Concentrations Of Alpha 2-macroglobulin Research Articles

Use of quantitative mass spectrometry-based proteomics and ELISA to compare the alpha 2 macroglobulin concentration in equine blood-based products processed by three different orthobiologic devices.

Alpha 2 macroglobulin (A2M), a multi-functional protein in the plasma protease inhibitor class, regulates proinflammatory cytokines and the clearance of chondrodestructive enzymes in cases of joint injury and osteoarthritis (OA). The purpose of this study was to compare A2M concentrations in equine plasma samples processed by three commercial devices developed for stall-side regenerative joint therapy. Plasma samples were obtained from healthy adult horses (N = 13). Mass spectrometry analysis was used to determine the concentration of protein analytes in each sample. Selected reaction monitoring measured a specific A2M peptide as a surrogate of the whole A2M protein. A2M concentrations produced by each test device were compared for two sample types: a pre-concentrate or platelet-poor (PP) component and a final component for use in the horse. There was no significant difference (p > 0.05) in the geometric mean (GM) concentration of A2M in the final concentration samples produced by the Alpha2EQ® device (N horses = 13) and the single-centrifugation PP samples produced by the Pro-Stride® APS (autologous protein solution) device (N = 13) and the Restigen® PRP (platelet-rich plasma) device (N = 11). When A2M content in final concentration samples produced by each device was compared, the Pro-Stride APS and Restigen PRP samples had significantly greater GM A2M content (p < 0.0001) compared to the Alpha2EQ samples, and the Pro-Stride APS final concentration samples had significantly greater GM A2M concentration (p < 0.0001) versus that for the Restigen PRP final samples. This comparison demonstrated that the volume and A2M concentration of an Alpha2EQ final concentrate are no different than the volume and concentration of A2M in the PP from Pro-Stride or Restigen devices.

Quench me if you can: Alpha-2-macroglobulin trypsin complexes enable serum biomarker analysis by MALDI mass spectrometry

One of the main functions of alpha-2-macroglobulin (A2M) in human blood serum is the binding of all classes of protease. It is known that trypsin, after such interaction, possesses modified proteolytic activity. Trypsin first hydrolyzes two bonds in A2M’s ‘bait region’, and the peptide 705VGFYESDVMGR715 is released from A2M. In this work, specifics of the A2M-trypsin interaction were used to determine A2M concentration directly in human blood serum using MALDI mass-spectrometry. Following exogenous addition of trypsin to human blood serum in vitro, the concentration of the VGFYESDVMGR peptide was measured, using its isotopically-labeled analogue (18O), and A2M concentration was calculated. The optimized mass spectrometric approach was verified using a standard method for A2M concentration determination (ELISA) and the relevant statistical analysis methods. It was also shown that trypsin’s modified proteolytic activity in the presence of serum A2M can be used to analyze other serum proteins, including potential biomarkers of pathological processes. Thus, this work describes a promising approach to serum biomarker analysis that can be technically extended in several useful directions.

Age-Dependent Variation in Glycosylation Features of Alpha-2-Macroglobulin.

Alpha-2-macroglobulin (A2M) is a glycosylated broad spectrum inhibitor of numerous proteases, including those involved in blood coagulation. Glycosylation characteristics can affect protein structure and function. This study compares glycosylation characteristics of A2M in newborn umbilical cord (NUCP) and adult pooled plasmas. Peptide N-Glycosidase F treatment was used to evaluate the total N-glycan content of the molecules. Neuraminidase treatment, and affinity for Ricinus Communis Agglutinin I were used to examine terminal sialic acid and galactose content, respectively. Two-dimensional (2D) electrophoresis was used to determine charge-related isoform profiles and fluorophore-assisted carbohydrate electrophoresis (FACE) was used to characterize N-glycan profiles. Results revealed no difference in total N-glycan mass, however, a statistically significant difference was shown in the change in charge associated with sialic acid loss in the NUCP A2M population. 2D electrophoresis indicated a lower pI range for NUCP A2M isoforms. In addition, NUCP A2M displayed a trend toward higher terminal galactose quantities than adult A2M. FACE revealed an increased abundance of more branched, higher molecular weight glycans in NUCP A2M. These differences in glycan branching and charged residues may impact A2M receptor-based clearance and thus could be responsible for the increased A2M concentration seen in NUCP, and newborns.

