Evaluation of Mannose-Binding Lectin is a Useful Approach to Predict the Risk of Infectious Complications Following Autologous Hematopoietic Stem Cell Transplantation.

Hematopoietic stem cell transplantation (HSCT) associated immunocompromised state carries high risk of infectious complications. Mannose-binding lectin (MBL) is an acute phase protein involved in innate immune response. Serum MBL level is genetically determined and quite stable. According to literature, significant association was shown between low MBL concentrations and serious infections. The association between serum MBL level and frequency and severity of infections was studied in 186 patients following autologous HSCT. Double-monoclonal antibody sandwich enzyme-linked immunosorbent assay was used to determine MBL antigen level in sera. MBL levels were measured around 100 days following transplantation, in a period without active infection. Twenty-one patients (11%) were MBL deficient. The median time of first infection and number of infections during the first year post-transplantation were not significantly different between patients with MBL deficiency and those without MBL deficiency. The occurrence and number of infections after HSCT correlated with the MBL/C-reactive protein ratio. The number of severe infections was not higher among those with MBL deficiency. The occurrence of infections after the pre-engraftment period during the first year post-transplantation was significantly different in patient groups separated by MBL cut-off level. The MBL/C-reactive protein ratio might be a useful marker of infectious complications. MBL measurement may be helpful in antibiotic treatment. In case of MBL deficiency, earlier and more intensive treatment may be indicated.

Identification of a VWF peptide antagonist that blocks platelet adhesion under high shear conditions by selectively inhibiting the VWF-collagen interaction.

BACKGROUND: Because the collagen-VWF-GPIb/IX/V axis plays an important role in thrombus formation, it represents a promising target for development of new antithrombotic agents. OBJECTIVES: We used phage display to identify potential small peptides that interfere with the VWF-collagen binding and might serve as lead products for the development of possible oral antithrombotic compounds. METHODS: A random linear heptamer peptide library was used to select VWF-binding peptides. RESULTS: We identified a phage clone, displaying the YDPWTPS sequence, further referred to as L7-phage, that bound to VWF in a specific and a dose-dependent manner. This L7-phage specifically inhibited the VWF-collagen interaction under both static and flow conditions. Epitope mapping using deletion mutants of VWF revealed that the L7-phage does not bind to the known collagen-binding A3 domain within VWF, but to the more carboxyterminal situated C domain. This inhibition was not due to steric hindrance of the A3 domain-collagen interaction by the L7-phage. Indeed, a tetrabranched multi-antigen peptide (MAP) presenting four copies of the peptide, but not the scrambled MAP, also inhibited VWF-collagen interaction under conditions of high shear stress at a concentration of 148 nmol L(-1). CONCLUSIONS: Based on these results, we conclude that we have identified the first peptide antagonist that binds to the VWF C domain and by this specifically inhibits the VWF binding to collagen, suppressing platelet adhesion and aggregation under high shear conditions. As a consequence, this peptide and its future derivates are potentially interesting antithrombotic agents.

4-thio-deoxyuridylate-modified thrombin aptamer and its inhibitory effect on fibrin clot formation, platelet aggregation and thrombus growth on subendothelial matrix.

