Human blood plasma advanced oxidation protein products (AOPP) correlates with fibrinogen levels

In 1996 a novel oxidative stress biomarker, referred to as advanced oxidation protein products (AOPP) was detected in the plasma of chronic uremic patients. The aim of the present studies was to find out that which plasma fraction(s) is responsible for AOPP reactivity. Thermal treatment of pooled samples of human citrate-plasma or EDTA-plasma at 50 degrees C resulted in a rapid and parallel loss of fibrinogen concentration and AOPP reactivity. On the basis of time course and t1/2 values following thermal treatment, AOPP was indistinguishable from fibrinogen. There was a statistically significant (p < 0.0001) correlation between levels of blood plasma fibrinogen and AOPP in patients (n = 61) with various peripheral vascular or cardiovascular diseases. There was also a significant (p < 0.0001) relationship between plasma levels of fibrinogen and molar AOPP/fibrinogen ratio indicating that higher fibrinogen concentrations were associated with more oxidatively transformed groups on the molecule. Results of the present studies suggest that post-translationally modified fibrinogen is a key molecule responsible for human plasma AOPP reactivity. It remains to be elucidated what is the pathophysiological significance of the post-translationally modified fibrinogen in the inflammation-associated events of atherosclerosis, in platelet aggregation, and as a cardiovascular risk biomarker.     

Plasma fibrinogen levels in type II diabetics

Plasma fibrinogen levels measured by an immunoassay method on 170 type II diabetic patients exhibited a bimodal distribution with one small population demonstrating levels greater than those of the normal reference range. The mean plasma level of fibrinogen in the type II diabetics was higher than that of the normal population. Spearman's correlations demonstrated statistically significant positive relationships in type II diabetic patients between fibrinogen levels and fasting glucose levels, serum cholesterol, glycosylated hemoglobin and urinary albumin excretion rate. These relationships suggest that increased plasma fibrinogen may be another marker for coronary heart disease complications encountered by diabetics.     

Human blood plasma advanced oxidation protein products (AOPP) correlates with fibrinogen levels

In 1996 a novel oxidative stress biomarker, referred to as advanced oxidation protein products (AOPP) was detected in the plasma of chronic uremic patients. The aim of the present studies was to find out that which plasma fraction(s) is responsible for AOPP reactivity. Thermal treatment of pooled samples of human citrate-plasma or EDTA-plasma at 50°C resulted in a rapid and parallel loss of fibrinogen concentration and AOPP reactivity. On the basis of time course and t_1/2 values following thermal treatment, AOPP was indistinguishable from fibrinogen. There was a statistically significant (p < 0.0001) correlation between levels of blood plasma fibrinogen and AOPP in patients (n = 61) with various peripheral vascular or cardiovascular diseases. There was also a significant (p < 0.0001) relationship between plasma levels of fibrinogen and molar AOPP/fibrinogen ratio indicating that higher fibrinogen concentrations were associated with more oxidatively transformed groups on the molecule. Results of the present studies suggest that post-translationally modified fibrinogen is a key molecule responsible for human plasma AOPP reactivity. It remains to be elucidated what is the pathophysiological significance of the post-translationally modified fibrinogen in the inflammation-associated events of atherosclerosis, in platelet aggregation, and as a cardiovascular risk biomarker.     

The Quantitative Determination of Fibrinogen in Normal Bovine Plasma and in Cows with Inflammatory Conditions

The quantitative determination of fibrinogen in normal plasma and in cows with inflammatory conditions. A rapid method for the quantitative determination of fibrinogen in bovine plasma is described. The method was employed in the determination of normal values in a material consisting of 100 cows and 50 calves and young animals of various ages. The mean value of the groups of cows was approximately 0.550 g/100 ml. For young animals it was somewhat lower and for cows in the last month of gestation moderately higher than in the other groups. The last part of the experiment involves the determination of the fibrinogen and γ-globulin levels in the plasma of 28 hospitalized cows with various inflammatory conditions. Group A in the material contained animals which were clinically cured and Group B animals that died or were killed. Both groups showed a considerable increase in the fibrinogen level. In Group A the mean value fell back to approximately the normal range while in Group B it remained constantly elevated. The sedimentation rate, SR, in human blood is primarily influenced by the fibrinogen content of the plasma. The SR in bovine blood is very low, and the test is therefore of little significance in diagnostic work. In conclusion, the possibility of using the fibrinogen determination in cattle for the same purpose as the SR in human blood is discussed.     

