Plasma C1-inhibitor Research Articles

Hereditary angioneurotic oedema: characterization of plasma kinin and vascular permeability-enhancing activities.

The mediator(s) responsible for localized enhanced vascular permeability that characterizes an exacerbation of hereditary angioneurotic oedema (HAE) is thought to be a product of either contact or complement system activation. In contrast to normal individuals, plasma from these patients generates both kinin and vascular permeability-enhancing activity following incubation at 37 degrees C. Depletion of C1 inhibitor in both normal and C2-deficient plasma, but not in contact factor-deficient plasmas, resulted in generation of these activities. The kinin activity from incubated HAE plasma was susceptible to kininase inactivation and was blocked by a Bk2 receptor antagonist. Furthermore, this activity was isolated from HAE plasma; amino acid sequence analysis proved it to be bradykinin. Similarly, the vasopermeability-enhancing activity from ethanol-fractionated or boiled HAE plasma, collected during either attack or remission, co-eluted with bradykinin on reverse-phase high performance liquid chromatography (HPLC). These studies conclusively demonstrate that bradykinin is the major kinin and mediator of enhanced vascular permeability generated during incubation of HAE plasma. The role of other bioactive products, such as the C2 kinin, at local sites of oedema formation remains to be further defined.

C1-Esterase Inhibitor Protects Against Neointima Formation After Arterial Injury in Atherosclerosis-Prone Mice

Although activation of the complement system has been implicated in the progression of human atherosclerosis, its function during arterial remodeling after injury has not been investigated. Here, we examined the contribution of the complement cascade to neointima formation in apolipoprotein E-deficient mice using a C1-esterase inhibitor (C1-inhibitor). Apolipoprotein E-deficient mice fed an atherogenic diet were subjected to wire-induced endothelial denudation of the carotid artery and treated with C1-inhibitor (Berinert; 15 IU i.v.) or vehicle perioperatively and subsequently every 2 days. The effectiveness of C1-inhibitor treatment was confirmed by measurement of plasma C1-inhibitor activity. A significant reduction in serum triglyceride levels was observed in C1-inhibitor-treated mice, whereas cholesterol levels did not differ. After 3 weeks, neointimal area was significantly reduced in C1-inhibitor-treated mice versus controls, whereas medial area was unaltered. This was associated with a significant decrease in neointimal and medial macrophage and CD3+ T-cell content. Expression of C3 mRNA was significantly reduced in plaques of C1-inhibitor-treated mice 10 days after injury, as assessed by reverse-transcription polymerase chain reaction. The peak in serum C3 levels after injury was markedly downregulated by C1-inhibitor, as evidenced by ELISA. Immunohistochemistry revealed strong expression of C3 and C3c, which colocalized to plaque macrophages and was reduced in C1-inhibitor-treated mice. C1-inhibitor impaired monocyte arrest on activated endothelium and platelets under flow conditions in vitro and leukocyte recruitment to carotid arteries 1 day after injury in vivo. C1-inhibitor limits neointimal plaque formation and inflammation. This may involve blockade of complement activation, inhibition of leukocyte recruitment, and reduced triglyceride levels, thus providing a multimodal approach to treat arterial disease.

C1-inhibitor deficiencies (hereditary angioedema): Where are we with therapies?

Hereditary angioedema, an autosomal dominant disorder, presents clinically as recurrent episodes of swelling. It results from either deficient production or function of C1 inhibitor. Acquired angioedema is associated with lymphoproliferative or autoimmune disease. Conventionally attenuated androgens and antifibrinolytics have been used for prophylaxis, both for the long term and presurgically. Fresh frozen plasma and plasma-derived C1 inhibitor concentrate have been used primarily for treatment of acute attacks. All have drawbacks in side effects or potential for infection transmission. New treatments (recombinant C1 inhibitor, icatibant, DX-88, and for acquired angioedema, rituximab) so far show good safety profiles. Early data suggest these may be effective treatment alternatives. The efficacy of current treatment and the potential held by newer agents that target specific elements in complement or kinin pathways are examined. Some agents are likely to have a wider role in treatment of other, more common, forms of angioedema.

C1-inhibitor prevents non-specific plasminogen activation by a prourokinase mutant without impeding fibrin-specific fibrinolysis.