Validation of Serum Test for Advanced Liver Fibrosis in Patients With Nonalcoholic Steatohepatitis

We analyzed markers of fibrosis in serum samples from patients with nonalcoholic fatty liver disease (NAFLD), assessed by liver biopsy. We used serum levels of markers to develop an algorithm to discriminate patients with advanced fibrosis from those with mild or moderate fibrosis and validated its performance in 2 independent cohorts of patients with NAFLD. We performed a retrospective analysis of serum samples from 396 patients with NAFLD and different stages of fibrosis (F0-F4), collected from 2007 through 2017 on the day of liver biopsy (training cohort 1). We measured serum concentrations of alpha-2 macroglobulin (A2M), hyaluronic acid (HA), and TIMP metallopeptidase inhibitor 1 (TIMP1), and used measurements to develop an algorithm that could discriminate patients with NAFLD with advanced fibrosis (F3-F4; 24.1% of cohort) from those with mild or moderate fibrosis (F0-F2; 79.5% of cohort). We validated the algorithm using serum samples collected from a separate 396 patients from the same time period and location (validation cohort 1), as well as 244 patients with NAFLD evaluated at a separate location, from 2011 through 2017, within a median of 11 days of liver biopsy (cohort 2). The algorithm identified patients with advanced fibrosis vs mild or moderate fibrosis in training cohort 1 with an area under the receiver operating characteristic (AUROC) curve of 0.867 (95% CI, 0.827-0.907), 84.8% sensitivity (95% CI, 75.5%-91.0%), and 72.3% specificity (95% CI, 66.9%-77.3%), at a cutoff score of 17. The AUROC for the combined validation cohorts 1 and 2 (n=640) was 0.856 (95% CI, 0.820-0.892), identifying patients with 79.7% sensitivity (95% CI, 71.9%-86.2%) and 75.7% specificity (95% CI, 71.8%-79.4%) at the predetermined cutoff score of 17. The algorithm had negative predictive values that ranged from 92.5% to 94.7% in the validation cohorts; it correctly classified 90.0% of F0 samples, 75.0% of F1 samples, 77.4% of F3 samples, and 94.4% of F4 samples. We developed an algorithm that identifies patients with advanced fibrosis from those with mild to moderate fibrosis in patients with NAFLD with an AUROC value of approximately 0.86, based on levels of serum biomarkers. We validated the findings in 2 separate sets of patients with biopsy-proven NAFLD. The algorithm can be used non-invasively to determine risk of advanced fibrosis in patients with NAFLD.

Alpha-2 macroglobulin in Alzheimer's disease: a marker of neuronal injury through the RCAN1 pathway.

Preclinical changes that precede the onset of symptoms and eventual diagnosis of Alzheimer's disease (AD) are a target for potential preventive interventions. A large body of evidence suggests that inflammation is closely associated with AD pathogenesis and may be a promising target pathway for such interventions. However, little is known about the association between systemic inflammation and preclinical AD pathophysiology. We first examined whether the acute-phase protein, alpha-2 macroglobulin (A2M), a major component of the innate immune system, was associated with cerebrospinal fluid (CSF) markers of neuronal injury in preclinical AD and risk of incident AD in the predictors of cognitive decline among normal individuals (BIOCARD) cohort. We find that A2M concentration in blood is significantly associated with CSF concentrations of the neuronal injury markers, tau and phosphorylated tau, and that higher baseline serum A2M concentration is associated with an almost threefold greater risk of progression to clinical symptoms of AD in men. These findings were replicated in the Alzheimer's Disease Neuroimaging (ADNI) study. Then, utilizing a systems level approach combining large multi-tissue gene expression datasets with mass spectrometry-based proteomic analyses of brain tissue, we identified an A2M gene network that includes regulator of calcineurin (RCAN1), an inhibitor of calcineurin, a well-characterized tau phosphatase. A2M gene and protein expression in the brain were significantly associated with gene and protein expression levels of calcineurin. Collectively these novel findings suggest that A2M is associated with preclinical AD, reflects early neuronal injury in the disease course and may be responsive to tau phosphorylation in the brain through the RCAN1-calcineurin pathway.