BACKGROUND: The consensus thrombin aptamer C15-mer is a single-stranded DNA of 15 nucleotides [d(GGTTGGTGTGGTTGG)] that was identified by the selection of thrombin-binding molecules from a large combinatorial library of oligonucleotides. It is capable of inhibiting thrombin at nanomolar concentrations through binding to a specific region within thrombin exosite 1. As has been shown in our earlier studies, the 4-thio-deoxyuridylate (s4dU)-containing oligonucleotides have high affinity for a number of proteins, due to the reduced hydrophilic character of the modified oligonucleotide. METHODS: Three different analogs of the original thrombin-inhibiting sequence, in which some of the thymidylate residues were replaced by 4-thio-deoxyuridylates, were synthesized. The inhibitory effect of modified aptamers was tested on thrombin-catalyzed fibrin clot formation and fibrinopeptide A release from fibrinogen, thrombin-induced platelet aggregation/secretion, and the formation of thrombus on coverslips coated with human collagen type III, thrombin-treated fibrinogen or subendothelial matrix of human microvascular endothelial cells. RESULTS: As compared with the C15-mer, the analog with the sequence GG(s4dU)TGG(s4dU)G(s4dU)GGT(s4dU)GG (UC15-mer) showed a 2-fold increased inhibition of thrombin-catalyzed fibrin clot formation, fibrinopeptide A release, platelet aggregation and secretion in human plasma and thrombus formation on thrombin-treated fibrinogen surfaces under flow conditions. Concerning the inhibition of thrombin-induced fibrin formation from purified fibrinogen and activation of washed platelets, UC15-mer was 3-fold and twelve-fold more effective than C15-mer, respectively. CONCLUSION: The replacement of four thymidylate residues in C15-mer by 4-thio-deoxyuridylate resulted in a new thrombin aptamer with increased anticoagulant and antithrombotic properties.

A monoclonal antibody directed against human von Willebrand factor induces type 2B-like alterations.

We have previously described a monoclonal antibody (mAb), 1C1E7, against von Willebrand factor (VWF), that increases ristocetin-induced platelet aggregation (RIPA) and induces a preferential binding of the high-molecular-weight multimers of VWF to platelet GPIb. Further investigations using a rotational viscometer at a shear rate of 4000 s(-1) could now demonstrate that shear-induced platelet aggregation (SIPA) is significantly increased with 1C1E7 and that this could be completely inhibited by the anti-GPIb mAb 6D1. In contrast, platelet adhesion to a collagen surface at a shear rate of 2600 s(-1), using a rectangular perfusion chamber, was significantly inhibited in the presence of 1C1E7. When citrated whole blood was incubated with 1C1E7, a spontaneous binding of VWF to the platelet GPIb could be demonstrated by flow cytometric analysis. Parallel to this, a decrease of the highest molecular weight multimers of VWF in the plasma was found. Platelets with bound VWF on their surface were able to form macroaggregates but were no longer able to adhere. These phenomena are very similar to the alterations described in von Willebrand's disease type 2B. The epitope of this mAb could be localized to the N-terminal part of the subunit; therefore a distant conformational change in the A1 domain of VWF is suggested.

The most severe forms of Perthes' disease associated with the homozygous Factor V Leiden mutation.

It has recently been postulated that thrombophilia may have a role in the aetiology of Perthes' disease. The published reports, however, remain conflicting. In this study a retrospective analysis of the coagulation parameters was made in 47 patients with Perthes' disease and the results compared with the clinical data. Five patients with Factor V Leiden mutation were found (10.6%) and surprisingly four of them had a homozygous pattern. These four patients showed the most severe form of the disease, Catterall group IV, with flattening of the entire epiphysis, involvement of the metaphysis, shortening and broadening of the femoral neck, trochanteric overgrowth and developed mushroom-shaped aspherical laterally displaced femoral heads in dysplastic acetabula. We would like to suggest that the homozygous form of Factor V Leiden mutation has some role in the clinical course of Perthes' disease and particularly its most severe form.

Factor XIII mediates adhesion of platelets to endothelial cells through alpha(v)beta(3) and glycoprotein IIb/IIIa integrins.

