Biomedical subjects

R Mischke

Publications and source records attributed to R Mischke.

At least 19 recordsLinked to original sources

PubMed2000

Influence of factor VIII:C and factor IX activity in plasmas of haemophilic dogs on the activated partial thromboplastin time measured with two commercial reagents.

The present study is based on 145 plasma samples with a reduced activity of factor VIII:C (range: 0.009-0.62 IU mL-1) and 28 samples with a reduced factor IX activity (range: 0.035-0.55 IU mL-1). The samples were collected from dogs with haemophilia A (n=22) or haemophilia B (n=3), some of these during substitution therapy. For all samples the activated partial thromboplastin time (APTT) was measured with two commercial reagents containing kaolin as a contact activator. In each case, the deficiency of factor VIII:C or IX was reflected in abnormal results of the APTT. This was true for both reagents. A significant correlation (P < 0.001) was found between factor VIII:C activity and APTT (reagent 1, Pathromtin(R); Spearman's rank correlation coefficient, rS=-0.731, reagent 2, PTT-Reagenz; rS=-0.875) as well as between factor IX activity and APTT (reagent 1, rS=-0.819; reagent 2, rS=-0.955]. In each case, the relationship between coagulation factor activity and APTT could be proven most precisely by geometric regression. The results of this study illustrate the applicability of commercial APTT test kits as a sensitive screening test of factor VIII:C and IX deficiencies in canine plasma.

Animals↗

PubMed2000

[Effect of a high single subcutaneous dose of unfractionated heparin on platelet function in dogs].

The influence of heparin on different tests of platelet function was investigated in 5 healthy dogs receiving subcutaneously 1000 I.E./kg BW of a commercial unfractionated Sodium heparin preparation. Blood samples were collected before and 4 hours after heparin injection. Besides plasma activity of heparin platelet count, capillary bleeding time with two different methods, platelet aggregation according to Breddin and to Born with the inductors ADP, collagen, and thrombin as well as in vitro bleeding time were measured. The activity of heparin four hours after the subcutaneous heparin application was 1.20 +/- 0.11 I.E./ml. Compared to the starting values no significant influence of this heparin level could be demonstrated on platelet count, platelet aggregation according to Born with the inductors ADP and collagen, platelet aggregation according to the Breddin method as well as in vitro bleeding time (p > 0.05). Only one of the two methods used for measuring the capillary bleeding time showed a slight prolongation compared to the starting value (mean = 77 sec) to 88 sec (p < 0.05). The most significant influence was seen on the platelet aggregation induced by 1 I.U./ml thrombin, whereby the aggregation maximum decreased from 92.0% (mean) to 12.2% (p < 0.001). Whereas the latter result has to be interpreted primarily as a consequence of the anti-thrombin effect of heparin, the results of the other tests in summary illustrate the clinical unimportant influence of heparin on platelet function in dogs.

Adenosine Diphosphate↗

PubMed2000

[Phamacokinetics of low-molecular-weight heparins Fragmin D in dogs].

The pharmacokinetics of a low molecular weight heparin (LMWH; Fragmin D) was studied in dogs after intravenous and subcutaneous administration, based on antifactor Xa- (anti-fXa-) activity. Each dosage was examined in 5 adult Beagles. After intravenous application of 25, 50 and 100 anti-fXaU./kg body weight (BW) the mean peak plasma heparin activity of 0.52 +/- 0.12 (x +/- s), 1.08 +/- 0.23 and 1.86 +/- 0.17 anti-fXaU./ml, respectively, was measured. After subcutaneous application of 50, 100 and 200 anti-fXaU./kg BW maximum heparin activity in the plasma was determined after 144-216 minutes (mean values) of 0.28 +/- 0.01, 0.52 +/- 0.06 or 1.09 +/- 0.20 anti-fXaU./ml. Intravenous application of LMWH has a short plasma terminal half-life (t50) between 49 and 76 minutes which depended on the dosage. After administration of 50 anti-fXaU./kg BW (74 minutes) and 100 anti-fXaU./kg BW (76 minutes) no essential difference was shown. A distinctly longer t50 was found after subcutaneous injection. After injection of 50, 100 and 200 anti-fXaU./kg BW t50 values of 81, 123 and 182 minutes were calculated. According to this, with increasing dosage a decrease of the total clearance was found for both application routes. The apparent volume of distribution after intravenously applicated LMWH ranged between 50 and 70 ml/kg BW. The absolute bioavailability calculated for the subcutaneous NMH-injection of 50 and 100 anti-fXaU./kg BW was 107% and 104%, respectively.

