PA Antigen Research Articles

EVALUATION OF THREE COVID-19 ANTIGEN ASSAY VERSUS PCR DETECTION IN ROUTINE PRACTICE

According to WHO, molecular testing is the "Gold Standard" for the diagnosis of SARS-CoV-2 infection. However, these tests have some limitations in practice (well trained staff, specific equipment requirements, organization of area and time frame for reporting results). Thus, rapid antigen test (RAT) for the detection of one or several SARS-CoV-2 antigens have emerged. We evaluated the analytical performance of 3 RAT used in our laboratory : FREND® Ag COVID-19 (NanoEntek) (FA), STANDARD Q COVID-19 Ag (SD Biosensor) (BA), PANBIO™ COVID-19 Ag RAPID TEST DEVICE (Abbott) (PA) in comparaison to rRT-PCR results. Our study indicates that PA and SA antigen tests have a very good sensitivity to identify infected patients with COVID-19 specifically between in the 0-5 days time-window post onset of symptoms. Nevertheless, one out of three antigen tests (FA) showed very poor clinical performance.

A serological survey of Bacillus anthracis reveals widespread exposure to the pathogen in free-range and captive lions in Zimbabwe.

Numerous unknown factors influence anthrax epidemiology in multi-host systems, especially at wildlife/livestock/human interfaces. Serology tests for anti-anthrax antibodies in carnivores are useful tools in identifying the presence or absence of Bacillus anthracis in a range. These were employed to ascertain whether the disease pattern followed the recognized high- and low-risk anthrax zonation in Zimbabwe and also to establish whether anthrax was absent from Hwange National Park in which there have been no reported outbreaks. African lions (Panthera leo) (n=114) drawn from free-range protected areas and captive game parks located in recognized high- and low-risk zones across Zimbabwe were tested for antibodies to anthrax PA antigen using the ELISA immunoassay. A random selection of 27 lion sera samples comprising 17 seropositive and 10 seronegative sera was further tested in the species-independent toxin neutralization assay (TNA) in order to validate the former as a surveillance tool for anthrax in African lions. Using the ELISA-PA immunoassay, 21.9% (25/114) of the lions tested positive for antibodies to anthrax. Seropositivity was recorded in all study areas, and there was no significant difference (p=.852) in seropositivity between lions in high- and low-risk anthrax zones. Also, there was no significant difference (McNemar's chi-square test=0.9, p=.343) in the proportion of lions testing positive to anti-PA anthrax antibodies on ELISA-PA immunoassay compared with the TNA, with fair agreement between the two tests [kappa (K) statistic=0.30; 0.08<K<0.613]. Results of this study indicate that anthrax could be more widespread than 42 currently realized in Zimbabwe, and present in recognized high- and low-risk zones, including 43 where it has not been reported in over 20years such as Hwange National Park. This is also the 44 first report documenting the presence of anthrax lethal toxin-neutralizing antibodies in naturally 45 infected carnivores, further confirming exposure to B. anthracis. The research results point to a 46 need for revisiting the currently recognized anthrax risk zones in Zimbabwe. This should be based 47 on improved surveillance of the disease in both wild and domestic animals for better understanding and control of the disease.

In situgastrointestinal protection against anthrax edema toxin by single-chain antibody fragment producing lactobacilli