Coagulation factor XIII (FXIII) is a transglutaminase that catalyzes crosslink formation in fibrin clots. Endothelial cells (EC) were demonstrated to bind FXIII via their alpha(v)beta3 integrin receptor. FXIII was also shown to bind platelet glycoprotein IIb/IIIa receptor. In the present study, we analyzed if FXIII can mediate platelet-EC interaction. Both FXIII and activated FXIII (FXIIIa) bound to EC monolayers; this binding was enhanced by the addition of Mn2+ and was inhibited by the monoclonal antibody L609 against alpha(v)beta3 integrin. Normal washed platelets also bound surface-immobilized or soluble FXIII and FXIIIa, and the binding was GPIIb/IIIa dependent. The effect of FXIII concentrate (Fibrogammin-P) treatment on the interaction of ECs with platelets from six FXIII-deficient patients was studied. Patients' platelets were radiolabeled with 3H-Adenine, washed, resuspended in autologous plasma and allowed to adhere to immortalized EC line EAhy926. Adhesion of platelets from FXIII-deficient patients to ECs increased 1.7+/-0.4-fold (P=.01) following intravenous infusion of FXIII concentrate. Similarly, addition of 1 U/ml of FXIII concentrate to the patients' PRP in vitro increased the adhesion 1.8+/-0.5-fold (P=.008). Preincubation of the EC monolayers with increasing concentrations of either FXIII or FXIIIa augmented the adhesion of normal washed platelets to ECs in a dose-dependent manner. At 10 U/ml of EC-bound FXIII or FXIIIa, platelet adhesion enhanced 1.7+/-0.25-fold (P=.03) and 2.5+/-0.5-fold (P=.02), respectively. The increase in platelet adhesion was completely abolished by pretreatment of ECs with the anti-alpha(v)beta3 antibody L609 or by preincubation of the platelets with the GPIIb/IIIa inhibitor Abciximab. Taken together, our data indicate that FXIII mediates the interaction of platelets with ECs by bridging between endothelial alpha(v)beta3 and platelet GPIIb/IIIa integrins. This interaction may be relevant for tissue remodeling and wound repair after vascular injury in FXIII-deficient patients.

Organization of the glycoprotein (GP) IIb/IIIa heterodimer on resting human platelets studied by flow cytometric energy transfer.

Glycoprotein IIb/IIIa is a heterodimer of glycoproteins IIb and IIIa which serves as the inducible receptor for fibrinogen and other adhesive proteins at the surface of platelets. Although a model of the quaternary structure of the GPIIb/IIIa molecule has been constructed in solution by Calvete et al. [Biochem. J. 282 (1992) 523], a corresponding model at the surface of intact platelets is still missing. In the present work conformation and lateral distribution of the GPIIb/IIIa heterodimer were studied at a nanometer resolution on the surface of resting human platelets under physiological conditions. The experiments were based on dual wavelength flow cytometric detection of fluorescence resonance energy transfer and application of a panel of monoclonal antibodies raised against well described binding sites. Monodisperse distribution of the GPIIb/IIIa heterodimer has been observed and a detailed three-dimensional proximity map of antibody binding sites was constructed on the platelet membrane, under physiological conditions, for the first time. Our data support the view that the GPIIb subunit is in a bent conformation. A detailed analysis of the K(d)-values and the number of binding sites for a set of monoclonal antibodies was also carried out giving supplementary data for the topology of the binding sites. Our results provide a refinement of the membrane-topology of the GPIIb/IIIa heterodimer.

Selection of phages that inhibit vWF interaction with collagen under both static and flow conditions.

Phages from a pentadecamer phage display library were selected for binding to vWF by affinity panning. Bound phages were selectively eluted with human collagen type I. After the third round of panning 95% of individual phage clones bound to vWF. The B8-phage inhibited the binding of collagen to vWF with an IC50 of 0.6 x 10(10) phages/ml, and of vWF to collagen with an IC50 of 1.0 x 10(10) phages/ml at 0.5 microg/ml vWF. Under flow conditions, 1.5 x 10(11) B8-phage/ml nearly completely inhibited platelet deposition on a human collagen type I coated surface at a shear rate of 1200 s(-1), while phages without an insert had no effect. The peptide corresponding to the one displayed on the B8-phage competed with the phage for binding to vWF with an IC50 of 30 microg/ml (16 microM). The peptide furthermore inhibited vWF-binding to collagen with a maximum of 40% at a concentration of 1.25 mg/ml (650 microM), higher concentrations of peptide could not improve this. We thus have selected phages that are potent vWF-binders and that can be used as tools to detect vWF, to inhibit vWF-collagen interaction and to further analyse the role of vWF-collagen binding.