Interactions of prostaglandin E1, bradykinin and histamine and the increase of plasma fibrinogen in rats

Considering that tissue injury caused by laparotomy significantly increases the liver synthesis of plasma fibrinogen, and that PGE1, bradykinin and histamine are released into the injured tissues, the effect of above mentioned inflammatory agents and of the adrenal medulla on plasma fibrinogen levels in rats was studied. The subcutaneous administration of PGE1, bradykinin or histamine does not modify plasma fibrinogen levels acting independently comparing with non-injected animals or injected with the drug vehicle. Bradykinin + histamine did not modify plasma fibrinogen levels either. However the administration of prostaglandin E1 + bradykinin + histamine reproduced the increase of fibrinogen characteristics of laparotomy. This increase was partially but significantly inhibited in rats that had undergone bilateral removal of the adrenal medulla or administration of PGE1 + bradykinin + histamine + bupivacaine (a local anesthetic), but it was not modified when the adrenal medullectomy was unilateral. It is concluded that plasma fibrinogen increase is obtained only when PGE1 acts in presence of bradykinin or histamine and the adrenal medulla should be partially responsible for said increase.     

A synergistic effect of albumin and fibrinogen on immunoglobulin-induced red blood cell aggregation

Therapeutic administration of immunoglobulins (Ig) has the potential to precipitate thrombotic events. This phenomenon may be explained by red blood cell (RBC) aggregation, which can be potentiated by Ig. The contribution of plasma albumin and fibrinogen to Ig-induced RBC aggregation is unclear. We examined RBC aggregation in three settings: 1) patients receiving therapeutic infusions of Ig; 2) patients receiving plasma supplemented in vitro with Ig; and 3) patients receiving RBC suspensions in standard buffer with varying concentrations of albumin, Ig, and fibrinogen. Ig infusion augmented aggregation of RBCs from patients with normal or high plasma levels of albumin but decreased aggregation in those with lower plasma albumin concentrations. In vitro, RBC aggregation was significantly increased only when all three components, fibrinogen, albumin, and Ig, were present at or above normal concentrations in the suspension but was unaffected when any one of the components was absent from the suspension. Our results suggest a three-way interaction among fibrinogen, Ig, and albumin that synergistically induces RBC aggregation in plasma. Understanding these interactions may help predict clinically important phenomena related to RBC aggregation, such as thrombotic complications of Ig infusion.     

Interactions of prostaglandin E1, bradykinin and histamine and the increase of plasma fibrinogen in rats

Considering that tissue injury caused by laparotomy significantly increases the liver synthesis of plasma fibrinogen, and that PGE₁, bradykinini and histamine are released into the injured tissues, the effect of above mentioned inflammatory agents and of adrenal medulla on plasma fibrinogen levels in rats was studied. The subcutaneous administration of PGE₁, bradykinin or histamine does not modify plasma fibrinogen levels acting independently comparing with non-injected animals or injected with the drug vehicle. Bradykinin + histamine did not modify plasma fibrinogen levels either. However the administration of prostaglandin E₁ + bradykinin + histamine reproduced the increase of fibrinogen characteristics of laparotomy. This increase was partially but significantly inhibited in rats that had undergone bilateral removal of the adrenal medulla or administration of PGE₁ + bradykinin + histamine + bupivacaine (a local anesthetic), but it was not modified when the adrenal medullectomy was unilateral. It is concluded that plasma fibrinogen increase is obtained only when PGE₁ acts in presence of bradykinin or histamine and the adrenal medulla should be partially responsible for said increase.     