Animals↗

PubMed1999

Characterization of catalytic centre mutants of macrophage migration inhibitory factor (MIF) and comparison to Cys81Ser MIF.

Macrophage migration inhibitory factor (MIF) displays both cytokine and enzyme activities, but its molecular mode of action is still unclear. MIF contains three cysteine residues and we showed recently that the conserved Cys57-Ala-Leu-Cys60 (CALC) motif is critical for the oxidoreductase and macrophage-activating activities of MIF. Here we probed further the role of this catalytic centre by expression, purification, and characterization of the cysteine-->serine mutants Cys60Ser, Cys57Ser/Cys60Ser, and Cys81Ser of human MIF and of mutants Ala58Gly/Leu59Pro and Ala58Gly/Leu59His, containing a thioredoxin (Trx)-like and protein disulphide isomerase (PDI)-like dipeptide, respectively. The catalytic centre mutants formed inclusion bodies and the resultant mutant proteins Cys57Ser/Cys60Ser, Ala58Gly/Leu59Pro, and Als58Gly/Leu59His were only soluble in organic solvent or 6 m GdmHCl when reconstituted at concentrations above 1 microgram.mL-1. This made it necessary to devise new purification methods. By contrast, mutant Cys81Ser was soluble. Effects of pH, solvent, and ionic strength conditions on the conformation of the mutants were analysed by far-UV CD spectropolarimetry and mutant stability was examined by denaturant-induced unfolding. The mutants, except for mutant Cys81Ser, showed a close conformational similarity to wild-type (wt) MIF, and stabilization of the mutants was due mainly to acid pH conditions. Intramolecular disulphide bond formation at the CALC region was confirmed by near-UV CD of mutant Cys60Ser. Mutant Cys81Ser was not involved in disulphide bond formation, yet had decreased stability. Analysis in the oxidoreductase and a MIF-specific cytokine assay revealed that only substitution of the active site residues led to inactivation of MIF. Mutant Cys60Ser had no enzyme and markedly reduced cytokine activity, whereas mutant Cys81Ser was active in both tests. The Trx-like variant showed significant enzyme activity but was less active than wtMIF; PDI-like MIF was enzymatically inactive. However, both variants had full cytokine activity. Together with the low but nonzero cytokine activity of mutant Cys60Ser, this indicated that the cytokine activity of MIF may not be tightly regulated by redox effects or that a distinguishable receptor mechanism exists. This study provides evidence for a role of the CALC motif in the oxidoreductase and cytokine activities of MIF, and suggests that Cys81 could mediate conformational effects. Availability and characterization of the mutants should greatly aid in the further elucidation of the mechanism of action of the unusual cytokine MIF.

Base Sequence↗

PubMed1999

[Changes in platelet concentrates from dogs due to storage. II. Biochemical changes in concentrate plasma].