Epitope mapping of inhibitory antibodies against platelet glycoprotein Ibalpha reveals interaction between the leucine-rich repeat N-terminal and C-terminal flanking domains of glycoprotein Ibalpha.

The interaction of von Willebrand factor (vWF) with the platelet receptor glycoprotein Ibalpha (GPIbalpha) is important for platelet adhesion at high shear stress. Two functionally important antigenic areas within GPIbalpha were identified through the characterization of 5 new inhibitory anti-GPIb monoclonal antibodies (mAbs). The binding sites of 3 of these anti-GPIb mAbs, which were intercompeting and potently inhibiting shear stress-induced binding of vWF, were mapped within the N-terminal amino acid (aa) 1-59 area by the use of canine-human chimeras. These antibodies, however, had little or no effect (approximately 40% inhibition) on the binding of vWF induced by either botrocetin or ristocetin. On the other hand, the anti-GPIb mAbs 24G10 and 6B4, which blocked GPIb-vWF binding under all conditions examined, bound to 2 different regions of GPIbalpha, aa 1-81 and aa 201-268, respectively. The epitope for 6B4 was further narrowed by phage display revealing 2 sets of peptide sequences aligning within aa 259-262 and aa 230-242. In the latter region of GPIbalpha, the gain-of-function platelet-type von Willebrand disease (PT-vWD) mutations have been identified. Alignment was partially confirmed because the binding of 6B4 to recombinant GPIbalpha fragments carrying either one of the PT-vWD mutations was considerably impaired but not completely abolished. In contrast, mAb 24G10 bound more strongly to mutant PT-vWD GPIbalpha. However, although 24G10 competed with 6B4 for binding to platelets, it bound to an epitope within aa 1-81 of GPIbalpha. In conclusion, 2 functionally important areas within GPIbalpha were identified: one localized within the leucine-rich repeat N-terminal aa 1-59 area and one composed of residues aa 1-81 in close contact with aa 201-268. Moreover, further support is provided for the existence of an intramolecular interaction between the N-terminal flanking (aa 1-81) and C-terminal flanking (aa 201-268) regions. (Blood. 2001;98:652-660)

The subendothelium of the HMEC-1 cell line supports thrombus formation in the absence of von Willebrand factor and collagen types I, III and VI.

The macromolecular composition of the extracellular matrix (ECM) produced by the human microvascular endothelial cell line (HMEC-1) was determined by ELISA and its thrombogenicity was studied in blood perfusion assays. Results were compared with those obtained with the ECM produced by human umbilical vein endothelial cells (HUVEC). The HMEC-1's ECM contains collagen type IV, fibronectin, laminin and thrombospondin, but no detectable levels of collagen types I, III and VI, or von Willebrand factor (vWF), whereas all these components were found in the ECM synthesized by HUVEC. HMEC-1's ECM was perfused with low-molecular-weight heparin-anticoagulated blood at two wall shear rates (650/s and 2,600/s), representative of moderate and high arterial wall shear rates, in parallel plate flow chambers for 5 min. This resulted in the formation of large platelet aggregates, compared to essentially a monolayer of adherent platelets on HUVEC's ECM. Interestingly, large thrombi were formed at 2,600/s when HMEC-1's ECM was perfused with the blood of a patient with severe type III von Willebrand disease lacking both plasma and platelet vWF, indicating that vWF was not absolutely required for thrombus formation on this matrix. Thrombin generated on the HMEC-1's ECM contributed importantly to the large platelet thrombi formed, shown by performing blood perfusion experiments in the presence of thrombin inhibitors. Our results indicate that 1) platelet adhesion and aggregate formation on a subendothelium may occur at a high shear rate (2600/s) without the participation of collagen types I, III and VI, and vWF; and 2) the HMEC-1 cell line may prove useful for in vitro studies of the thrombogenic properties of microvascular subendothelium which in most cases does not contain fibrillar collagens and vWF.

