angiogenic inhibitor. Previously, we have evaluated the effect of K5 on the vascular leakage and neovascularization in a rat model of oxygen-induced retinopathy. In this study, we expressed K5 and a deletion mutant of K5 (K5 mutant) in a prokaryocyte expression system and purified them by affinity chromatography. K5 mutant was generated by deleting 11 amino acids from K5 while retaining the three disulfide bonds. The anti-angiogenic activity of intact K5 and K5 mutant were compared in endothelial cells and retinal neovascularization rat model. K5 mutant inhibited the proliferation of primary human retinal capillary endothelial cells (HRCEC) in a concentration-dependent manner, with an apparent EC50 of approximate 35 nmol/L, which is twofold more potent than intact K5. In the even higher concentration range, K5 mutant did not inhibit pericytes from the same origin of HRCEC, which suggested an endothelial cell-specific inhibition. K5 mutant had no effect on normal liver cells and Bel7402 hepatoma cells even at high concentration range either. Intravitreal injection of the K5 and mutant in the oxygen-induced retinopathy rat model both resulted in significantly fewer neovascular tufts and nonperfusion area than controls with PBS injection, as shown by fluorescein angiography. Furthermore, K5 mutant exhibited more strong inhibition effect on neovascularization than intact K5 by quantification of vascular cells. These results suggest that this K5 deletion mutant is a more potent angiogenic inhibitor than intact K5 and may have therapeutic potential in the treatment of those disorders with neovascularization, such as solid tumor, diabetic retinopathy, age-related macular degeneration, rheumatoid arthritis, and hyperplasia of prostate.
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PMID:Enhanced anti-angiogenic effect of a deletion mutant of plasminogen kringle 5 on neovascularization. 1616 44

Diabetic nephropathy, a major complication of diabetes mellitus that leads to mortality, has been shown to involve a dysregulation of the coagulation system. Annexin-2, a co-receptor for plasminogen and tissue plasminogen activator on endothelial cells, is one of the molecules required to maintain the antithrombogenic properties of endothelial cells. Previously, we showed that recombinant annexin-2 protein (rAN II) modulated impaired fibrinolytic activity in the carotid arteries of rats. In the present study, to investigate its protective effects against diabetic nephropathy, rAN II was administered to KK-Ay mice, a murine model of type 2 diabetes, for eight weeks, and albuminuria, kidney size, and histological glomerular lesions were investigated. The mean weight of kidneys from KK-Ay mice treated with rAN II was significantly less than that of those treated with PBS (control) (p < 0.02). Furthermore, the level of albuminuria observed in rAN II-treated KK-Ay mice was significantly less than that of the control group (rAN II, 0.90+/-0.12 microg/day; PBS, 1.55+/-0.31 microg/day; p < 0.01); also, the area of diffuse glomerular lesions was significantly smaller (rAN II, 41.51+/-4.54%; PBS, 81.81+/-8.10%; p < 0.01). Bleeding time, prothrombin time (PT), and active partial thromboplastin time (APTT) did not significantly differ between the two groups. Our results suggest that rAN II may inhibit the progression of diabetic nephropathy in KK-Ay mice without influencing the coagulation system, indicating that annexin-2 may be considered as a possible new therapeutic tool for patients with diabetic nephropathy.
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PMID:Recombinant annexin-2 inhibits the progress of diabetic nephropathy in a diabetic mouse model via recovery of hypercoagulability. 1720 Jul 79