Measurement of Protease Activity and Concentration of a Broad Spectrum Protease Inhibitor; Alpha 2-Macroglobulin (A2m) in Plasma of Severely Chronic Ill Patients in Bangladesh

Different types of proteases may play influential roles across a spectrum of diseases. Correspondingly, protease inhibitors have been known to play crucial roles in the patho-physiology of diseases. We hypothesize the activity of the broad spectrum protease inhibitor, alpha-2-macroglobulin (A2M) may hold unexplored roles in diseases as well. Clinically measuring levels of variant proteases in combination with the monitoring of A2M, in plasma, can be a novel approach to further our understanding of diseases related to protease activity. There is evidence showing administration of purified human A2M to animal models with bacteria induced septic shock using Pseudomonas aeruginosa resulted in the reversal of the pathological effect. In addition, recent evidence has also shown the beneficial effects of treatment with autologous A2M in post-traumatic osteoarthritis patients. We investigated the relationship of protease activity and concentration of A2M in plasma of chronic ill patients in Bangladesh. Blood was collected to prepare plasma from patients (n=20) of different categories: severe burn, cancer of different types, enteric fever with psychosis, chronic pancreatitis, chronic liver diseases, different types of kidney diseases, different types of pulmonary diseases, diabetes and hypertension. As control, plasma was prepared from healthy volunteers (n=20) without any known disease(s). Plasma concentrations of A2M were measured by sandwich ELISA (Enzyme– Linked Immunosorbent Assay) and protease activity assay were done by protease activity assay kit. In this study, we have found that all chronically ill patients (n=20) with a verity of diseases, A2M concentration went down significantly (p<0.001) compared to the concentration of normal healthy individuals (n=20). On contrary, the protease activity level were increased significantly (p<0.001) in all chronically ill patients (n=20) compared the activity of normal healthy individuals (n=20). These results clearly indicated an inverse relationship of A2M to the protease activity in chronically ill patients in Bangladesh which may be a novel approach to further our understanding of pathophysiology of diseases related to protease activity.

Novel biomarkers predict liver fibrosis in hepatitis C patients: alpha 2 macroglobulin, vitamin D binding protein and apolipoprotein AI

BackgroundThe gold standard of assessing liver fibrosis is liver biopsy, which is invasive and not without risk. Therefore, searching for noninvasive serologic biomarkers for liver fibrosis is an importantly clinical issue.MethodsA total of 16 healthy volunteers and 45 patients with chronic hepatitis C virus (HCV) were enrolled (F0: n = 16, F1: n = 7, F2: n = 17, F3: n = 8 and F4: n = 13, according to the METAVIR classification). Three serum samples of each fibrotic stage were analyzed by two-dimension difference gel electrophoresis (2D-DIGE). The differential proteins were identified by the cooperation of MALDI-TOF/TOF and MASCOT; then western blotting and Bio-Plex Suspension Array were used to quantify the protein levels.ResultsThree prominent candidate biomarkers were identified: alpha 2 macroglobulin (A2M) is up regulated; vitamin D binding protein (VDBP) and apolipoprotein AI (ApoAI) are down regulated. The serum concentration of A2M was significantly different among normal, mild (F1/F2) and advanced fibrosis (F3/F4) (p < 0.01). The protein levels of VDBP and ApoAI were significantly higher in normal/mild fibrosis, when compared to those in advanced fibrosis (both p < 0.01).ConclusionsThis study not only reveals three putative biomarkers of liver fibrosis (A2M, VDBP and ApoAI) but also proves the differential expressions of those markers in different stages of fibrosis. We expect that combination of these novel biomarkers could be applied clinically to predict the stage of liver fibrosis without the need of liver biopsy.