Antithrombotic effect of platelet glycoprotein Ib-blocking monoclonal antibody Fab fragments in nonhuman primates.

Platelet adhesion in arterial blood flow is mainly supported by the platelet receptor glycoprotein (GP) Ib, which interacts with von Willebrand factor (vWF) that is bound to collagen at the site of vessel wall injury. Antibody 6B4 is a monoclonal antibody (MoAb) raised against purified human GPIb. MoAb 6B4 inhibits both ristocetin- and botrocetin-induced, vWF-dependent human platelet agglutination. MoAb 6B4 furthermore blocks shear-induced adhesion of human platelets to collagen I. We studied the antithrombotic effect of this inhibitory murine MoAb 6B4 in a baboon model of arterial thrombosis. When injected into baboons, intact IgG and its F(ab')(2) fragments caused almost immediate thrombocytopenia, whereas injection of the Fab fragments alone did not. Fab fragments were subsequently used to investigate their in vivo effect on platelet deposition onto a thrombogenic device, consisting of collagen-rich, glutaraldehyde-fixed bovine pericardium (0.6 cm(2)), at a wall shear rate ranging from 700 to 1000 s(-1). Baboons were either pretreated with Fabs to study the effect of inhibition on platelet adhesion or treated 6 minutes after placement of the thrombogenic device to investigate the effect on interplatelet cohesion. Pretreatment of the animals with bolus doses ranging from 80 to 640 microgram/kg Fab fragments significantly reduced (111)In-labeled platelet deposition onto the collagen surface by approximately 43% to 65%. Only the highest dose caused a significant prolongation (doubling) of the bleeding time. Ex vivo ristocetin-induced platelet agglutination was equally reduced. Treatment with a bolus of 110 microgram/kg Fab fragments after a thrombus was allowed to form for 6 minutes had no effect on further platelet deposition. We therefore conclude that Fab fragments or derivatives of inhibitory anti-GPIb antibodies may be useful compounds to prevent thrombosis.

Acquired Bernard-Soulier syndrome: a case with necrotizing vasculitis and thrombosis.

We describe a patient with positive antinuclear antibodies, polyclonal gammopathy and high level of circulating immunocomplexes, resulting in vascular purpura. In addition, the patient had a slightly prolonged bleeding time and an isolated defect of ristocetin-induced platelet aggregation (RIPA) in platelet-rich plasma (PRP). The patient's plasma also inhibited RIPA in normal PRP and in normal platelet suspension. The activity and multimeric structure of plasmatic von Willebrand factor showed no alteration. We could demonstrate an autoantibody against platelet membrane glycoprotein (GP) Ib, using an ELISA-type assay. These data suggest an acquired Bernard-Soulier syndrome. We suggest that the patient had an immunocomplex-mediated leukocytoclastic vasculitis accompanied by production of antinuclear autoantibodies as well as the presence of an autoantibody against GPIb. The titer of the anti-GPIb antibody, however, was too low to induce significant platelet-type bleeding tendency, only laboratory alterations were found. Moreover, in a later stage of her disease, she developed a severe necrotizing vasculitis which was followed by a deep venous thrombosis.

Increased platelet activation and decreased fibrinolysis in the pathogenesis of aseptic necrosis of the femoral head.

