[Cutaneous lesions in COVID-19 patients]. / Huidafwijkingen bij covid-19.

Aside from the typical respiratory symptoms resulting from an infection with SARS-CoV-2, there are reports of cutaneous lesions in patients diagnosed with a SARS-CoV-2 infection. There are reports of multiple groups of skin lesions presenting in different stages of this diagnosis. The most common reported groups are chilblains, vesicular eruptions, morbilliformexanthems, acute urticaria and livedo. It is unlikely that all these groups of skin lesions are distinctive of an infection with SARS-CoV-2. Chilblains of new onset, however, could possibly be a distinctive symptom of a mild/asymptomatic infection with SARS-CoV-2. It is recommended to consider an infection with SARS-CoV-2 in the differential diagnosis in patients presenting with these groups of skin lesions. Consider testing for SARS-CoV-2 and consult the dermatologist if needed, especially in case of chilblains, to ensure histopathological evaluation of the skin lesions to increase knowledge of the underlying pathophysiology.

Amount of Brain Edema Correlates With Neurologic Recovery in Pediatric Cerebral Malaria.

BACKGROUND: Cerebral malaria (CM) remains a leading cause of mortality and morbidity in children in sub-Saharan Africa. Recent studies using brain magnetic resonance imaging have revealed increased brain volume as a major predictor of death. Similar morphometric predictors of morbidity at discharge are lacking. The aim of this study was to investigate the utility of serial cranial cisternal cerebrospinal fluid (CSF) volume measurements in predicting morbidity at discharge in pediatric CM survivors. METHODS: In this case-control study, 54 Malawian pediatric CM survivors with neurologic sequelae evident at discharge who underwent serial magnetic resonance imaging scans while comatose were matched to concurrently admitted children with serial imaging who made full recoveries. Serial cranial cisternal CSF volume quantified by radiologists blinded to outcome was evaluated as a predictor of neurologic deficits at discharge. The probability of neurologic sequelae was determined using a model that included coma duration and changes in cisternal CSF volume over time. RESULTS: Coma duration before admission was similar between cases and controls (16.1 vs. 15.3; P = 0.81), but overall coma was longer among children with sequelae (60 vs. 38 hours; P < 0.01). Lower initial CSF volumes and decreased volumes over time were both associated with a higher probability of neurologic sequelae at discharge. CONCLUSIONS: Among pediatric CM survivors with prolonged coma, lower initial CSF volume and decreasing volume during coma is associated with neurologic sequelae at discharge. These findings suggest that cerebral edema is an underlying contributor to both morbidity and mortality in pediatric CM.

PP107. Cardiovascular parameters 40 years after hypertensive pregnancies.

INTRODUCTION: Epidemiological data indicate an increased cardiovascular risk in women with previous hypertensive pregnancies. There are few clinical investigations regarding the mechanisms that could mediate this increased risk. OBJECTIVES: The aim of the present study was to clarify if any deterioration in the cardiovascular, metabolic or neuroendocrine status is present in women 40 years after pregnancies complicated by hypertension. METHODS: Three hundred and nineteen women were invited to take part in a follow up investigation regarding cardiovascular regulation. One hundred and five women accepted to participate - 50 with previously hypertensive pregnancies (HTP) and 55 with normotensive pregnancies (NTP). Office and ambulatory blood pressure levels, central blood pressure and pulse wave velocity, echocardiographic measurements (RWT, LVMI, LA, LA-RA, diastolic function, strain) and P-glucose, HbA1c, S-leptin, S-hsCRP, P-renin, P-Noradrenaline and NT-proBNP were examined. Women who choose not to participate (n=214) were followed up with a questionnaire regarding their previous pregnancies and present cardiovascular health. RESULTS: The investigations did not reveal differences in any examined variables regarding blood pressure, echocardiographic parameters or blood analysis for metabolic and neurohumoral balance. Twenty-five individuals were diagnosed with hypertension in the HTP group (mean BP 145/86mmHg) and 17 subjects in the NTP group (mean BP 145/87mmHg). The questionnaire was answered by 79% of the participants and revealed that these women had an impaired cardiovascular health compared to the group investigated. CONCLUSION: Blood pressure, metabolic and neuroendocrine parameters are not permanently worsened in all women with previous hypertensive pregnancies. There exist disparities within the group of women with previous hypertensive pregnancies and there are women without obvious cardiovascular or metabolic dysfunction 40 years after the hypertensive manifestation during pregnancy.