Prourokinase (prouPA) is unstable in plasma at therapeutic concentrations. A mutant form, M5, made more stable by reducing its intrinsic activity was therefore developed. Activation to two-chain M5 (tcM5) induced a higher catalytic activity than two-chain urokinase plasminogen activator (tcuPA), implicating an active site functional difference. Consistent with this, an unusual tcM5 complex with plasma C1-inhibitor was recently described in dog and human plasma. The effect of C1-inhibitor on fibrinolysis and fibrinogenolysis by M5 is the subject of this study. Zymograms of tcM5 and tcuPA incubated in plasma revealed prominent tcM5-C1-inhibitor complexes, which formed within 5 min. The inhibition rate by purified human C1-inhibitor (250 microg mL(-1)) was about 7-fold faster for tcM5 than it was for tcuPA (10 microg mL(-1)). The effect of the inhibitor on the stability of M5 and prouPA was determined by incubating them in plasma at high concentrations (10-20 microg mL(-1)) +/- C1-inhibitor supplementation. Above 10 microg mL(-1), depletion of all plasma plasminogen occurred, indicating plasmin generation and tcM5/tcuPA formation. With supplemental C1-inhibitor, M5 stability was restored but not prouPA stability. Clot lysis by M5 +/- supplemental C1-inhibitor showed no attenuation of the rate of fibrinolysis, whereas fibrinogenolysis was prevented by C1-inhibitor. Moreover, because of higher dose-tolerance, the rate of fibrin-specific lysis reached that achievable by non-specific fibrinolysis without inhibitor. Plasma C1-inhibitor stabilized M5 in its proenzyme configuration in plasma by inhibiting tcM5 and thereby non-specific plasminogen activation. At the same time, fibrin-specific plasminogen activation remained unimpaired. This unusual dissociation of effects has significant implications for improving the safety and efficacy of fibrinolysis.

A Direct Role for C1 Inhibitor in Regulation of Leukocyte Adhesion

Plasma C1 inhibitor (C1INH) is a natural inhibitor of complement and contact system proteases. Heterozygosity for C1INH deficiency results in hereditary angioedema, which is mediated by bradykinin. Treatment with plasma C1INH is effective not only in patients with hereditary angioedema, but also in a variety of other disease models, in which such therapy is accompanied by diminished neutrophil infiltration. The underlying mechanism has been explained primarily as a result of the inhibition of the complement and contact systems. We have shown that C1INH expresses the sialyl-Lewis(x) tetrasaccharide on its N-linked glycan, via which it binds to E- and P-selectins and interferes with leukocyte-endothelial adhesion in vitro. Here we show that both native C1INH and reactive center cleaved C1INH significantly inhibit selectin-mediated leukocyte adhesion in several in vitro and in vivo models, whereas N-deglycosylated C1INH loses such activities. The data support the hypothesis that C1INH plays a direct role in leukocyte-endothelial cell adhesion, that the activity is mediated by carbohydrate, and that it is independent of protease inhibitory activity. Direct involvement of C1INH in modulation of selectin-mediated cell adhesion may be an important mechanism in the physiologic suppression of inflammation, and may partially explain its utility in therapy of inflammatory diseases.

Clinical studies of recombinant human C1 inhibitor in subjects with hereditary angioedema

RATIONALE: Recurrent attacks of subcutaneous and/or submucosal angioedema in subjects with hereditary angioedema (HAE) results from uncontrolled activation of the contact and complement systems due to a congenital deficiency of functional C1 inhibitor in plasma. Recombinant human C1 inhibitor (rhC1INH) was isolated from the milk of transgenic rabbits for replacement therapy in subjects with HAE. RhC1INH was highly similar to plasma-derived C1INH and appeared safe in preclinical studies.

C1 inhibitor gene expression in patients with hereditary angioedema: Quantitative evaluation by means of real-time RT-PCR