Thrombotic events, increased tendency toward intravascular thrombosis and decreased fibrinolysis seem to be possible pathologic causes for aseptic necrosis of the femoral head (ANFH) in adults. This project was to study whether either increased platelet activation or decreased fibrinolytic activity and/or any other thrombogenic factor may be implicated in the evolvement of ANFH. The speed of the in vitro lysis was significantly lower in patients (both in primary and in secondary cases) compared with healthy controls. The platelet activation (measuring with beta TG) proved to be significantly higher in the primary group as well as in the secondary group compared with healthy controls. Lp(a) levels were elevated in primary and secondary cases. This alteration was more characteristic in the primary cases. Fibrinogen levels were also elevated in the primary group, but the difference was not significant. The data shown here may further support that hypofibrinolysis and increased thrombogenesis are major causes of ANFH. Early diagnosis of ANFH increases the chances of modifying the course of this disabling disease.

Effect of L-arginine on in vitro plasmin-generation and fibrinogenolysis.

L-arginine ahs received much attention in numerous aspects of the regulation of vascular tone and haemostasis. L-arginine seems to be capable to bind to plasminogen, too. The aim of the present paper is to investigate the action of L-arginine on in vitro plasmin generation and fibrino(geno)lysis by chromogenic, kinetic plasmin generation assay and electrophoretic analysis. The acceleration of tPA-induced plasmin generation in the presence of low concentration of L-arginine, along with augmentation of in vitro fibrinogenolysis have been documented. L-arginine may have a role in the modification of fibrinogenolysis, and this role should be considered if arginine is used as an element of some novel antithrombotic agents.

Recirculated normal platelets adhere to surfaces coated with plasma from patients with immune thrombocytopenia.

Immune thrombocytopenic purpura (ITP) patients have characteristic anti-platelet antibodies in their circulation. To assess the interaction between such antibodies adhering on to a non-physiological surface and human platelets, normal anticoagulated blood was perfused over ITP patient plasma-coated surfaces in a parallel plate flow chamber. At 300 s-1, platelet adhesion to patient plasma-coated glass coverslips (24.0 +/- 10%) was significantly higher than the adhesion to normal plasma-coated surfaces (9.8 +/- 7%). When perfused at 1300 s-1, the adhesion to patient plasma-(5.1 +/- 1.3%) and to normal plasma-(2.5 +/- 1.2%) coated coverslips were significantly weaker. Furthermore, patient platelet binding depended on simultaneous contributions by antibodies and fibrinogen present on the plasma-coated surface, since adherence was antagonized both by normal immunoglobulins added to the perfusate, as well as by the anti-GPIIb/IIIa monoclonal antibody 16N7C2, which competes with fibrinogen for binding to its receptor on the platelet. Accordingly, platelet adhesion was only observed to coverslips coated with the plasma but not the serum of ITP patients. Hence, perfusion of normal platelets over surfaces coated with ITP patient plasma enables a functional assessment of the presence in this plasma of anti-platelet antibodies.

New pentamidine related substances which simultaneously inhibit platelet aggregation and accelerate plasmin generation and in vitro clot lysis.

Pentamidine, a highly toxic drug, possesses RGD-peptide (Arg-Gly-Asp)-like antiplatelet actions. The objective of this investigation was to study the anticipated profibrinolytic and antiplatelet actions of pentamidine and of pentamidine (bearing guanidino-like groups)-related synthetic peptidomimetic compounds. Platelet aggregation inhibition was assessed using ADP, thrombin, collagen, arachidonic acid and epinephrine as inducers, by aggregometry. In vitro chromogenic plasmin generation tests and clot lysis assays were also performed. Two (assigned as D-2 and D-3) of the synthetic pentamidine-guanidino related molecules were able to inhibit platelet aggregation and simultaneously accelerate in vitro plasmin generation and clot-lysis in the nM range. These dual action antithrombotic agents now need to be tested further to assess their antithrombotic actions in vivo.

Altered lysis resistance of platelet-rich clots in patients with insulin-dependent diabetes mellitus.