BackgroundAnthrax is caused by the bacterium Bacillus anthracis and is regarded as one of the most prominent bioterrorism threats. Anthrax toxicity is induced by the tripartite toxin complex, composed of the receptor-binding anthrax protective antigen and the two enzymatic subunits, lethal factor and edema factor. Recombinant lactobacilli have previously been used to deliver antibody fragments directed against surface epitopes of a variety of pathogens, including Streptococcus mutans, Porphyromonas gingivalis, and rotavirus. Here, we addressed whether or not anthrax toxins could be targeted and neutralised in the gastrointestinal tract by lactobacilli producing recombinant antibody fragments as a model system for toxin neutralisation in the gastrointestinal lumen.ResultsThe neutralising anti-PA scFv, 1H, was expressed in L. paracasei as a secreted protein, a cell wall-anchored protein or both secreted and wall-anchored protein. Cell wall display on lactobacilli and PA binding of the anchored constructs was confirmed by flow cytometry analysis. Binding of secreted or attached scFv produced by lactobacilli to PA were verified by ELISA. Both construct were able to protect macrophages in an in vitro cytotoxicity assay. Finally, lactobacilli producing the cell wall attached scFv were able to neutralise the activity of anthrax edema toxin in the GI tract of mice, in vivo.ConclusionWe have developed lactobacilli expressing a neutralising scFv fragment against the PA antigen of the anthrax toxin, which can provide protection against anthrax toxins both in vitro and in vivo. Utilising engineered lactobacilli therapeutically for neutralising toxins in the gastrointestinal tract can potential be expanded to provide protection against a range of additional gastrointestinal pathogens. The ability of lactobacilli to colonise the gastrointestinal tract may allow the system to be used both prophylactically and therapeutically.

Decreased Peritoneal Tissue Plasminogen Activator During Prolonged Laparoscopic Surgery

Peritoneal fibrinolysis is crucial in the peritoneal healing processes and subsequent adhesion formation. During conventional surgery, the peritoneal fibrinolytic system is rapidly disturbed. Short-term laparoscopy does not seem to affect peritoneal fibrinolysis. The aim of the present study was to assess the effect of prolonged laparoscopic surgery on peritoneal fibrinolysis. Twelve consecutive patients undergoing laparoscopic gastric bypass surgery for morbid obesity were included in the study. During the procedure, biopsies of the parietal peritoneum were taken at the start of the procedure and each 45 min afterward. Tissue samples were homogenized and tissue-type plasminogen activator (tPA) antigen, tPA activity, urokinase-type PA antigen, and plasminogen activating inhibitors type 1 antigen were measured using commercial assay techniques. Both tPA antigen and its activity progressively decreased during the procedure, reaching significant levels after 90 min of surgery. The levels of uPA antigen and plasminogen activating inhibitors antigen did not significantly change throughout the procedure. As for conventional surgery, prolonged laparoscopic surgery causes a decreased fibrinolytic activity in the peritoneum due to decreased tPA levels.

EMILINs interact with anthrax protective antigen and inhibit toxin action in vitro

The informational spectrum method (ISM) is a virtual spectroscopy method for the fast analysis of potential protein–protein relationships. By applying the ISM approach to the GeneBank protein database the vascular proteins EMILIN1 ( Elastin Microfibril Interface Located Prote IN), EMILIN2, MMN1, and MMN2 were identified as additional anthrax PA antigen interacting molecules. This virtual molecular interaction was formally proven by solid phase assays using recombinant proteins. The interaction is independent of the presence of divalent cations and does not involve PA aspartic residue at 683, a critical residue in receptor binding. In fact, the D683A point mutation fully prevented the cell intoxication ability of PA in the presence of Lethal Factor, but it was fully ineffective on the binding of mutated PA to EMILIN1 and EMILIN2. The ISM approach also led to the identification of the potential interaction sites between PA and EMILINs. A PA mutant with a deletion at residue D425 and solid phase protein–protein interaction studies as well as deletion mutant of EMILIN2 confirmed the hypothesized interaction site. Our findings imply that the PA-cell surface receptor interaction is not likely to provide the full explanation for the vascular lesions and prominent hemorrhages that follow Bacillus anthracis infection and spreading and call into play vascular associated proteins such as EMILINs as potential inhibitory proteins.