Hereditary angioedema (HAE) is caused by heterozygous defects in the C1 inhibitor (C1-INH) gene (SERPING1/C1NH). In patients' plasma C1-INH levels range between 5% and 30% of normal levels (ie, far from the 50% expected for an autosomal dominant defect). Most patients have antigenic and functional deficiency (type I HAE), and 15% have reduced C1-INH function but normal to increased antigen because of the presence of a dysfunctional protein (type II HAE). We sought to contribute to the understanding of the pattern of C1-INH gene expression in patients with HAE. We used real-time quantitative RT-PCR to measure C1-INH mRNA levels in PBMCs of 57 patients with HAE typed for mutations in the SERPING1/C1NH gene. Thirty-six different mutations were identified in genomic DNA. Compared with healthy control subjects, C1-INH mRNA was significantly and similarly reduced in patients with type I and type II HAE (40% and 47%, respectively; P <.0001). By means of direct sequencing of cDNAs, we found that 74% of patients with type I HAE carrying small mutations presented significant amounts of mutated transcripts at the mRNA level, suggesting that both allelic mRNA products were reduced to approximately 50%. In 4 patients carrying large deletions expected to fully inactivate expression from the mutant allele, C1-INH mRNA was 23% on average compared with that seen in control subjects, confirming that normal mRNA was strongly underexpressed. These new findings, combined with previous evidence of increased C1-INH consumption, might explain the plasma levels of normal C1-INH that are markedly less than the expected 50%.

Practical genetics: alpha-1-antitrypsin deficiency and the serpinopathies.

Alpha-1-antitrypsin (alpha(1)-antitrypsin) is the archetypal member of the serine proteinase inhibitor or serpin superfamily. The most common severe deficiency variant is the Z allele, which results in the accumulation of mutant protein within hepatocytes. This 'protein overload' causes neonatal hepatitis, cirrhosis and hepatocellular carcinoma. The lack of circulating plasma alpha(1)-antitrypsin results in early-onset panlobular emphysema. The mechanism underlying the deficiency of Z alpha(1)-antitrypsin is due to an aberrant conformational transition within the protein and the formation of chains of polymers that tangle within the secretory pathway of hepatocytes. This mechanism also underlies the plasma deficiency of other members of the serpin superfamily to cause a class of diseases called the serpinopathies. Specifically mutant alleles of antithrombin, C1-inhibitor and alpha(1)-antichymotrypsin have been reported that favour the spontaneous formation of polymers and the retention of protein within hepatocytes. The consequent lack of plasma antithrombin, C1-inhibitor and alpha(1)-antichymotrypsin results in thrombosis, angio-oedema and emphysema, respectively. Moreover, the polymerisation of mutants of neuroserpin results in the retention of polymers within neurones to cause the inclusion body dementia, familial encephalopathy with neuroserpin inclusion bodies or FENIB. We review here the genetic and molecular basis and clinical features of alpha(1)-antitrypsin deficiency, and show how this provides a platform to understand the other serpinopathies.

Recombinant human C1-inhibitor produced in Pichia pastoris has the same inhibitory capacity as plasma C1-inhibitor

Therapeutic application of the serpin C1-inhibitor (C1-Inh) in inflammatory diseases like sepsis, acute myocardial infarction and vascular leakage syndrome seems promising, but large doses may be required. Therefore, a high-yield recombinant expression system for C1-Inh is very interesting. Earlier attempts to produce high levels of C1-Inh resulted in predominantly inactive C1-Inh. We describe the high yield expression of rhC1-Inh in Pichia pastoris, with 180 mg/l active C1-Inh at maximum. On average, 30 mg/l of 80–100% active C1-Inh was obtained. Progress curves were used to study the interaction with C1s, kallikrein, coagulation factor XIIa and XIa, and demonstrated that rhC1-Inh had the same inhibitory capacity as plasma C1-Inh. Structural integrity, as monitored via heat stability, was comparable despite differences in extent and nature of glycosylation. We conclude that the P. pastoris system is capable of high-level production of functionally and structurally intact human C1 inhibitor.

Changes in cardiovascular risk factors in obesity following bariatric surgery

The incubation of normal human plasma in the presence of sulphatide vesicles results in the generation of amidolytic activity due to factor XIIa (FXIIa) and to kallikrein (KRN). The progress of the generation of the enzymes distinguished a high initial rate of enzyme generation, a decline of this rate to maximum amidolytic activity ([FXIIa]m and [KRN]m) and a negative pseudo-first-order rate attributed to enzyme inactivation by plasma C1-inhibitor (C1INH). [FXIIa]m and [KRN]m were determined after the treatment of various dilutions of plasma in the presence of 4, 15, or 40 μM sulphatide vesicles. At all levels of sulphatides, [FXIIa]m and [KRN]m initially increased with the concentration of plasma, to reach a plateau at higher concentration of plasma. The plateau activities of the generated enzymes and the optimal concentration of plasma both increased with the level of sulphatide vesicles. The pseudo-first-order inactivation rate for KRN increased progressively with the concentration of plasma but the respective rate for FXIIa was independent of the plasma concentration. The data suggest that contiguous binding of plasma FXIIa, factor XII (FXII), and the complexes of high molecular weight kininogen (HK) with prekallikrein (HK-PKRN) and factor XI (HK-FXI) to an electronegative surface induces a rapid generation of FXIIa and KRN. The concentration of the electronegative surface controls the levels of generated FXIIa and KRN and their release to the bulk phase. The released FXIIa and KRN are both inactivated by C1INH.