Increased synthesis rate of fibrinogen as a basis for its elevated plasma levels in obese female adolescents

Increased concentrations of plasma fibrinogen, an independent risk factor for cardiovascular disease (CVD), in obese children have been reported. The underlying mechanism for this, however, remains to be defined. In the current study, we measured the fractional synthesis rates (FSR) of plasma fibrinogen in six healthy postpubertal obese girls [body mass index (BMI) 36.6 +/- 1.8 kg/m(2); age 16.6 +/- 0.5 yr] and six age-matched lean normal control girls (BMI 20.8 +/- 0.7 kg/m(2); age 16.4 +/- 0.4 yr) during a primed, continuous infusion of L-[1-(13)C]leucine in the postabsorptive state. The method involved purification of plasma fibrinogen by use of immunoaffinity chromatography followed by measurement of [(13)C]leucine enrichment using gas chromatography-combustion-isotope ratio mass spectrometry. The FSR of fibrinogen in obese girls (35.06 +/- 2.61%/day) was almost double that in lean girls (17.02 +/- 1.43%/day), and this increase was associated with a relative increase in plasma concentration of fibrinogen as well as BMI in the subjects studied. Obese subjects had high fasting insulin levels (138 +/- 47 pmol/l) compared with lean subjects (54 +/- 11 pmol/l), whereas their glucose concentrations were similar (4.5 +/- 0.3 mmol/l in obese and 4.4 +/- 0.4 mmol/l in lean subjects), suggesting insulin resistance. The doubling of the FSR of fibrinogen provides novel insight into the mechanism of elevated levels of plasma fibrinogen and suggests a primary role for increased synthesis in producing the hyperfibrinogenemia associated with obesity. This finding may have important implications in the design of therapies for modulating plasma fibrinogen levels in obesity and/or CVD in childhood.     

The Fibrinogen Concentration in Blood of Dairy Cows and its Influence on the Interpretation of the Glutaraldehyde and Formol-Gel Test Reactions

It has earlier been shown that the formol-gel test on serum and glutaraldehyde test on whole blood are simple and rapid methods for evaluation or the immunoglobulin status in the cow. Both tests function as coagulation tests in which aldehyde groups oross-link basic blood globulins at their NH2-groups, forming polymerisates. The glutaraldehyde has in whole blood the capacity to polymerize not only immunoglobulins but also fibrinogen. This investigation was made in order to study whether the fibrinogen level may influence the result of the glutaraldehyde test, so revealing any differences between the results of that and the formol-gel test carried out on serum. In 92 cows with a variety of clinical disorders (most of them with inflammatory processes) the total protein, albumin, total globulin concentration and albumin/globulin ratio in serum and fibrinogen concentration in plasma were recorded. The material was grouped according to glutaraldehyde and formol-gel test reactions. It is shown that increases in the fibrinogen level have an effect on the results of the glutaraldehyde test. A positive glutaraldehyde test in more acute processes is ascribed to a heavy rise of plasma fibrinogen in its capacity of acute-phase protein. A positive glutaraldehyde test in chronic diseases may be viewed as a result of interaction between high immunoglobulin concentrations and elevated fibrinogen concentration. In conclusion the fibrinogen and immunoglobulin status of blood is important to assess in many diseases of cattle. The semiquantitative tests described for field use can separately, or especially in parallel use, provide valuable information about the character and development of a disease and may be regarded as good substitutes for the sedimentation rate (SR), which is not demonstrable in cattle. kw|Keywords|k]bovine fibrinogen; k]bovine serum proteins; k]formol-gel reaction; k]glutaraldehyde test; k]acute and chronic inflammations     

Advanced oxidation protein products (AOPP): novel uremic toxins, or components of the non-enzymatic antioxidant system of the plasma proteome?

In 1996, a novel oxidative stress biomarker, referred to as advanced oxidation protein products (AOPP), was detected in the plasma of chronic uremic patients. It was suggested that AOPP measure highly oxidized proteins, especially albumin. Recent data in turn appear to indicate that oxidized fibrinogen is the key molecule responsible for the AOPP reaction in the human plasma. Since fibrinogen is an acute-phase reactant, it is evident that during each episode of inflammatory response, the antioxidant capacity of the plasma is enhanced. In this context, fibrinogen can be regarded as a component of the antioxidant system of the plasma proteome. It was also demonstrated that oxidized fibrinogen is bound to apolipoprotein(a) of lipoprotein(a) via lysine binding sites. Thus, apo(a) could compete with plasminogen (and/or tissue plasminogen activator) for its binding sites of fibrin(ogen), causing inhibition of fibrinolysis, and thereby promote atherosclerosis and cardiovascular disease.     