Western blot antibody determination in sera from patients diagnosed with Anisakis sensitization with different antigenic fractions of Anisakis simplex purified by affinity chromatography

Using Western blot techniques, the specificities of crude and purified (PAK and PAS) Anisakis simplex antigens were compared against 24 sera from patients diagnosed with Anisakis sensitization. All patients recognized a 60 kDa protein against the A. simplex crude extract, while 37.5% and 12.5% reacted with proteins of 40 and 25 kDa, respectively, when IgG was tested. In the case of IgE determination, 41.6% of sera were negative, while 12.5% and 20.8% appeared to cross-react against Toxocara canis and Ascaris suum, respectively. When the PAK antigen (A. simplex antigen purified by means of a column of IgG anti-A. simplex) was tested, immune recognition towards the 60, 40 and 25 kDa proteins increased in 83.3%, 16.7% and 4.2%, respectively, when the Ig antibodies were tested. In the case of the PAS antigen (PAK antigen purified by means of a column of IgG anti-A. suum), the reaction against the 40 and 25 kDa proteins increased to 45.8% and 25%, respectively, when Ig antibodies were used. Finally, when the EAS antigen (eluted from the anti-A. suum column after PAK purification) was tested, 83.3% of the assayed sera reacted against the 14 kDa protein, when the Ig antibodies, IgG and IgM immunoglobulins were measured. With the IgE determination, the reactions were observed in 41.7% of patients with proteins between 60 and 35 kDa against the PAS antigen. With the EAS antigen, reactive bands of 184, 84 and 14 kDa appeared. In conclusion, in the purification process of the A. simplex larval crude extract, the proteins implicated in cross-reactions with Ascaris and Toxocara were eliminated, with an important concentration of proteins responsible for the induction of specific responses.

Detection and Quantification of Pharaoh Ant Antigens in Household Dust Samples as Newly Identified Aeroallergens

Background:The widespread house ant, Monomorium pharaonis (pharaoh ant, PA), was recently identified as a potential cause of respiratory allergies. However, there are no reports of the distribution of PA allergens in various environments. We developed specific ELISA inhibition assays and measured the distribution and amount of PA antigens in household dust samples. Methods:Floor dust was collected at 3-month intervals from 56 homes in Seoul over a 1-year period. PA antigens in fine dusts were quantified by ELISA inhibition assays using rabbit anti-PA sera, and specific IgE to PA antigens in residents’ serum was measured by ELISA. Results: In 18 of the 56 homes (32.1%), PA antigen was detected in at least 1 floor dust sample either from the living room or the kitchen. Levels of PA antigens showed seasonal variations with peaks in autumn and winter. The detection rate of PA antigens was significantly higher in homes with visual evidence of PA infestations (70%) than in homes without such infestations (23.9%; p < 0.05). However, a significant amount of PA antigens was still detected in uninfested homes. Thirteen of 113 (11.5%) residents were positive for PA-specific IgE. PA-specific IgE was detected more frequently in residents living in PA antigen-positive homes (19.6%) than in antigen-negative homes (4.8%; p < 0.05). Conclusion: A considerable level of PA antigens is distributed in the indoor environment. Therefore, inhalant exposure to PA antigens can occur during domestic activities. These results suggest that PAs might be a significant source of aeroallergens in households.

Expression of furin-linked Fab fragments against anthrax toxin in a single mammalian expression vector

Human anti-recombinant protective antigen (rPA) Fab genes were previously cloned from single B cells of a donor immunized with anthrax vaccine using fluorescence activated cell sorting with fluorescein labeled rPA and single-cell PCR. The light and heavy chains were sub-cloned individually into mammalian expression vectors pSecTag2B or pEXPR44, respectively, and expressed in the same CHOK1 cells. Alternatively, the same heavy and light chains were linked together, using PCR, with an in-frame sequence coding for a furin cleavage site. This construct was cloned into pSecTag2B and expressed in CHOK1 cells. Once expressed, the individual chains combined in vivo to form a Fab fragment which was purified as a single protein when either method was utilized. The human Fab antibodies produced by this technique were functional when tested in Western blots using the recombinant PA antigen as the target.