High Affinity Binding of Factor XIIa to an Electronegative Surface Controls the Rates of Factor XII and Prekallikrein Activation in Vitro

The incubation of normal human plasma in the presence of sulphatide vesicles results in the generation of amidolytic activity due to factor XIIa (FXIIa) and to kallikrein (KRN). The progress of the generation of the enzymes distinguished a high initial rate of enzyme generation, a decline of this rate to maximum amidolytic activity ([FXIIa]m and [KRN]m) and a negative pseudo-first-order rate attributed to enzyme inactivation by plasma C1-inhibitor (C1INH). [FXIIa]m and [KRN]m were determined after the treatment of various dilutions of plasma in the presence of 4, 15, or 40 μM sulphatide vesicles. At all levels of sulphatides, [FXIIa]m and [KRN]m initially increased with the concentration of plasma, to reach a plateau at higher concentration of plasma. The plateau activities of the generated enzymes and the optimal concentration of plasma both increased with the level of sulphatide vesicles. The pseudo-first-order inactivation rate for KRN increased progressively with the concentration of plasma but the respective rate for FXIIa was independent of the plasma concentration. The data suggest that contiguous binding of plasma FXIIa, factor XII (FXII), and the complexes of high molecular weight kininogen (HK) with prekallikrein (HK-PKRN) and factor XI (HK-FXI) to an electronegative surface induces a rapid generation of FXIIa and KRN. The concentration of the electronegative surface controls the levels of generated FXIIa and KRN and their release to the bulk phase. The released FXIIa and KRN are both inactivated by C1INH.

C1-Inhibitor substitution as ultima ratio therapy in septic shock: experience with 15 patients

Plasma C1-Inhibitor (C1-INH) is the major inhibitor of both the classical pathway of complement system and contact activation. A relative deficiency of C1-INH has been proposed as an important contributor to the development of shock and organ failure [1,2]. Among others, our working group has published two cases of successful C1-INH substitution in septic patients recently [3].

Activation of the plasma contact system and hemodynamic changes after graft revascularization in liver transplantation.

In this study, the relation between activation of the plasma contact system and hemodynamic changes during orthotopic liver transplantation was evaluated. Nineteen consecutive courses of OLT in 17 adult patients were investigated. Veno-venous bypass was used in all patients. Blood samples were drawn through all phases of the procedure, and analyzed for the following parameters using functional techniques (chromogenic peptide substrate assays): plasma kallikrein (KK), prekallikrein, functional plasma kallikrein inhibition, C1 inhibitor, and alpha 2-macroglobulin. Plasma high molecular weight kininogen (HK) degradation was evaluated using the immunoblotting technique. An abrupt rise in KK activities occurred within 1 min after portal reperfusion of the liver graft (7-16 U/L, P < 0.05). Simultaneously, proteolytic breakdown of HK was seen. The elevated KK activities were maintained the next 1 1/2 hr. Ten min after graft reperfusion, a significant increase in cardiac output compared with the anhepatic phase (7.2-12.4 L/min, P < 0.05) was found. At the same time, systemic vascular resistance fell significantly (817-408 dynes x sec/cm-5, P < 0.05). The increase in plasma KK activities accompanied by simultaneous degradation of HK seen immediately after reperfusion of the liver graft may be due to contact activation as recipient blood contacts with the underlying basement membrane of injured sinusoidal endothelium in the transplanted liver. We suggest that hemodynamic changes associated with the postreperfusion syndrome seen after revascularization of the liver in OLT could at least be caused in part by bradykinin release due to contact activation.