Increased thrombin-induced polymerization of fibrinogen associated with high protein carbonyl levels in plasma from patients post myocardial infarction

Increased levels of protein carbonyls have been measured in plasma of patients following a myocardial infarction (Mocatta et al. J. Am. Coll. Cardiol. 49:1993–2000; 2007). In this study, we have examined representative plasma samples from this group of patients. We show that carbonyls are formed preferentially on fibrinogen and that there is a strong correlation between fibrinogen and total plasma protein carbonyls. Functional properties of fibrinogen isolated from post myocardial plasmas were investigated by measuring thrombin-catalyzed polymerization. Fibrinogen from plasma with upper quartile protein carbonyls (mean 0.16 nmol/mg protein) polymerized ～1.4 times more rapidly and gave 1.4-fold higher maximum turbidity (12 per group, P < 0.001) than fibrinogen from lower quartile carbonyl plasma (mean 0.007 nmol/mg), which behaved similarly to control plasma. Significant differences were also apparent when related to the carbonyl content of the fibrinogen itself. These changes in the high carbonyl plasma reflect a faster rate of lateral aggregation of small oligomers to form fibrin polymers that comprise thicker, more loosely woven fibers. In vivo this could be translated into a tendency to clot faster and form more fragile clots. High protein carbonyls in fibrinogen were not associated with an increased content of nitrotyrosine or chlorotyrosine. Nitrotyrosine levels were significantly lower in fibrinogen than total plasma protein, with no difference between patients and controls. Chlorotyrosine levels in fibrinogen (but not total protein) were significantly higher for the post myocardial patients. Our data suggest that high fibrinogen protein carbonyls in heart disease could be a prothrombotic risk factor.     

双歧杆菌对迁延性及慢性腹泻患儿部分胃肠激素的影响

目的：探讨双歧杆菌对迁延性及慢性腹泻患儿血清胃泌素(GAS)、血浆胃动素(MOT)和血浆生长抑素(SS)水平的影响。方法：应用放免法测定20例用双歧杆菌和19例未用微生态制剂治疗的迁延性及慢性腹泻患儿治疗前和治疗7d后空腹及餐后1h的血清GAS、血浆MOT和血浆SS水平,结果与20例正常儿童比较,并分析三种胃肠激素间的相关性。结果：治疗前两组病例空腹和餐后的血清GAS、血浆MOT和血浆SS水平高于正常儿童。除空腹未用微生态制剂病例的血清GAS水平外,治疗前两组病例血中的三种胃肠激素水平高于治疗7d后。治疗7d后双歧杆菌治疗病例空腹和餐后的血清GAS和血浆MOT水平低于未用微生态制剂病例,且血清GAS水平与正常儿童差异无显著性。治疗7d后血浆SS水平三组间差异无显著性。另两组病例治疗前空腹和餐后三种胃肠激素间均无关联,治疗7d后的双歧杆菌治疗病例和正常儿童GAS与SS和MOT与SS则呈正相关。结论：双歧杆菌制剂对迁延性及慢性腹泻儿童胃肠激素的分泌有调理作用,应用双歧杆菌治疗的患儿血清GAS、血浆MOT和血浆SS水平较未用微生态制剂的患儿更快恢复正常。     

Effect of chronic consumption of alcohol on the hypothalamo-pituitary-testicular axis in the rat

An experimental model of chronic alcohol abuse is developed, in order to study the hypothalamic-pituitary testicular axis in the rat. For this purpose basal plasma prolactin, gonadotropins, testosterone and estradiol have been measured. Also these hormones were studied after LHRH or hCG stimulation. This experimental model allows us to study the role of alcohol in hypogonadism induction. Chronic alcohol administration resulted in an inconstant decrease in plasma testosterone levels and very diminished response of it to hCG. Along with these modifications, there was an increase in basal plasma estrogen levels, as has been shown in the human. The decrease in plasma LH levels in alcoholic rats together with a normal response to LHRH suggest a toxic role of alcohol at higher levels than the pituitary. The existence of a hyperprolactinemic state under chronic alcohol ingestion is confirmed. The decrease in plasma prolactin levels after LHRH administration suggests that prolactin and gonadotropin secretion are very closely related.     