ELISA antibody determination in patients with anisakiosis or toxocariosis using affinity chromatography purified antigen

One of the fundamental aspects of a parasitic infection diagnosis is the use of adequate antigens to develop specific and sensitive immunoassays. This fact is especially complicated in nematode infection cases because of the high cross-reactivity among different parasites in this group. We performed an evaluation of Anisakis simplex antigens purified by affinity chromatography. We used sera from 38 patients diagnosed with Anisakis sensitization and sera from 35 patients with clinical suspicion of visceral larva migrans (VLM). These sera were assayed by the ELISA method against the crude extracts (CEs) and the purified antigens. When the sera from patients diagnosed with Anisakis sensitization were tested against the A. simplex CE, the IgG was the most abundant immunoglobulin. When the A. simplex larval antigens were purified using a column of IgG anti-A. simplex (PAK) or a column of IgG anti-Ascaris suum (PAS) were tested, we observed a higher diminution in the IgG levels, which coincides with the augmentation of the mean values against the "eluted of Ascaris" (EAS antigen). When the IgE was detected, only 18.4% of the sera reacted with the PAS antigen. We have observed that in the purification process of A. simplex antigen by affinity chromatography, the majority of the proteins that produced cross-reactivity against A. suum and Toxocara canis were eliminated.

Advances in alfalfa mosaic virus-mediated expression of anthrax antigen in planta

Plant viruses show great potential for production of pharmaceuticals in plants. Such viruses can harbor a small antigenic peptide(s) as a part of their coat proteins (CP) and elicit an antigen-specific immune response. Here, we report the high yield and consistency in production of recombinant alfalfa mosaic virus (AlMV) particles for specific presentation of the small loop 15 amino acid epitope from domain-4 of the Bacillus anthracis protective antigen (PA-D4s). The epitope was inserted immediately after the first 25 N-terminal amino acids of AlMV CP to retain genome activation and binding of CP to viral RNAs. Recombinant AlMV particles were efficiently produced in tobacco, easily purified for immunological analysis, and exhibited extended stability and systemic proliferation in planta. Intraperitional injections of mice with recombinant plant virus particles harboring the PA-D4s epitope elicited a distinct immune response. Western blotting and ELISA analysis showed that sera from immunized mice recognized both native PA antigen and the AlMV CP.

Coagulation and Fibrinolysis in Preeclampsia and Neonates

Coagulation and fibrinolysis were determined in 67 Indonesian women admitted to the University Hospitals for delivery in Medan. They were diagnosed to be at term gestation (mean 39.3 +/- 1.1 weeks) with moderate and severe preeclampsia (n=32) and in labor, and 8 had preterm labor (gestation mean 33.5 +/- 2.6 weeks). Twenty-seven normal pregnant women in labor (gestation mean 39.7 +/- 1.0 weeks) served as controls. Cord blood from 23 neonates from normal pregnancy and 31 neonates from preeclampsia was also evaluated. Preeclamptic women in labor showed further enhanced coagulation activation (F(1+2)) with raised urokinase-like plasminogen activator (u-PA) activity and reduced plasminogen activator inhibitor-2 (PAI-2) levels. In preterm preeclampsia, significantly reduced antithrombin III (ATIII) and PAI-2 levels with further elevated tissue-type PA (t-PA) antigen and plasminogen activator inhibitor-1 (PAI-1) antigen were seen compared to normal pregnancy. These would suggest a state of enhanced thrombin generation with elevated fibrinolytic/inhibitor proteins in preterm preeclampsia. The reduced PAI-2 levels seen in preeclampsia have been suggested to be associated with reduced placental function. Neonates born to mothers of either normal pregnancy or preeclampsia at term showed similar hemostatic changes with reduced fibrinogen, ATIII, t-PA, u-PA antigen, PAI-1 levels, and coagulation activation compared to their respective maternal plasma levels. No significant differences in hemostatic parameters studied between the neonates of both cohorts were seen, and this would suggest that the neonates were protected from the adverse effects of preeclampsia and their hemostatic system was physiologically balanced.