Swedish hypertension open care retrospective study in men and women (SHOW).

The aim of this retrospective study in primary health care was to study gender differences in blood pressure levels in response to treatment of new onset hypertension. Gender difference in blood pressure control and pharmacological treatment was also recorded. A total of 334 women and 332 men aged ≥50 years and <80 years at baseline, with blood pressure ≥140 mm Hg systolic and/or ≥90 mm Hg were included. Men were younger, had a higher frequency of type II diabetes mellitus and a higher body mass index compared with women at baseline. There was no difference between women and men in systolic blood pressure (SBP) before or after treatment. Women however had a lower diastolic blood pressure (DBP) before and after intervention and as a result a higher pulse pressure (PP). Approximately 50% of the patients reached target blood pressure (≤140/90 mm Hg) in both women and men. Beta blocker was the most commonly used antihypertensive treatment in both genders, whereas diuretics were predominately used in women. In conclusion; women and men reached target blood pressure to the same extent but with different antihypertensive treatment strategies. Differences at baseline in risk factor pattern may explain the finding.

Aerobic exercise program reduces anger expression among overweight children.

This study tested the effect of a structured aerobic exercise program on anger expression in healthy overweight children. Overweight sedentary children were randomly assigned to an aerobic exercise program or a no-exercise control condition. All children completed the Pediatric Anger Expression Scale at baseline and posttest. Anger Out and Anger Expression scores were lower for the exercise condition at posttest. Fitness improvements contributed significantly to final models, and points earned for adherence correlated negatively with posttest Anger Out. An aerobic exercise program might be an effective strategy to reduce anger expression, including reduction of aggressive behavior, in overweight children.

Classification and management of skin, hair, nail and mucosal side-effects of epidermal growth factor receptor (EGFR) inhibitors.

Inhibitors of epidermal growth factor receptor (EGFR) are commonly used as therapeutic agents in oncology. In contrast to currently used oncological treatments, these inhibitors almost always cause skin- and skin adnex toxicity. About 85% of treated patients develop to a more or lesser extent an acneiform eruption. Xerosis cutis and painful nail disorders occur in, respectively, 35% and 10-15% of all treated patients. Also hair and mucosal changes have been reported, although to a lesser extent. These skin- and skin adnex toxicities are reversible after withdrawal of treatment, but are seldom a reason to stop or interrupt therapy. This review outlines the classification, the pathogenesis and therapy of these skin, hair, nail and mucosal changes due to EGFR inhibition. Informing the patient and management of these side-effects is very important to reduce discomfort and as such to increase compliance to therapy.

[Long-term beneficial effects of cetuximab in a woman with metastasised rectal carcinoma without expression of the epidermal growth factor receptor]. / Langdurig gunstig klinisch effect van cetuximab bij een patiënte met een gemetastaseerd rectumcarcinoom zonder expressie van epidermale-groeifactorreceptor.

A 37-year-old woman presented with an epidermal growth factor-(EGFR)-negative rectum carcinoma with liver metastases. After extensive treatment, consisting of first-line chemotherapy, low anterior resection, isolated liver perfusion, second- and third-line chemotherapy and a pericardiodesis with bleomycin, she was subsequently treated with combination irinotecan and cetuximab therapy. At her last follow-up she had had long-term stable disease for 18 months with clinical benefit. Cetuximab is a monoclonal antibody which targets EGFRR. This exceptional case illustrates that treatment with cetuximab may be of benefit to a patient with EGFR-negative colorectal cancer. The exact mechanism of action and the role ofcetuximab in the treatment of advanced colorectal cancer have still to be determined.

Evaluating personal health care and health promotion web sites.

OBJECTIVE: An exemplary sample of web sites relevant to personal health care and health promotion was chosen and evaluated. METHODS: Both quantitative and qualitative data were converged to assess and rank the sites on nine attributes. RESULTS: The sites provided a definitive range of value and variety of presentations, health care and health promotion information, and services covering the virtual choices currently available to users of the Internet. CONCLUSION: Discussion focused on methodological approaches and issues of web site evaluation serving the public interest, health care, and health promotion.

Role of fibrin matrix in angiogenesis.