Platelet concentrate (PC) obtained from dogs with an automatic cell separator was stored in C4-cell separation sets with low gasdiffusionable Polyvinylchlorid (PVC) storage containers or in C4L-sets developed for storage with high gasdiffusionable Polyolefin(PO) containers, respectively. PC were stored for 10 days under permanent agitation at 22 degrees C (C4/22 degrees C, n = 10; C4L/22 degrees C, n = 11) or at 4 degrees C (C4L/4 degrees C, n = 6), respectively. Measurements were carried out directly after production of the PC, after 6 hours and then daily during the 10-day storage period. In the second part of this paper the results of pH, the concentration of bicarbonate, glucose, lactate and potassium ions as well as the activity of lactate dehydrogenase (LDH) are presented. The varying duration and intensity of the energy metabolism of the platelets and different part of glycolysis became obvious by the consumption of glucose and production of lactate, which differed significantly between the different storage conditions. Resulting from this, the mean pH decreased under the limit prescribed for human PC (pH = 6.3) already after a storage period of 3 days due to the slight capacity of gas diffusion in PVC-containers (C4/22 degrees C). In the PO-containers the pH fell below this limit at 22 degrees C (C4L/22 degrees C) after a storage period of 5 days and at 4 degrees C (C4L/4 degrees C) after 10 days. The latter reflects the high gas diffusion capacity of the PO-containers and the decreased metabolism activity at 4 degrees C. The increase of activity of LDH and of the concentration of potassium ions, which are localized in the cytosol of platelets, depended also on the different storage conditions and, thereby, reflected the different rapidity of increasing membrane permeability or the destruction of the cell membrane, respectively. The results of this study nearly are in agreement with the changes of platelet function shown in part I. Biochemical changes occur in canine platelet concentrates similar to those in human platelet concentrates during storage in dependency of the storage conditions, in part even with a higher rate or in a higher extent.

Animals↗

PubMed1999

[Effect of unfractionated and low-molecular-weight heparin on platelet aggregation and in vitro bleeding time in dogs].

The aim of the present study was to examine the influence of unfractionated heparin (UFH) and low molecular weight heparin (LMWH) on the function of canine thrombocytes. This was performed by adding different concentrations of UFH or LMWH to platelet rich plasma (PRP) or blood of healthy dogs: 0.2, 0.5, 1, 2, 5, 10, 20, and 50 I.U./ml UFH or Anti-FXaU/ml LMWH, respectively (aggregation induced by thrombin additionally: 0.025, 0.05, and 0.1 I.U./ml UFH or Anti-FXaU/ml LMWH.) Platelet aggregation induced by ADP, collagen or thrombin with the BORN method (n = 11) as well as the in vitro bleeding time using the analyzer PFA-100 (n = 5) were examined. Additionally, a specific test assay for ADP induced platelet aggregation was performed which enabled an individual adjustment of the aggregation maximum at 30-40% in the control measurements (n = 6). The most prominent effect was noted in the platelet aggregation induced by thrombin. The aggregation maximum of the platelet aggregation induced by 1 I.U./ml thrombin (final concentration) was significantly lower in all of the testet UFH concentrations > or = 0.025 I.U./ml UFH in comparison to control measurements. If the aggregation was induced by 10 I.U./ml thrombin a significant reduction of the aggregation maximum was restricted to UFH concentrations > or = 0.5 I.U./ml. The addition of LMWH to canine PRP resulted in a distinct decrease (p < 0.01) of the maximum aggregation induced by 1 I.U./ml thrombin in concentrations > or = 0.2 Anti-FXaU/ml LMWH. A slight decrease of the maximum aggregation induced by collagen was only found for UFH at activities > or = 20 I.U./ml. No significant systematic influence could be demonstrated for LMWH on the aggregation induced by collagen as well as for LMWH and UFH on the platelet aggregation induced by ADP. Capillary in vitro bleeding time (closure time) was prolonged only after adding high concentrations of UFH (> or = 10 I.U./ml) and LMWH (> or = 50 Anti-FXaU/ml) to the sample material. The results document the unimportant influence of therapeutic levels of UFH and LMWH on platelet function in dogs. Therefore, the remarkable inhibition of the aggregation induced by thrombin reflects mainly the antithrombin effect.

Animals↗

PubMed1999

[Pharmacokinetics of an unfractionated heparin (Liquemin) in the dog after intravenous and subcutaneous application based on heparin activity].