α1-acid glycoprotein and α1-antitrypsin as early markers of treatment response in patients receiving the intensive phase of tuberculosis therapy

The identification of early markers that predict the response to anti-tuberculosis treatment would facilitate evaluation of new drugs and improve patient management. This study aimed to determine whether selected acute phase proteins and micronutrients measured at the time of diagnosis and during the first weeks of treatment could predict treatment responses during the 2-month standard intensive phase of therapy. For this purpose, alpha1-antitrypsin, alpha1-acid glycoprotein, alpha2-macroglobulin, C-reactive protein, C3, C4, zinc, copper and selenium concentrations were measured in Brazilian patients with smear-positive tuberculosis at the time of diagnosis and 1, 3, 5 and 8 weeks after initiation of therapy. Patients were classified into fast (n=29), intermediate (n=18) and slow responders (n=10) if they were smear-negative at 3, 5 or 8 weeks of treatment. alpha1-acid glycoprotein on enrolment and 1 week of treatment, alpha1-antitrypsin at week 1 and C-reactive protein and C3 after 3 weeks of therapy were higher in slow responders than in fast responders. alpha1-antitrypsin and alpha1-acid glycoprotein may be helpful in predicting treatment response at the time of initiation of therapy, and could be used as early markers to identify patients with an increased likelihood of treatment failure.

Effects of Fibrinogen and α2‐Macroglobulin and Their Apheretic Elimination on General Blood Rheology and Rheological Characteristics of Red Blood Cell Aggregates

Methods of therapeutic apheresis, such as plasma exchange or rheopheresis eliminate moderately aggregating macromolecules like fibrinogen, as well as strongly aggregating substances like alpha2-macroglobulin from blood. In order to examine the specific effect of eliminating alpha2-macroglobulin as a highly aggregating macromolecule, this study aimed to analyze the different rheological properties of: (i) moderately aggregating red blood cells (RBCs; inducible by fibrinogen); and (ii) strongly aggregating RBCs (inducible by alpha2-macroglobulin). In vitro, RBC aggregate geometry was determined in the presence of strong and moderate aggregation inducing macromolecules. In vivo, flow behavior of RBC aggregates was analyzed by intravital microscopy. Using network scanning, the number of perfused and non-perfused microvessels was determined. In vitro, the higher adhesive forces of strongly aggregating RBCs led to both a higher packing density of single RBCs within aggregates, expressed as a significantly reduced thickness of individual RBCs, and greater deformation, expressed as a significantly diminished offset between RBCs and an increased curvature of RBCs at the ends of the aggregates. In vivo rheoscopy showed that only high aggregating RBCs persisted in the precapillary bed and led to the absence of RBCs in up to 40% of nutritive capillaries. These novel findings are of importance regarding recent developments in clinical hemorheology, specifically the clinical use of hemapheretic therapies for diseases in which impaired microcirculation plays a role in either their development or progression, such as age-related macular degeneration and complications of diabetes mellitus. Our data support that procedures reducing the concentration of alpha2-macroglobulin in blood by extracorporeal elimination might provide a more efficient improvement of overall blood fluidity in microcirculatory beds.

Influence of an established acute phase response on the severity of experimental nephritis.