Angiogenesis, the formation of new blood vessels from existing vessels, plays an important role during development. In the adult, it is limited to the female reproductive system and to tissue repair and pathological conditions. Repair associated angiogenesis is usually accompanied by the presence of inflammatory cells, vascular leakage, and fibrin deposition. The temporary fibrin matrix acts, not only as a sealing matrix, but also as a scaffolding for invading leukocytes and endothelial cells during tissue repair. We have used a three-dimensional fibrin matrix to study the outgrowth of human microvascular endothelial cells in capillary-like tubular structures. This process is induced by the simultaneous addition of an angiogenic growth factor (bFGF or VEGF) and the cytokine TNF alpha, and is enhanced by hypoxia. It involves proteolytic activities, in particular cell bound urokinase/plasmin and matrix metalloproteinase activities. Modulation of the fibrin structure markedly affects the extent and stability of capillary tube formation in vitro. Preparation of fibrin at different pH (7.0-7.8) or crosslinking of the fibrin matrix induces differences in fibrin matrix rigidity and structure. This is accompanied by a change in capillary ingrowth. Heparins, in particular low molecular weight heparins, modulate the fibrin structure and by this action affect angiogenesis in vitro. A mutant fibrinogenNieuwegein, which lacks the terminal part of the A alpha chain of fibrin harboring an RGD sequence and the transglutaminase sequence, provided additional evidence that the structure of fibrin is an important determinant for angiogenesis. These findings may have impact on improving wound healing and on influencing angiogenesis in malignancies with a fibrinous stroma.

Genetically engineered peptide fusions for improved protein partitioning in aqueous two-phase systems. Effect of fusion localization on endoglucanase I of Trichoderma reesei.

Genetic engineering has been used for fusion of the peptide tag, Trp-Pro-Trp-Pro, on a protein to study the effect on partitioning in aqueous two-phase systems. As target protein for the fusions the cellulase, endoglucanase I (endo-1,4-beta-Dglucan-4-glucanohydrolase, EC 3.2.1.4, EGI, Cel7B) of Trichoderma reesei was used. For the first time a glycosylated two-domain enzyme has been utilized for addition of peptide tags to change partitioning in aqueous two-phase systems. The aim was to find an optimal fusion localization for EGI. The peptide was (1) attached to the C-terminus end of the cellulose binding domain (CBD), (2) inserted in the glycosylated linker region, (3) added after a truncated form of EGI lacking the CBD and a small part of the linker. The different constructs were expressed in the filamentous fungus T. reesei under the gpdA promoter from Aspergillus nidulans. The expression levels were between 60 and 100 mg/l. The partitioning behavior of the fusion proteins was studied in an aqueous two-phase model system composed of the thermoseparating ethylene oxide (EO)-propylene oxide (PO) random copolymer EO-PO (50:50) (EO50PO50) and dextran. The Trp-Pro-Trp-Pro tag was found to direct the fusion protein to the top EO50PO50 phase. The partition coefficient of a fusion protein can be predicted with an empirical correlation based on independent contributions from partitioning of unmodified protein and peptide tag in this model system. The fusion position at the end of the CBD, with the spacer Pro-Gly, was shown to be optimal with respect to partitioning and tag efficiency factor (TEF) was 0.87, where a fully exposed tag would have a TEF of 1.0. Hence, this position can further be utilized for fusion with longer tags. For the other constructs the TEF was only 0.43 and 0.10, for the tag fused to the truncated EGI and in the linker region of the full length EGI, respectively.

Genetic engineering of the Trichoderma reesei endoglucanase I (Cel7B) for enhanced partitioning in aqueous two-phase systems containing thermoseparating ethylene oxide--propylene oxide copolymers.