In this study pharmacokinetic data for the unfractionated heparin Liquemin were obtained after intravenous and subcutaneous application. Each dosage was examined in 5 healthy, adult Beagle dogs. After intravenous application of 25, 50 and 100 I.U./kg body weight heparin plasma activity of 0.65 +/- 0.15 I.U./ml (mean +/- s), 0.91 +/- 0.10 I.U./ml and 1.94 +/- 0.22 I.U./ml was measured. Subcutaneous applications of 250, 500 or 750 I.U./kg revealed maximum plasma heparin activities of 0.25 +/- 0.10, 0.60 +/- 0.15 and 1.29 +/- 0.24 I.U./ml. The maximum heparin activity in the plasma was observed after 3.8 +/- 1.1 (250 und 500 I.E./kg) or 4.0 +/- 1.0 hours (750 I.E./kg), respectively. Intravenously applicated heparin has a short terminal half-life time (t50) between 22 and 44 minutes. The t50 after subcutaneous application of heparin was distinctly longer. After 250, 500 or 750 I.U./kg the t50 was 3.7 +/- 2.4, 3.5 +/- 1.2 or 5.3 +/- 2.4 hours. Corresponding to this result a lower total clearance (Cltot) was found with increasing doses. Especially the Cltot after subcutaneous injection decreased from 2.08 +/- 0.73 ml/min/kg (250 I.E./kg) to 0.83 +/- 0.27 ml/min/kg (750 I.E./kg). The volume of distribution of heparin corresponded approximately to the plasma volume. The total bioavailability of subcutaneously administered UFH was 53-100% depending on the dosage.

Animals↗

PubMed1998

Disulfide analysis reveals a role for macrophage migration inhibitory factor (MIF) as thiol-protein oxidoreductase.

The molecular mechanism of action of macrophage migration inhibitory factor (MIF), a cytokine with a critical role in the immune and inflammatory response, has not yet been identified. Here we report that MIF can function as an enzyme exhibiting thiol-protein oxidoreductase activity. Using a decapeptide fragment of MIF (MF1) spanning the conserved cysteine sequence motif Cys57-Ala-Leu-Cys60 (CALC), Cys-->Ser mutants (C57S MIF, C60S MIF, and C57S/C60S MIF) of human MIF (wtMIF), and alkylated wtMIF, we show that this activity is mediated by the CALC region and is important for the macrophage-activating properties of MIF. Both wtMIF and MF1 were demonstrated to form an intramolecular disulfide bridge. Using two common oxidoreductase assays, MIF was shown to enzymatically catalyze the reduction of insulin and 2-hydroxyethyldisulfide (HED). Examination of wtMIF and the mutants by far-UV circular dichroism spectroscopy (CD) together with denaturation studies showed that substituting or reducing the cysteine residues of CALC led to a reduced conformational stability of MIF but did not significantly change its overall conformation. A functional role for the CALC region was revealed by subjecting the mutants and alkylated wtMIF to the enzymatic assays. Mutant C60S did not have any enzymatic activity while mutant C57S had a reduced activity. Thiol-modified wtMIF that was alkylated under oxidizing conditions was found to have full enzymatic activity, whereas alkylation of wtMIF under reducing conditions completely eliminated MIF-mediated redox activity. Importantly, further physiological relevance of the disulfide motif was obtained by examining the mutants and alkylated MIF in an immunological assay that involved the macrophage-activating properties of MIF. In this test, mutant C60S was essentially inactive and mutant C57S was partly active, indicating together that at least some of the cytokine-like biological activities of MIF are dependent on the presence of cysteine 57 and 60. Again, use of the alkylated MIF species confirmed the role of the cysteine motif for this MIF activity. In conclusion, our results argue (a) that MIF exhibits enzymatic oxidoreductase activity, (b) that this activity is dependent on the presence of the catalytic center that is formed by cysteine residues 57 and 60, and (c) that certain MIF-mediated immune processes are due to the cysteine-mediated redox mechanism.

Animals↗

PubMed1998

Specific reduction of insulin disulfides by macrophage migration inhibitory factor (MIF) with glutathione and dihydrolipoamide: potential role in cellular redox processes.