Small doses of lipopolysaccharide (LPS) induced an acute phase response (APR), and a number of studies have also shown that this greatly enhances the severity of glomerular injury in the heterologous phase of nephrotoxic nephritis (hNTN), an experimental model of anti-glomerular basement membrane (GBM) disease. Here, we examined the influence of pre-existing subclinical infection and raised APR, assessed by plasma alpha 2-macroglobulin (alpha 2-M) concentration, on the degree of injury in this model of nephritis. Studies were initially performed to determine the normal range of alpha 2-M in rats and its modulation by IL-6 and different doses of LPS. Plasma concentration of alpha 2-M was found to be variable and dependent on the weight of the rats. Single injections of either LPS or IL-6 had a comparable effect, and continuous perfusions of LPS caused a progressive increase in alpha 2-M which peaked at 48 h and declined gradually over 1 week. Following induction of nephritis with 10 mg of anti-GBM antibody, rats with raised alpha 2-M had 14 +/- 3 mg/24 h albuminuria compared with 4 +/- 1 mg/24 h in rats with normal alpha 2-M (P < 0.001, Wilcoxon). Injection of 20 mg anti-GBM antibody caused 36 +/- 11 mg/24 h albuminuria compared with 16 +/- 4 mg/24 h (P < 0.001), respectively. However, all these rats remained active and none of them died. In contrast, injection of 0.25 microgram LPS before induction of nephritis with 10 mg anti-GBM antibody, in rats with raised alpha 2-M, caused severe albuminuria (115 +/- 23 mg/24 h) compared with rats having normal levels of alpha 2-M (72 +/- 15 mg/24 h, P < 0.05). Furthermore, rats with raised alpha 2-M also had severe systemic manifestations characterized by pulmonary haemorrhage and extensive glomerular thrombosis, and many of them died. These results demonstrate the potential effect of pre-existing subclinical infection and raised APR on severity of glomerular injury which may affect the outcome of experimental studies.

Isovolämische Hämodilution bei peripherer arterieller Verschlußkrankheit im Stadium IIb: Prospektiver randomisierter Doppelblindvergleich von mittelmolekularer Hydroxyäthylstärke mit Elektrolytlösung

Blood-letting to a haematocrit of 0.40 to 0.42 was performed over a period of six weeks on 30 randomized patients with peripheral vascular disease (intermittent claudication), stage IIb (after Fontaine), in addition to standardized gymnastics and physical exercise (walking). Subsequently, 15 randomly selected patients received after each blood-letting middle-molecular hydroxyethyl starch solution, another 15 randomized patients received isotonic sodium chloride solution, both groups under a double-blind protocol, gymnastics and walking exercise being continued. At the beginning of the study both groups were comparable with respect to clinical and vascular parameters and results of laboratory tests. Those of the former group had a significantly greater increase in pain-free walking distance than the latter. Blood pressure levels remained unchanged in the former, but significantly rose in the latter. Peak flow during reactive hyperaemia increased significantly in both legs in the first group, remaining unchanged in the second. An increase in spontaneous platelet aggregation occurred only in the second group; alpha 2-macroglobulin concentration rose only in patients of the first group, while fibrinogen decreased significantly in both.

Edema de la mucosa bronquial y rapidez de instauración de la crisis de asma

The plasma protein leakage produced in the bronchial mucose of patients with acute asthma might be associated to the quickness of the onset exacerbation. The aim of this study was to study this association. 22 patients with acute asthma were recruited, and the magnitude of plasma protein leakage was measured by the concentrations of albumin and alpha2-macroglobulin in sputum. Days of onset in acute asthma correlated negatively with albumin sputum concentration (r = -0.563; p = 0.006), alpha2-macroglobulin sputum concentration (r = -0.603; p = 0.003), and related relative coefficients of albumin sputum/serum (r = -0.538; p = 0.01) and alpha2-macroglobulin sputum/serum (r = -0.514; p = 0.014). When the sample was divided by the following daily cutoff: < or = 4, patients suffering from a shorter onset of acute asthma showed a higher concentration of alpha2-macroglobulin in sputum: mean (standard deviation) of 14.4 (18) versus 5.3 (5.4) (p = 0.028). Plasma protein leakage seems to play an important role in the inflammatory pathogenesis of asthma exacerbation. The quicker onset of asthma the more plasma protein extravasation to the bronchial lumen.