Endoglucanases (endo-1,4-beta-D-glucan-4-glucanohydrolase, EC 3.2.1.4) are industrially important enzymes. In this study endoglucanase I (EGI or Cel7B) of the filamentous fungi Trichoderma reesei has been genetically engineered to investigate the influence of tryptophan rich peptide extensions (tags) on partitioning in an aqueous two-phase model system. EGI is a two-domain enzyme and is composed of a N-terminal catalytic domain and a C-terminal cellulose binding domain, separated by a linker. The aim was to find an optimal tag and fusion position, which further could be utilised for large scale extractions. Peptide tags of different length and composition were attached at various localisations of EGI. The fusion proteins were expressed from T. reesei with the use of the gpdA promoter from Aspergillus nidulans. Variations in secreted levels between the engineered proteins were obtained. The partitioning of EGI in an aqueous two-phase system composed of a thermoseparating ethylene oxide-propylene oxide random copolymer (EO(50)PO(50)) and dextran, could be significantly improved by relatively minor genetic engineering. The (Trp-Pro)(4) tag added after a short stretch of the linker, containing five proline residues, gave in the highest partition coefficient of 12.8. The yield in the top phase was 94%. The specific activity was 83% of the specific activity of unmodified EGI on soluble substrate. The efficiency of a tag fused to a protein is shown by the tag efficiency factor (TEF). A hypothetical TEF of 1.0 would indicate full tag exposure and optimal contribution to the protein partitioning by the fused tag. The location of the fusion point after the sequence of five proline residues in the linker of EGI is the most beneficial in two-phase separation. The highest TEF (0.97) was obtained with the (Trp-Pro)(2) tag at this position, indicating full exposure and intactness of the tag. However, the peptide tag composed of (Trp-Pro)(4) improved the partition properties the most but had lower TEF in comparison to (Trp-Pro)(2).

Aberrant fibrin formation and cross-linking of fibrinogen Nieuwegein, a variant with a shortened Aalpha-chain, alters endothelial capillary tube formation.

A congenital dysfibrinogenemia, fibrinogen(Nieuwegein), was discovered in a young man without any thromboembolic complications or bleeding. A homozygous insertion of a single nucleotide (C) in codon Aalpha 453 (Pro) introduced a stop codon at position 454, which resulted in the deletion of the carboxyl-terminal segment Aalpha 454-610. The ensuing unpaired cysteine at Aalpha 442 generated fibrinogen-albumin complexes of different molecular weights. The molecular abnormalities of fibrinogen(Nieuwegein) led to a delayed clotting and a fibrin network with a low turbidity. Electron microscopy confirmed that thin fibrin bundles were organized in a fine network. The use of fibrinogen(Nieuwegein)-derived fibrin (fibrin(Nieuwegein)) in an in vitro angiogenesis model resulted in a strong reduction of tube formation. The ingrowth of human microvascular endothelial cells (hMVEC) was independent of alpha(v)beta(3), indicating that the reduced ingrowth is not due to the absence of the RGD-adhesion site at position Aalpha 572-574. Rather, the altered structure of fibrin(Nieuwegein) is the cause, since partial normalization of the fibrin network by lowering the pH during polymerization resulted in an increased tube formation. Whereas factor XIIIa further decreased the ingrowth of hMVEC in fibrin(Nieuwegein), tissue transglutaminase (TG), which is released in areas of vessel injury, did not. This is in line with the absence of the cross-linking site for TG in the alpha-chains of fibrinogen(Nieuwegein). In conclusion, this newly discovered congenital dysfibrinogenemia has a delayed clotting time and leads to the formation of an altered fibrin structure, which could not be cross-linked by TG and which is less supportive for ingrowth of endothelial cells.

Parameters influencing protein extraction for whole broths in detergent based aqueous two-phase systems.

The parameters important for an optimisation of cloud point extraction in technical scale were investigated using a genetically engineered fusion protein derived from endoglucanase I expressed in Trichoderma reesei and the nonionic polyoxyethylene Agrimul NRE 1205. The key parameters are temperature, detergent concentration, and additional salts. These parameters are interdependent, thus there is an optimum in the partition coefficient with respect to detergent concentration and a maximum for the partition coefficient and the yield with respect to temperature. These results were confirmed for the detergent C12E5 to demonstrate that these optima are due to the nature of polyoxyethylenes. Cloud point extraction was found to be only slightly affected by pH. In the case studied extraction of whole broth is favourable for a high yield and partition coefficient, since fusion protein adhering to the cells can be solubilized. However some loss of detergent which remains in the fungal biomass was observed.

Unfractionated and low molecular weight heparin affect fibrin structure and angiogenesis in vitro.