The molecular mechanism of action of MIF, a cytokine that plays a critical role in the host immune and inflammatory response, has not yet been identified. We recently demonstrated that MIF is an enzyme that exhibits oxidoreductase activity by a cysteine thiol-mediated mechanism. Here we further investigated this function by examining the reduction of insulin disulfides by wild-type human MIF (wtMIF) using various substrates, namely glutathione (GSH), dihydrolipoamide, L-cysteine, beta-mercaptoethanol and dithiothreitol. The activity of wtMIF was compared to that of the relevant cysteine mutants of MIF and to two carboxy-truncated mutants. Only GSH and dihydrolipoamide were found to serve as reductants, whereas the other substrates were not utilized by MIF. Reduction of insulin disulfides by MIF was closely dependent on the presence of the Cys57-Ala-Leu-Cys60 (CALC) motif-forming cysteines C57 and C60, whereas C81 was not involved (activities: 51+/-13%, 14+/-5%, and 70+/-12% of wtMIF, respectively, and 20+/-3% for the double mutant C57S/C60S). Confirming the notion that the activity of MIF was dependent on the CALC motif in the central region of the MIF sequence, the C-terminal deletion mutants MIF(1-105) and MIF(1-110) were found to be fully active. The favored use of GSH and dihydrolipoamide indicated that MIF may be involved in the regulation of cellular redox processes and was supported further by the finding that MIF expression by the cell lines COS-1 and RAW 264.7 was significantly induced upon treatment with the oxidant hydrogen peroxide.

Amino Acid Sequence↗

PubMed1998

Cross-linking and mutational analysis of the oligomerization state of the cytokine macrophage migration inhibitory factor (MIF).

The structure of the cytokine MIF has been investigated by X-ray crystallography, NMR, and biochemical methods with conflicting results regarding the structural and functional oligomerization state of this protein. Determination of the oligomeric state(s) is important for understanding more precisely the molecular mechanism of MIF action. To address this issue, we performed cross-linking of human and mouse MIF and selected mutants by various methods and analyzed the oligomerization by SDS-PAGE and gel filtration. MIF was found to form a mixture of monomeric, dimeric, and trimeric states at physiological concentrations, with the monomer and dimer representing the major species. Similar results were obtained when the carboxy-truncated mutants MIF(1-104) and MIF(1-109) were examined, indicating that the C-terminus of MIF is not critical for trimer stabilization. Cross-linking analysis of the isosteric Cys --> Ser mutants C56S and C80S of human MIF resulted in a similar oligomer distribution, whereas substitution of Cys59 led to a significant reduction in the dimeric and trimeric forms, indicating that the hydrophobic region around Cys59 is important for the oligomerization of MIF. Together, our data argue that physiological MIF solutions contain a mixture of monomers, dimers, and trimers.

Animals↗

PubMed1998

Trisomy 1 in a canine acute leukemia indicating the pathogenetic importance of polysomy 1 in leukemias of the dog.

We report on a canine acute myeloid leukemia showing a trisomy 1 and a t(X;8) as clonal cytogenetic abnormalities. Comparing these results with similar cases of canine leukemias reported in the literature, one realizes that trisomy 1 is a recurrent cytogenetic finding in canine acute leukemia. Trisomy 1 may be a specific anomaly associated with either etiology or progression of this disease. As with comparable human neoplasms, cytogenetic investigations could be of diagnostic and prognostic significance for canine hematopoietic diseases. Moreover, as trisomies are recurrent cytogenetic findings in human leukemias, as well, comparative gene mapping in future studies may help to focus the etiologic basis of the disease to particular chromosomal segments instead of whole chromosomes.

Acute Disease↗

PubMed1998

[Hemostatic disorders as a complication of autoimmune hemolytic anemia in dogs].