Bronchial Exudate of Serum Proteins During Asthma Attack

Although altered vascular permeability and edema of the bronchial mucosa are associated with asthma attack, their influence on its severity remains unknown. We address this issue by comparing relative indices for the concentration of albumin (RIAlb) and alpha2-macroglobulin (RIalpha2M) in induced sputum and peripheral blood from patients with exacerbated asthma, patients with stable asthma, and control subjects. Forty-six volunteers participated in the study: 14 with exacerbated asthma (forced expiratory volume in the first second [FEV1] 74.3% [SD, 20.8%] of reference), 23 with stable asthma (FEV1 93.6% [7.5%]), and 9 controls (FEV1 101.1% [9.9%]). The concentrations of albumin and alpha2-macroglobulin were quantified by immunoturbidimetry and immunonephelometry, respectively. The relative index was then calculated by dividing the concentration in sputum supernatant by the concentration in peripheral blood. The mean RIAlb was 1.2 (1.1) in the control group, 2.9 (3.1) in the stable asthma group, and 6.0 (6.7) in the exacerbated asthma group. The RIalpha2M values were 11.7 (10.9), 11.9 (14.7), and 3.2 (3.8) for the control group and stable and exacerbated asthma groups, respectively. The increases in the RIAlb values between all groups, and the decrease in the RIalpha2M value between the exacerbated asthma and control groups were statistically significant (P<.05). The percentage of neutrophils, but not of eosinophils, in sputum was correlated with the RIAlb (r=0.39; P=.008) but not the RIalpha2M (r=-0.035; P=.82). FEV1 displayed an inverse relationship with the RIAlb (r=-0.43; P=.009) but not with the RIalpha2M (r=-0.206; P=.24). No correlation was found between oxyhemoglobin saturation and either the RIAlb (r=-0.33; P=.19) or the RIalpha2M (r=-0.12; P=.84). Vascular permeability is altered during asthma exacerbations and appears to be correlated with the presence of neutrophils and the degree of bronchial obstruction.

C-reactive protein and alpha2-macroglobulin plasma activity in medium-severe and severe psoriasis.

An acute phase of psoriasis can be induced by cytokines involved in psoriatic pathogenic phenomena. Activation of the acute phase reaction by proinflammatory cytokines (including interleukins 1 and 6, tumour necrosis factor alpha) may account for systemic symptoms in severe psoriasis, especially in pustular and arthropathic forms of this disease. To study the activity of selected acute phase proteins in psoriatics before and after effective treatment. Plasma concentrations of C-reactive protein (CRP) and alpha2-macroglobulin (alpha2-MG) were examined in 175 males with medium-severe and severe psoriasis in the active stage, before treatment and in remission achieved with various treatments (local, acitretin, psoralen + ultraviolet A, retinoid + psoralen + UVA [Re-PUVA] methotrexate, cyclosporin A). Clinical activity of psoriasis was evaluated using the Psoriasis Area and Severity Index score, ranging from 18 to 70.8 (mean 29.2). Measurements were done using the immunoenzymatic method. The control group consisted of 30 healthy male volunteers. In the active stage of disease highly increased plasma levels of both CRP and alpha2-MG were found (P < 0.001). It appeared that efficacious treatment was correlated with a considerable decrease of the proteins examined towards the control values. Mean levels of alpha2-MG after treatment did not differ significantly from those of controls (P > 0.15). However, despite their strong decrease following all methods of treatment, mean plasma levels of CRP remained significantly elevated during remission compared with those of the healthy controls (P < 0.001). The results of this study indicate that in medium-severe and severe psoriasis the acute phase response can be initiated and is not completely extinguished in phases of clinical remission.

Circulating alpha-2-macroglobulin levels and depression scores in patients who underwent abdominal cancer surgery

Background Although depression is a predominant health care problem, it is difficult to diagnose in patients who underwent abdominal cancer surgery. Relevance of elevated levels of serum of alpha-2-macroglobulin (A2M) to psychological depression has been recently suggested. Patients and methods Serum A2M levels were measured and depression scores were determined by the Hospital Anxiety and Depression Scale before and 1 year after abdominal cancer surgery in 45 patients. Results Serum levels of A2M and depression scores were significantly ( P = 0.0006, P = 0.0045, respectively) increased after total gastrectomy compared with preoperative values, whereas there was no interval change in both measurements of patients having colorectal resection. Of the patients who had undergone distal or total gastrectomy for gastric cancer, serum A2M concentrations in a depressed group were significantly higher ( P = 0.028) than those in non-depressed group. Conclusions Patients with total gastrectomy are predisposed to depression, and circulating A2M elevation may be implicated in the development of postoperative depression in these patients.