Alterations in the common pathway of coagulation during weight loss induced by gastric bypass in severely obese patients

The objective of this study was to establish the relationship between the plasminogen activator inhibitor-1 (PAI-1), antithrombin-III (ATIII), fibrinogen, and white blood cell (WBC) levels in severely obese patients. We analyzed various plasma parameters implicated in the intrinsic and extrinsic coagulation pathway from 34 severely obese patients before and 1, 6, and 12 months after gastric bypass. In obese people, ATIII, fibrinogen, and WBC levels were in the upper limit of the normal range, and all were higher and significantly different from nonobese people. After bariatric surgery, the ATIII level continued to be high during the first month and increased until 12 months, while fibrinogen decreased only at that time. PAI-1 plasma protein and PAI-1 mRNA levels in liver and adipose tissue show similar profiles and had a strong positive correlation (r = 0.576, P = 0.0003 in liver; r = 0.433, P = 0.0004 in adipose tissue). They were higher in obese patients compared with nonobese control, but tended to recover normal values 1 month after surgery. Thus, the liver and adipose tissue could be an important source of PAI-1 protein in plasma. Gastric bypass surgery leads to a normalization of the hematological profile and a decrease in PAI-1 levels, which entails a decrease of risk for thromboembolism in severely obese.     

Fibrinogen-fibrin degradation products: hemagglutination inhibition immunoassay in plasma.

Immunoassay for fibrinogen and/or fibrin degradation products (FDP) is generally in the clot and hence assay of serum may not reveal the true concentration of FDP in blood. We have developed a hemagglutination inhibition immunoassay for FDP in human plasma. D fragment appears to possess an antigenic determinant, called D-neoantigen, not found in native fibrinogen. Rabbit antiserum produced against D fragment was absorbed with immunosorbent columns coupled with fibrinogen and normal human serum, respectively, so that it contained only those antibodies directed against the neoantigenic determinant of D fragment. In this immunoassay, sheep erythrocytes (SRBC) were stabilized with glutaraldehyde and subsequently sensitized with D fragment by means of tannic acid. Hemagglutination of absorbed anti-D-neoantigen serum against SRBC sensitized with D fragment was titered to be 1:256. The hemagglutination was inhibited by D fragment but not by fibrinogen; the sensitivity of detecting D fragment was 8 mug/ml. Human plasma from normal subjects did not inhibit. This appears to be the first report of a hemagglutination inhibition immunoassay for FDP in plasma.     

Regenerative Surface Plasmon Resonance (SPR) biosensor: real-time measurement of fibrinogen in undiluted human serum using the competitive adsorption of proteins

Epidemiological studies suggest that elevated plasma fibrinogen levels are associated with an increased risk of cardiovascular disorders. Normal fibrinogen level is in the range of 1.5-4.5mg/mL, depending upon both genetic (intrinsic) and environmental (extrinsic) factors. An increase of 0.25mg/mL from the normal level can often be correlated with a high risk of cardiovascular disease. Thus, it is useful to monitor fibrinogen level in serum of a patient for clinical diagnosis. We report a regenerative biosensor that measures real-time fibrinogen levels in undiluted serum. The biosensor uses Surface Plasmon Resonance (SPR), highly sensitive optical technique. The biosensor does not use bio-receptors (i.e., antibodies, enzymes, DNA, etc.) unlike conventional biosensors, and deploys the nature of competitive adsorption of proteins to achieve selective detection of fibrinogen. We measured fibrinogen-spiked serum samples with a concentration of 1.5-4.5 mg/mL, and repeated six measurement trials to obtain statistical distribution of the measurements using the regeneration method of the sensing surface. The SPR biosensor has a sensitivity of 42 mDeg/(mg/mL) for a fibrinogen concentration in the range of 0.5-2.5 mg/mL, whereas it was hard to correlate the measurements to the spiked-fibrinogen samples of above 2.5 mg/mL.     

R?le of histamine on plasma fibrinogen levels in rats with surgical injury (laparotomy)

The probable r?le of endogenous histamine in the increase of plasma fibrinogen in rats submitted to tissue injury (laparotomy) was studied. In laparotomized rats with 10 mg kg-1 day-1 of diphenhydramine (a H1-histamine receptor blocker) plasma fibrinogen decreased significantly as compared to the group of rats laparotomized only (P less than 0.02), reaching values similar to those observed in rats laparotomized with removal of the adrenal medulla or laparotomized with severing of splanchnic nerves. There is a significant difference between these latter groups and the normal noninjured group (P less than 0.01). Plasma fibrinogen did not modify (as compared with the uninjured group) in rats injected only with histamine (1 mg kg-1 day-1) or with diphenhydramine. Taking into account the results obtained and the mechanism of action of diphenhydramine, it would seen that endogenous histamine takes part in the increase of plasma fibrinogen in laparotomized rats, perhaps indirectly through stimulation of the adrenal medulla secretion.     