In eleven of 15 investigated dogs suffering from autoimmune haemolytic anaemia an increased concentration of soluble fibrin (p < 0.001) and in all of these 15 dogs an increased concentration of fibrin(ogen) degradation products was observed. The increased turnover indicated by these markers of intravascular coagulation and hyperfibrinolysis was decompensated in approximately the half of the patients which was shown by a decrease of platelet count, activity of different coagulation factors as well as of the inhibitor antithrombin III. The consumption induced decrease of activity was especially related to prekallikrein and high molecular weight kininogen reflecting an activation mainly of the initial intrinsic system by the haemolytic anaemia and was also indicated by the activated partial thromboplastin time which was distinctly prolonged in some cases. The results of this study demonstrate the necessity of an anticoagulant therapy in dogs suffering from autoimmune haemolytic anaemia.

Anemia, Hemolytic, Autoimmune↗

PubMed1998

[Changes in hemostasis of dogs with acute lymphoblastic leukemia].

Twelve dogs suffering from acute lymphoblastic leukaemia were investigated concerning the following tests: platelet count, prothrombin time (PT, standard test, modified test), activated partial thromboplastin time (APTT), activity of the individual coagulation factors II, V, VII, X, VIII:C, IX, XI, XII, prekallikrein, and high-molecular weight kininogen, the activity of antithrombin III (AT III), protein C, plasminogen, and alpha 2-antiplasmin as well as concentration of fibrinogen, soluble fibrin and fibrin(ogen) degradation products (FDP). All patients showed a decreased platelet count due to suppression of megakaryopoesis by infiltration of the bone marrow with leukaemic cells. In addition, in most of the patients a moderate activity decrease of one or more individual coagulation factors has been found, especially regarding factor II (median, x0.50 = 51%, p = 0.0001), but also factors X (x0.50 = 71%, p = 0.0003) and XI (x0.50 = 68%, p = 0.0006). This was reflected by the APTT and the PT activity (modified test), which were prolonged or decreased, respectively, in the majority of the cases. Furthermore, the activity of AT III and of plasminogen was distinctly diminished (p < 0.001). Like the concentration of FDP, the plasma level of soluble fibrin was significantly higher than in normal dogs (p < 0.001). This indicates that besides thrombocytopenia disseminated intravascular coagulation occurs frequently in dogs with acute lymphoblastic leukaemia and is a main cause for the decreased activity of several plasmatic components of the haemostatic system. The lack of correlation between the concentration of soluble fibrin as an indicator of intravascular coagulation and the total blast cell count (rS = 0.011) shows the importance of other factors like degree of cell lysis as well as participation of organs such as the liver for generation of consumption coagulopathy in dogs with acute lymphoblastic leukaemia.

Angiotensins↗

PubMed1998

[Alterations of hemostasis in liver cirrhosis of the dog].

In seven dogs with histologically proven liver cirrhosis the activity of the single coagulation factors with the exception of factor VIII:C, of the inhibitors antithrombin III and protein C as well as plasminogen and alpha 2-antiplasmin was distinctly lower than in the control group (p < 0.0001). The changes of the factors VII [median (x0.50) = 17 %] and X (x0.50 = 18 %) as well as of protein C (x0.50 = 15 %) were particularly pronounced. Diminution of activity certainly exceeded also in nearly all of the remaining haemostatic proteins the decrease of albumin concentration. Besides the shorter half life time, this reflected an increased consumption in consequence of intravascular coagulations and fibrinolysis. The latter could also be seen from the significantly increased concentrations of soluble fibrin and fibrin(ogen) degradation products. Therefore, the alterations of the haemostatic system measured in dogs in many details were in accordance with findings in human beings suffering from liver cirrhosis.

Animals↗

PubMed1998

[Determination of fibrinogen levels in the horse with the heat-precipitation methods of Schalm and Millar].

The purpose of this study was to investigate the usefulness of two heat-precipitation techniques (Schalm- and Millar-method) as screening tests to measure plasma fibrinogen concentration in horses. Based on the measurement of samples from 108 different horses, the coefficient of correlation (CC) for the relationship between the results with the Schalm- and with the reference-method (Jacobsson) were much lower (r = 0.78) than between the Millar- and Jacobsson-method (r = 0.94). Furthermore the Schalm-method was less precise as reflected by the greater coefficient of variation (CV, within-run precision) with a sample of low-limit fibrinogen concentration (CV = 47.4 %) and with a sample of high fibrinogen concentration (CV = 35.6 %) than the Millar-method (CV = 11.1 resp. 2.9 %). In 40 healthy horses, aged 3 to 19 years, fibrinogen values ranged from 1.82 to 4.94 g/l (2.5-97.5 %-quantil). The Millar-Method is recommended as a simple and suitable heat-precipitation assay for fibrinogen determination in the horse.