Urinary plasma protein patterns in acute prostatitis.

We evaluated the diagnostic utility of urinary alpha1-microglobulin, alpha2-macroglobulin and albumin in the diagnosis of acute prostatitis. We studied 133 men (43 +/- 17 years) with, and a reference population (n=36, 41 +/- 16 years) without, urinary tract infection. Prostatectomy samples were used to study the potential interference between prostatic proteins and protein analysis. Urinary alpha2-macroglobulin/albumin ratio was significantly lower in prostatitis compared to the reference population, cystitis or acute pyelonephritis (p < 0.0001). Low alpha2-macroglobulin concentrations in prostatitis are due to inhibition (p = 0.0001) of the immune reaction between alpha2-macroglobulin in presence of polyclonal rabbit antibodies (used for immunonephelometry) by soluble prostatic proteins (+/- 60 kDa) which appear in urine in acute prostatitis. The urinary alpha1-microglobulin/creatinine ratio diagnoses acute pyelonephritis (sensitivity 100% and specificity 87%) and the urinary alpha2-macroglobulin/albumin ratio diagnoses acute prostatitis (sensitivity 100% and specificity of 90%). Stepwise multinomial logistic regression analysis reveals that urinary alpha1-microglobulin, alpha2-macroglobulin, albumin and creatinine provide optimal differentiation between acute pyelonephritis and acute prostatitis (pseudo R2=0.83; Loglikelihood -30.55, p < 0.000001). In conclusion, the combination of hematuria and absence of urinary alpha-2-macroglobulin is diagnostic for acute prostatitis. Even without hematuria, alpha2-macroglobulin remains lower compared to patients without prostatitis.

Ozone exposure and the production of reactive oxygen species by bronchoalveolar cells in humans.

Exposure to ozone injures respiratory epithelium, and the mechanisms may involve the generation of reactive oxygen species (ROS). This study tested the hypothesis that ozone exposure increases the airway burden of ROS to a greater degree in smokers than nonsmokers, and that this effect is independent of ozone-induced changes in spirometry. Healthy subjects were selected as either responders (decrement in FEV1 > 15%) or nonresponders (decrement in FEV1 < 5%) to ozone; each underwent 2 exposures to ozone and 1 to air, with bronchoalveolar lavage (BAL) performed 30 min (early) and 18 h (late) after exposure. Release of superoxide anion (O2(-)) was used as a measure of ROS release by all BAL cells, and flow cytometry was used to detect ROS production in alveolar macrophages (AM) only. Recovery of AM was approximately threefold greater in smokers than nonsmokers. Unstimulated, but not stimulated, cells obtained by BAL from smokers released approximately twofold greater amounts of O2(-) than cells from nonsmokers, both early and late after ozone exposure (p =.012 and p =.046, respectively). Stimulated, but not unstimulated, ROS generation by AM from smokers increased following ozone exposure, but the ozone effect was not significant. ROS production by AM decreased in nonsmokers (air vs. ozone late, p =.014). Total protein, albumin, and immunoglobulin M (IgM) increased in BAL fluid, consistent with an increase in epithelial permeability. In addition, the concentration of alpha2-macroglobulin increased approximately threefold 18 h after exposure in nonsmokers (p <.001). No relationship was found between measures of ROS production and lung function responsiveness to ozone. These studies suggest the airways of smokers experience a greater burden of ROS than those of nonsmokers following ozone exposure.

The concentration of protein-compounds in interstitial tissue of patients with chronic critical limb ischaemia and oedema.