Cancer patients treated for venous thromboembolism with low molecular weight heparin (LMWH) have a better survival rate than patients treated with unfractionated heparin (UFH). Because fibrin-associated angiogenesis is an important determinant in the progression and metastasis of many solid tumors, the effects of heparins on in vitro angiogenesis were investigated. Both UFH and LMWH inhibited bFGF-induced proliferation of human microvascular endothelial cells (hMVECs) to the same the extent (36-60%). VEGF165-induced proliferation was inhibited to a to a lesser extent (19-33%). Turbidity measurements and electron microscopy showed that the presence of LMWH during polymerization of the fibrin matrix led to a more transparent rigid network with thin fibrin bundles, whereas the presence of UFH resulted in a more opaque more porous network with thick fibrin fibers. We used a human in vitro angiogenesis model, which consisted of hMVECs seeded on top of a fibrin matrix, and stimulated the cells with basic fibroblast growth factor plus tumor necrosis factor a to induce capillary-like tubular structures. The formation of capillary-like tubular structures was retarded with matrices polymerized in the presence of LMWH (46% inhibition compared with a control matrix for both 1.5 and 10 units/ml LMWH), whereas matrices polymerized in the presence of UFH facilitated tubular structure formation (72 and 36% stimulation compared with a control matrix for 1.5 and 10 units/ml UFH, respectively). Similar results were obtained for cells stimulated with vascular endothelial growth factor plus tumor necrosis factor alpha. These data demonstrate the inhibitory effect of heparins on proliferation of hMVECs and provide a novel mechanism by which LMWH may affect tumor progression, namely reduced ingrowth of microvascular structures in a fibrinous stroma matrix by rendering it less permissive for invasion.

The inactivation of single-chain urokinase-type plasminogen activator by thrombin on cultured human endothelial cells.

Single-chain urokinase-type plasminogen activator (scu-PA) is cleaved by thrombin, resulting in an inactive molecule called thrombin-cleaved two-chain urokinase-type plasminogen activator (tcu-PA/T). There is no knowledge about cell-mediated inactivation of scu-PA. We have studied whether scu-PA bound to cultured human umbilical vein endothelial cells (HUVEC) could be inactivated by thrombin. High molecular weight scu-PA was bound to HUVEC and incubated with increasing amounts of thrombin for 30 min at 37 degrees C. Cell-bound urokinase-type plasminogen activator (u-PA) was released and levels of scu-PA, tcu-PA/T and active two-chain u-PA were measured using sensitive bioimmunoassays. Cell-bound scu-PA was efficiently inactivated by thrombin. Fifty percent inactivation of scu-PA occurred at about 0.2 nM thrombin. In the presence of monoclonal anti-urokinase receptor IgG, at least 50% of the binding of scu-PA to HUVEC was inhibited. The relative amount of tcu-PA/T that was generated by thrombin was not affected by the monoclonal antibody. These results indicated that scu-PA bound to HUVEC via the urokinase receptor can be inactivated by thrombin. The efficient inactivation of cell-bound scu-PA suggests that a cofactor for thrombin may be involved, like thrombomodulin or glycosaminoglycans. It is concluded that scu-PA bound to the urokinase receptor on a cell surface can be inactivated by thrombin, which may have profound effects on u-PA-mediated local fibrinolysis and extracellular proteolysis during processes in which thrombin is also involved.

Angiogenesis and anti-angiogenesis: perspectives for the treatment of solid tumors.

Angiogenesis is the formation of new blood vessels from preexisting ones. Many solid tumors depend on an extensive newly formed vascular network to become nourished and to expand. Tumor cells induce the formation of an extensive but aberrant vascular network by the secretion of angiogenic factors. A proper context is needed for the endothelial cells to respond. To create this proper context, the tumor often uses the body's own repair system to accelerate angiogenesis and the subsequent tumor expansion. The angiogenic response is governed by the interaction of angiogenic growth factors and cytokines with specific receptors on the endothelium, as well as by the interaction of these cells with their surrounding matrix, which is regulated by matrix-degrading proteases and adhesion molecules such as integrins. A number of agents have been discovered and developed that aim to inhibit angiogenesis and to convert the tumor to a dormant state. They have proven to be effective in animal studies. At present their efficacy in man is under evaluation.

Role and localization of urokinase receptor in the formation of new microvascular structures in fibrin matrices.