Plasma C1 Inhibitor in Malignant Diseases: Functional Activity versus Concentration

Plasma C1 inhibitor (C1-INH) was determined in 64 patients with different malignant diseases and in 58 healthy persons. C1-INH antigen concentration was measured by radial immunodiffusion (RID), whereas its functional activity was assayed with chromogenic substrate. Within-run and day-to-day precision of both methods were good, with CVs ranging from 3.6 to 5.4%. Plasma C1-INH antigen concentrations were significantly higher in the patients than in healthy controls (P = 4.0 x 10(-3)), as were their C1-INH functional activities (P = 3.5 x 10(-3)). C1-INH activities obtained in the patient plasma samples were in correlation with their antigen concentrations (r = 0.914), showing that C1-INH synthesized in malignant disease was functional. However, the specific activity of the C1-INH (functional activity/antigen concentration ratio) was significantly lower in the patient group as compared with the controls (P = 2.1 x 10(-6)), indicating partial inactivation of plasma C1-INH in malignant diseases. The C1-INH specific activity in patients was inversely proportional to its antigen concentration.

Inactivation of human plasma C1-inhibitor by human PMN leucocyte matrix metalloproteinases

Highly purified human polymorphonuclear (PMN) leucocyte matrix metalloproteinases, collagenase and gelatinase, cleaved human plasma C1-inhibitor at the carboxyl site of Ala 439 (P 6). This led to a concomitant loss of C1-inhibitor activity. An additional cleavage site at the carboxyl site of Ser 441 (P 4), was observed during PMN leucocyte gelatinase-induced inactivation, and a minor fragment of the plasma C1-inhibitor was generated.

POSSIBLE ROLE OF EXTRACELLULARLY RELEASED PHAGOCYTE PROTEINASES IN COAGULATION DISORDER DURING LIVER TRANSPLANTATION

Orthotopic liver transplantation is frequently associated with a complex coagulation disorder, influencing the outcome of the procedure. In this respect, disseminated intravascular coagulation (DIC) had been suggested to be of causative importance for bleeding complications after reperfusion of the liver graft. In 10 consecutive patients undergoing orthotopic liver transplantations, we studied the occurrence of two phagocyte proteinases of different origin in the graft liver perfusate and in systemic blood during the operation, as well as their effects on hemostasis. As compared with plasma samples taken at the end of the anhepatic phase, highly significant increases of cathepsin B and thrombin-anti-thrombin III complexes (TAT), as well as highly significant decreases in antithrombin III, protein C, and C1-inhibitor were observed in graft liver perfusate. Von Willebrand factor and fibrinogen were slightly decreased, whereas the elastase-alpha 1 proteinase inhibitor complexes (EPI) were elevated. In plasma the activity of cathepsin B remained unchanged during the prereperfusion phases, but immediately after revascularization of the graft this cysteine proteinase increased. The EPI showed a gradual increase in plasma during the preanhepatic and anhepatic phases but a more pronounced increase in the reperfusion phase. In parallel with the rise in these two proteinases TAT increased and the activities of antithrombin III and C1-inhibitor in plasma decreased after reperfusion. At 12 hr after revascularization plasma levels of TAT, antithrombin III, and C1-inhibitor had returned to the prereperfusion ranges, whereas cathepsin B and EPI were significantly above the baseline levels. These observations are consistent with the hypothesis that extracellularly released lysosomal proteinases may play a role in the development of a DIC-like constellation, including thrombin formation after revascularization of the liver graft. For the first time we could prove the occurrence of phagocyte proteinases in graft liver perfusate and evaluate the importance of these proteinases for the understanding of the pathophysiology leading to bleeding complications in patients undergoing orthotopic liver transplantation.

Plasminogen activators in dextran sulfate-activated euglobulin fractions: a molecular analysis of factor XII- and prekallikrein- dependent fibrinolysis

To elucidate the mechanism by which activation of the contact system of blood coagulation leads to expression of fibrinolytic activity, we have determined the molecular characteristics of the plasminogen activators present in dextran sulfate-treated euglobulin fractions by electrophoretic-zymographic analysis and specific immunoadsorption. In addition to free and protease inhibitor-bound tissue-type plasminogen activator (t-PA), dextran sulfate precipitates of euglobulins contained the complex formed between plasma kallikrein and C1-inhibitor, an indicator of prekallikrein activation. These precipitates also contained substantial fibrinolytic activity related to urinary-type plasminogen activator (u-PA). Autoradiographic analysis was then used to evaluate the cleavage of 125I-single-chain u-PA (prourokinase) in dextran sulfate euglobulins as well as after exposure to kallikrein or beta-factor XIIa. This analysis supported the conclusion that plasma kallikrein-mediated cleavage and activation of single-chain u-PA is the mechanism operative for the development of lytic activity in euglobulin precipitates following activation of the contact system.