Many of chronic critical limb ischaemia (CLI) patients have distal leg and foot oedema. Previous electronmicroscopic studies have shown that chronic severe ischaemia may cause hypoxic damage of the capillary endothelium, including morphological changes i.e. multiplicated/thickened basal lamina, and formation of interendothelial gaps. To assess the functional consequences of these morphologic derangements, where proteins can leak through, we investigated the composition of the interstitial fluid in oedematous ischaemic limbs. Nine female and 3 male patients with a mean age of 79 +/- 7.9 years were included. All had unilateral CLI and peripheral pitting oedema. Leg and foot volume was measured with water displacement volumetry. Blister suction technique was used to collect subcutaneous interstitial fluid. The concentration of albumin, transferrin, immunoglobulin G and alpha 2-macroglobulin in plasma and blister fluid was measured by immunoturbidimetry. Nine patients, 8 women and 1 man with a mean age of 83 +/- 5.5 years with a proximal femur fracture served as an age-matched control group. The mean concentration of albumin in blister fluid was significantly lower in the patients, whereas the mean concentration of alpha 2-macroglobulin in blister fluid did not differ between patients and controls. Mean ratio between concentrations in blister and serum of albumin, transferrin and immunoglobulin G in the limbs with CLI and oedema were significantly lower than respective values in the control group. However, there was no significant difference in the ratio of alpha 2-macroglobulin between these groups. A higher transcapillary concentration gradient for proteins in CLI limbs signifies an increase in the net osmotic pressure gradient across the capillary wall, which may be a potential oedema limiting factor.

Protein sgp 120 as a marker of an acquired angioedema.

Angioedema is not an event peculiar to an acute-phase response. We registered the appearance of angioedema in four nonatopic patients suffering from diverse inflammatory stimuli. Serum concentrations of C-reactive protein (CRP), haptoglobin, alpha1-antitrypsin, alpha2-macroglobulin, C3, C4, C1INH and protein sgp 120 (classified as a regulatory protein of the complement system) were assessed over 7 consecutive days, beginning with the onset of angioedema in these four patients. Instead of returning to the normal level, CRP peaked again on the 4th day (A-test, P<0.05) and declined significantly by the 7th day (A-test, P<0.05). Serum levels of sgp 120, C3c and C1INH increased gradually up to the 5th day but only sgp 120 was significantly raised in relation to the initial value (P< 0.05). In our patients, we registered four diverse profiles of the acute-phase proteins (Kruskal-Wallis test, P<0.05) probably due to the diverse nature of the pre-existing inflammatory stimuli. According to these results, the unexpected appearance of angioedema implies the contribution of sgp 120 as the main source of fragments with kinin-like properties.

Hepatocyte growth factor modulates the hepatic acute-phase response in thermally injured rats.

Hepatocyte growth factor (HGF) has been shown to modulate the acute-phase response in vitro. The specific in vivo role of HGF in this multifactorial response, however, remains unknown. This study examines the effects of exogenous HGF on the acute-phase response in thermally injured rats. Prospective, randomized, laboratory study. Shriners Hospital for Children and University of Texas Medical Branch laboratories. Fifty-six male Sprague-Dawley rats (weight range, 300-325 g). Animals received a 60% total body surface area third-degree scald burn and were randomly divided to receive either 400 microg/kg/day i.v. HGF or saline (control). Serum acute-phase proteins, cytokines, and insulin-like growth factor (IGF)-I concentrations, as well as liver weight, protein and triglyceride content, IGF-I concentrations, and cytokine gene expression were measured 1, 2, 5, or 7 days after burn. Serum albumin was increased on days 2, 5, and 7 after burn, and transferrin was increased on day 7 after burn in HGF-treated rats compared with controls (p<.05). HGF increased alpha2-macroglobulin concentrations on postburn days 2, 5, and 7 compared with controls (p<.05). Serum interleukin-6 and tumor necrosis factor-alpha were significantly higher within 2 days of burn in rats treated with HGF (p<.05). HGF increased the hepatic gene expression of tumor necrosis factor-alpha compared with controls (p<.05). Serum IGF-I decreased in rats receiving HGF 1, 2, and 5 days after burn, whereas liver IGF-I concentrations were higher on days 1 and 7 after burn compared with controls (p<.05). Hepatic protein concentrations were higher in the HGF group compared with controls on postburn days 1, 2, and 7, with a concomitant increase in total liver weight (p<.05). HGF exerted a strong mitogenic effect on hepatocytes 1 and 2 days after thermal injury compared with controls (p<.05). These findings suggest that HGF modulates the acute-phase response in vivo after burn and causes changes in liver morphology.