Animals↗

PubMed1998

[Effect of the evaluation parameter on sensitivity of resonance thrombography of thrombocytopenia in dogs].

Based on 109 blood samples taken from 36 dogs suffering from thrombocytopenia resonance thrombography with the resonance thrombograph RTG 801 (von Hoerner und Sulger Electronic GmbH, Schwetzingen; manufacturer: Fresenius AG, Bad Homburg) was distinctly more sensitive and more closely correlated to the platelet count using an optimized parameter of the resonance thrombogramm (RTG) in comparison to usual parameters. Nevertheless, clinical requirements regarding samples with platelet counts > 25,000/microliter were not fulfilled. Out of 13 samples with reduced platelet count and simultanous extended capillary bleeding time, depending on the used parameter a maximum of 9 samples could be detected as pathological by the RTG. The normal RTG in part of the cases with clearly altered primary haemostasis contrasts to the exclusive use of RTG in the screening of thrombocytopenia in dogs.

Animals↗

PubMed1998

[Disseminated intravascular coagulation and hyperfibrinolysis in dogs with metastasizing mammary carcinoma].

The alterations of the haemostatic system (platelet count, activated partial thromboplastin time [APTT], thromboplastin time [standard test, modified test], thrombin time, fibrinogen concentration, activity of the coagulation factors II, V, VII, X, VIII:C, IX, XI, XII, of prekallikrein, high molecular weight kininogen, antithrombin III, protein C, plasminogen and alpha 2-plasmin inhibitor, concentration of soluble fibrin and fibrin(ogen) degradation products [FDP], resonance thrombogram) were described in seven dogs with haemorrhagic diathesis in consequence of an infiltrative, growing mammary carcinoma with multifocal invasion of lymphatic and blood vessels. In most of the cases metastases in different organs could be demonstrated. In every case a serious stage of disseminated intravascular coagulation and hyperfibrinolysis was existent. This was indicated by the distinctly increased concentration (p < 0.0001) of soluble fibrin (27.7 [16.0-79.2] micrograms/ml, median [minimum-maximum], reference range [RR.]: < 9.4 micrograms/ml) and FDP (340 [50-860] micrograms/ml, RR.: < 18 micrograms/ml) as well as a diminished plasma level of nearly all components of the coagulation and fibrinolytic system concerning especially the concentration of fibrinogen (0.16 [0.01-0.46] g/l, RR.: 1.17-3.09 g/l), the activity of factors V (30 [21-40]%, RR.: 75-158%) and VIII:C (9 [4-16]%, RR.: 72-136%) as well as the activity of protein C (8 [3-13]%, RR.: 68-139%) (each: p < 0.0001).

Animals↗

PubMed1997

Structure activity studies of the cytokine macrophage migration inhibitory factor (MIF) reveal a critical role for its carboxy terminus.

Carboxy-truncated mutants of human MIF (MIF(1-104) and MIF(1-109)) were used in structure activity studies. CD spectroscopy revealed an overall structural similarity between the mutants and MIF. Denaturant-induced unfolding demonstrated that the C-terminus contributed significantly to the conformational stability of MIF. This appears to be due to the formation of two C-terminal beta-strands. The mutants were enzymatically active, exhibiting half of the enzymatic redox activity of MIF. However, immunological analysis showed that deletion of both 5 and 10 C-terminal residues resulted in loss of the macrophage activating properties of MIF, providing functional evidence that the C-terminus is important for immunological activity and trimer formation. A more detailed study of the C-terminus may assist in identifying the molecular basis for the immunological and enzymatic activities of MIF.

Amino Acid Sequence↗