Plasminogen Activators in Dextran Sulfate-Activated Euglobulin Fractions: A Molecular Analysis of Factor XII- and Prekallikrein-Dependent Fibrinolysis

To elucidate the mechanism by which activation of the contact system of blood coagulation leads to expression of fibrinolytic activity, we have determined the molecular characteristics of the plasminogen activators present in dextran sulfate-treated euglobulin fractions by electrophoretic-zymographic analysis and specific immunoadsorption. In addition to free and protease inhibitor-bound tissue-type plasminogen activator (t-PA), dextran sulfate precipitates of euglobulins contained the complex formed between plasma kallikrein and C1-inhibitor, an indicator of prekallikrein activation. These precipitates also contained substantial fibrinolytic activity related to urinary-type plasminogen activator (u-PA). Autoradiographic analysis was then used to evaluate the cleavage of 125I-single-chain u-PA (prourokinase) in dextran sulfate euglobulins as well as after exposure to kallikrein or beta-factor XIIa. This analysis supported the conclusion that plasma kallikrein-mediated cleavage and activation of single-chain u-PA is the mechanism operative for the development of lytic activity in euglobulin precipitates following activation of the contact system.

Effects of fibroblasts and endothelial cells on inactivation of target proteases by protease nexin-1, heparin cofactor II, and C1-inhibitor.

Previous studies have shown that glycosaminoglycans in the extracellular matrix accelerate the inactivation of target proteases by certain protease inhibitors. It has been suggested that the ability of the matrix of certain cells to accelerate some inhibitors but not others might reflect the site of action of the inhibitors. Previous studies showed that fibroblasts accelerate the inactivation of thrombin by protease nexin-1, an inhibitor that appears to function at the surface of cells in extravascular tissues. The present experiments showed that endothelial cells also accelerate this reaction. The accelerative activity was accounted for by the extracellular matrix and was mostly due to heparan sulfate. Fibroblasts but not endothelial cells accelerated the inactivation of thrombin by heparin cofactor II, an abundant inhibitor in plasma. This is consistent with previous suggestions that heparin cofactor II inactivates thrombin when plasma is exposed to fibroblasts and smooth muscle cells. Neither fibroblasts nor endothelial cells accelerated the inactivation of C1s by plasma C1-inhibitor.

Factor XII, Plasma Prekallikrein, α2-Macroglobulin and C1-Inhibitor Levels in Renal Allograft Recipients During Immunosuppression with Cyclosporin A – Sequential Measurements over Four Months in 17 Patients

Factor XII clotting activity (F XII), plasma prekallikrein amidolytic activity (PK), alpha 2-Macroglobulin (alpha 2-M) and C1-Inhibitor (C1-Inh) antigens have been measured in 17 patients immediately before and sequentially for up to four months after kidney transplantation. Before transplantation mean F XII and PK levels were normal (99 +/- 27% and 102 +/- 21%, respectively, mean +/- S.D.) and alpha 2-M and C1-Inh levels were slightly elevated (115 +/- 55% and 129 +/- 32%, respectively, mean +/- S.D.). In the first two weeks after transplantation a significant decrease of F XII to 65 +/- 27%, of PK to 67 +/- 20% and of alpha 2-M to 88 +/- 42%, and a rise of C1-Inh to 201 +/- 44% (mean +/- S.D.) were observed (2 p less than 0.005). F XII levels four month after operation remained significantly (2 p less than 0.05) lower than preoperatively. PK and alpha 2-M values, however, were significantly higher (2 p less than 0.05) at four months as compared to the pretransplant period. Mean F XII levels in the 17 patients at various time points after transplantation correlated positively with PK, alpha 2-M and serum albumin and negatively with CyA level and dose and serum bilirubin. PK and alpha 2-M correlated positively with each other and albumin and negatively with creatinine, bilirubin and CyA (2 p less than 0.01). Whether CyA has a direct influence on production or consumption of F XII, PK, alpha 2-M and C1-Inh, or whether the changes merely reflect altered protein metabolism awaits further study.