Fibrin assembly in human plasma and fibrinogen/albumin mixtures

Magnetic birefringence is used to monitor the kinetics of thrombin-catalyzed fibrin polymerization in model systems of increasing complexity (i.e., fibrinogen solutions, fibrinogen/albumin mixtures, and plasma anticoagulated with citrate) and in plasma containing free calcium which is the physiological condition. The introduction of albumin into fibrinogen solutions shortens the lag period and enhances fiber thickness. The polymerization progress curves are sigmoidal at zero or low albumin concentrations, but at physiological and higher concentrations, they become hyperbola-like from the end of the lag period. High albumin concentration has thus induced a change in the assembly kinetics. The progress curves from plasma in which the cascade is dormant are also hyperbola-like although they round off more quickly because of antithrombin activity. In plasma containing free calcium, thrombin is endogenously produced, and the progress curves are nearly linear; hence, the assembly kinetics are very different from those of the model systems. The curves are not influenced by calcium-dependent cross-linking involving factor XIIIa. The progress curves are also linear when polymerization is induced with Russell's viper venom, which by directly activating factor X circumvents earlier steps in the cascade. This implies that linear polymerization is caused by events posterior to factor X activation and are thus likely to be largely dependent on the functioning of the prothrombinase complex. Addition of thrombin to plasma containing free calcium reduces the lag period. At low exogenous thrombin levels, the polymerization rate is increased, and the progress curves remain linear. However, at higher levels, the curves become more complicated and, paradoxically, full polymerization takes longer.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fibrinolysis in Cholestatic Jaundice

The fibrinolytic system was studied in primary biliary cirrhosis (16 patients) and large bile duct obstruction (10 patients, nine of whom had carcinoma). Plasma fibrinolysis (plasminogen activator activity) was decreased and fibrinogen increased in both groups of patients, particularly in those with large duct obstruction. These changes were related to the degree of cholestasis. Plasminogen activator activity was inversely related to serum triglyceride levels in patients with primary biliary cirrhosis. Urokinase inhibitors were decreased in both groups and antiplasmins increased in patients with large duct obstruction; fibrin/fibrinogen degradation products were normal in primary biliary cirrhosis and moderately increased in large duct obstruction. None of these fibrinolytic indices was related to the degree of cholestasis. Fibrinolytic activity and fibrinogen returned almost to normal levels after palliative surgery in the three patients with large duct obstruction who were studied. The decreased plasma fibrinolysis and increased fibrinogen may be due to altered lipid metabolism in cholestatic jaundice. In patients undergoing surgery for large duct obstruction there may be an increased risk of thrombosis.     

Stimulation of fibrinogen synthesis in cultured rat hepatocytes by fibrinogen fragment E

Because of the inherent difficulties of experimentation in intact animals, we used primary monolayer cultures of non-proliferating adult rat hepatocytes to study the effects of fibrinogen degradation products on fibrinogen biosynthesis. The freshly isolated hepatocytes obtained by collagenase perfusion of the liver in situ were cultured in a chemically defined serum-free medium. The rate of fibrinogen synthesis in control cultures was $\text{40–50}\text{pmol}\text{2.5·10}{}^6$ cells per 24 h. Additions of 20, 60 or 100 μg of homologous stage I fibrinogen degradation products had no effect on fibrinogen synthesis. In contrast, addition of the same amounts of homologous or heterologous (human) stage III fibrinogen degradation products resulted in a concentration-dependent increase in fibrinogen biosynthesis without affecting the rate of synthesis of albumin. When purified stage III fibrinogen degradation products D and E (human) were tested in 10, 30 or 50 μg/3 ml medium only fragment E showed a significant increase in fibrinogen biosynthesis (1.9-, 2.8- and 5.6-fold, respectively, over the control cultures). The presence of excess fibrinogen had no effect. These results suggest that fibrinogen fragment E may be a specific stimulator of fibrinogen biosynthesis which may play an important role in maintaining normal levels of plasma fibrinogen.