Fibrin or a fibrinous exudate can facilitate angiogenesis in many pathological conditions. In vitro, the outgrowth of capillary-like structures in fibrin can be mimicked by exposing human microvascular endothelial cells (hMVECs) to an angiogenic growth factor and tumor necrosis factor (TNF)-alpha. Urokinase-type plasminogen activator (u-PA) and plasmin activities are required for this angiogenic process. This study focuses on the role and localization of the u-PA receptor (u-PAR) in newly formed microvascular structures. The u-PAR-blocking monoclonal antibody (MAb) H-2 completely inhibited the formation of capillary-like tubular structures induced by exposure of hMVECs to basic fibroblast growth factor and TNF-alpha. This was accompanied by a several-fold increase in u-PA accumulation in the conditioned medium. The effect of MAb H-2 was not caused by blocking cellular activation by u-PA/u-PAR interaction, as the amino-terminal fragment (ATF) of u-PA, which also activates u-PAR, prevented tube formation. In addition, the inhibition by MAb H-2 was not due to an effect of the antibody on u-PAR-vitronectin binding. These data show that inhibition of tube formation can be caused not only by inhibition of u-PA or plasmin activities but also by unavailability of the u-PAR for cell-bound proteolysis. Immunohistochemical analysis showed that in in vitro angiogenesis u-PAR and u-PA were localized on the invading, tube-forming hMVECs and not on the endothelial cells that are located on top of the fibrin matrix. u-PAR and u-PA were also prominently expressed on endothelial cells of neovessels present in an atherosclerotic plaque. These data may give more insight into the role of u-PAR in repair-associated angiogenesis.

Comparison of different methods for plant regeneration and transformation of the legume Galega orientalis Lam. (goat's rue).

For the first time, regeneration and transformation have been achieved from the legume Galega orientalis Lam. (goat's rue). Two different regeneration protocols are described, one based on direct shoot induction from meristems and the other involving callus induction and shoot induction from callus with the plant growth regulator thidiazuron (TDZ). Different media and explants were evaluated. Three different transformation methods were compared: cocultivation with four different Agrobacterium tumefaciens strains, electroporation of embryos and apical meristems and particle bombardment of embryos. TDZ-promoted shoot induction on calli from immature embryos gave the best results. Transformation using this regeneration protocol was most successful with particle bombardment. Stable transformation has yet to be proven.

Influence of fibrin structure on the formation and maintenance of capillary-like tubules by human microvascular endothelial cells.

Fibrin is a temporary matrix which not only covers a wound, but also provides a structure for invading cells during healing. Changes in the polymerization conditions before gelation of the clot affect the structure of fibrin and thus might influence the interaction with invading cells. Therefore we tested whether changes in the fibrin structure influence the formation of capillary-like tubular structures by human microvascular endothelial cells (hMVEC) in an in vitro angiogenesis model. Opaque [125I]fibrin structures prepared at pH 7.0, fibrin matrices at pH 7.4 and transparent [125I]fibrin structures prepared at pH 7.8 were neutralized (pH 7.4) before seeding hMVEC on top of them in confluent density. Endothelial cells were stimulated with a growth factor [basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF)165] and a cytokine [tumor necrosis factor (TNF)-alpha] to induce the u-PA/u-PA receptor-dependent formation of capillary-like tubular structures. The formation of these structures was quantified by determining the length of the invasive structures by image analysis and by measuring the accompanying [125I]fibrin degradation. Ingrowth of tubular structures proceeded at a faster rate in opaque matrices consisting of thick fibrin fibers as compared to transparent gels with fine fibrin fibers. The more rapid ingrowth of tubular structures in opaque fibrin gels induced by bFGF/TNF-alpha or VEGF165/TNF-alpha was accompanied by a larger extent of fibrin degradation. Both processes were inhibited by aprotinin and epsilon-aminocaproic acid indicating the involvement of plasmin. They were also inhibited by anti-u-PA or anti-u-PA receptor IgG, but not by anti-t-PA IgG, suggesting the involvement of cell-bound u-PA activity. However, in the opaque fibrin gels, the tubular structures dissolved upon prolonged incubation due to excessive fibrin degradation. Simulation of hMVEC with bFGF alone did not induce tubular structures, but ca used a high degree of t-PA- and plasmin-dependent fibrin lysis, and, after several days, a partial detachment of sheets of cells. Gradual inhibition of the excessive fibrin degradation by a series of aprotinin concentrations did not lead to tube formation in bFGF-treated cells. These data indicate that the formation and stability of tubular structures by hMVEC in fibrin is accompanied by controlled fibrinolysis and depends critically not only on cell-bound u-PA-dependent plasminogen activation, but also on the fibrin structure. Because the fibrin structure is largely influenced by the conditions in which fibrin has been polymerized, these conditions may have considerable impact on angiogenesis during wound healing and vascularization of tumour stroma.