The fibrinolytic resistance of platelet-rich arterial thrombi received much attention. Clot lysis method was used to assess the in vitro fibrinolytic properties in diabetes mellitus. Platelet rich (PRP) clots were formed by addition of thrombin, and lysis was induced by tissue-plasminogen-activator. The coagulation and lysis was followed by the light scattering properties. A special pattern of good initial lysis followed by a second clotting phase was observed in more than half of insulin dependent diabetic patients, while a similar pattern of clot-lysis was only occasionally found in non-insulin dependent diabetes mellitus or in the healthy control group. Following the thrombin activation of washed, gel-filtered platelets, the supernatants possessed an inhibitory action on in vitro lysis of PPP-clots. This suppression was remarkably stronger in IDDM, along with the highest PAI-1 activity concentration ratio of the platelet lysates, compared to plasmatic levels. The relation of this special type of PRP clot-lysis resistance to diabetic vascular complications needs further clarifying and investigations.

Calin from Hirudo medicinalis, an inhibitor of von Willebrand factor binding to collagen under static and flow conditions.

Calin from the saliva of the medicinal leech, Hirudo medicinalis, is a potent inhibitor of collagen mediated platelet adhesion and activation. In addition to inhibition of the direct platelet-collagen interaction, we presently demonstrate that binding of von Willebrand to coated collagen can be prevented by Calin, both under static and flow conditions in agreement with the occurrence of binding of Calin to collagen, confirmed by Biospecific Interaction Analysis. To define whether Calin acted by inhibiting the platelet-collagen or the platelet-von Willebrand factor (vWF)-collagen-mediated thrombus formation, platelet adhesion to different types of collagens was studied in a parallel-plate flow chamber perfused with whole blood at different shear rates. Calin dose-dependently prevented platelet adhesion to the different collagens tested both at high- and low-shear stress. The concentration of Calin needed to cause 50% inhibition of platelet adhesion at high-shear stress was some fivefold lower than that needed for inhibition of vWF-binding under similar conditions, implying that at high-shear stress, the effect of Calin on the direct platelet-collagen interactions, suffices to prevent thrombus formation. Platelet adhesion to extracellular matrix (ECM) of cultured human umbilical vein endothelial cells was only partially prevented by Calin, and even less so at a high-shear rather than a low-shear rate, whereas the platelet binding to coated vWF and fibrinogen were minimally affected at both shear rates. Thus, Calin interferes with both the direct platelet-collagen interaction and the vWF-collagen binding. Both effects may contribute to the inhibition of platelet adhesion in flowing conditions, although the former seems to predominate.

[Changes in fat metabolism in acute carbon tetrachloride intoxication of rats]. / Zsíranyagcsere változások patkányok heveny széntetraklorid mérgezésében.

CCl4 is an organic solvent and a well known hepatotoxic agent. Injury is mediated by reactive free radicals, mainly-CCl3 (trichloromethyl). Liver lesion develops within one-two hours, however, late toxic effects may appear after a delay of several hours or two to three days. Some drugs and liver toxicants cause disturbances in synthesis and metabolism of triglycerides, cholesterol and lipoproteins, thus damaging the basic resource for living cells. In our experiments a single 1.25 ml/kg CCl4 dose was administered s.c. to male Wistar rats. 24 hours later triglyceride level increased in the plasma by 223%. Cholesterol content suffered no changes. HDL-C level decreased by 40%. LDL-C concentration was higher by 235%. Cholesterol/HDL-C ratio increased by 0.75% on account of the practically unchanged cholesterol amount in the blood. The most often used calculation (Friedewald equation) LDL-C/HDL-C ratio was higher by 305%. The increased triglyceride content in the blood is in correlation with the fatty degeneration of the liver. The high LDL-C/HDL-C ratio may point to incipient atherosclerotic complications. The pathological lipid levels measured 24 hours after intoxication claim for delayed toxic effects to be taken into consideration. It may be suggested to determine the main lipid parameters after carbon tetrachloride exposition.