Enzyme-linked immunosorbent assay and Western blot antibody determination in sera from patients diagnosed with different helminthic infections with Anisakis simplex antigen purified by affinity chromatography

An evaluation of the sensitivity and the specificity of the Anisakis simplex antigens purified by affinity chromatography was performed using sera from patients diagnosed with Anisakis sensitisation and sera from patients previously diagnosed with different helminthic infections. Only the sera of the patients diagnosed with Schistosoma mansoni or Onchocerca volvulus parasitic infections were negative against the A. simplex antigen and its purified fractions (PAK antigen: A. simplex antigen purified using columns prepared with anti-A. simplex rabbit IgG and PAS antigen: PAK antigen purified using columns prepared with anti-Ascaris suum rabbit IgG). However all the sera were positive against the A. suum antigen. In all the sera from the patients diagnosed with Anisakis sensitisation, the antibody levels detected using the purified antigens (PAK and PAS antigens) were lower than the observed using the A. simplex crude extract with the highest diminution in the case of the IgG. When these same sera were tested against the A. simplex crude extract by Western blot, several bands of high molecular masses were observed as well as, intense bands at 60 and/or 40 kDa. A concentration of these last proteins was observed in the PAK and the PAS antigens. When the sensitivity and the specificity determinations were performed, only seven of the 38 patients diagnosed of Anisakis sensitisation were positive, as well as, the sera from the patients diagnosed with parasitisms by Echinococcus granulosus or Fasciola hepatica.

Increasing expression of tissue plasminogen activator and plasminogen activator inhibitor type 2 in dog gingival tissues with progressive inflammation

Urokinase and tissue-type plasminogen activators (u–PA and t–PA) are serine proteases that convert plasminogen into plasmin, which degrades matrix proteins and activates metalloproteinases. The PAs are balanced by specific inhibitors (PAI–1 and PAI–2). Local production of t–PA and PAI–2 was recently demonstrated in human gingival tissues. The aim now was to investigate the production and localization of t–PA and PAI–2 in gingival tissues from dogs in three well-defined periodontal conditions; clinically healthy gingiva, chronic gingivitis and an initial stage of ligature-induced loss of attachment. At the start of the experiment the gingiva showed clear signs of inflammation. Clinically healthy gingiva were obtained after 21 days period of intense oral hygiene. Attachment loss was induced by placing rubber ligatures around the neck of some teeth. Biopsies were taken from areas representing the different conditions and prepared for in situ hybridization and immunohistochemistry. In clinically healthy gingiva both t–PA mRNA and antigen were expressed in a thin outer layer of the sulcular and junctional epithelia. No t–PA signals or staining were seen in connective tissue. Both mRNA signaling and immunostaining for t–PA were stronger in chronic gingivitis. In areas with loss of attachment, t–PA mRNA as well as antigen were found in the sulcular and junctional epithelia to a similar degree as in gingivitis. Occasionally the connective tissue was involved, especially in connection with vessels. PAI–2 mRNA was seen in a thin outer layer of the sulcular and junctional epithelia in clinically healthy gingiva, but no signals were seen in connective tissue. PAI–2 antigen was found primarily in the outer layer of the sulcular and junctional epithelia. Some cells in the connective tissue were stained. In gingivitis, PAI–2 signals were mainly found in the same locations, but more intense and extending towards the connective tissue. Immunostaining was seen in the outer half of the sulcular and junctional epithelia as well as in the upper part of the connective tissue, close to the sulcular epithelium. In sites with loss of attachment, PAI–2 mRNA was found throughout the sulcular and junctional epithelia, as was the antigen, which stained intensely. No PAI–2 mRNA was seen in connective tissue; the antigen was found scattered, especially near vessels. This study shows that the expression of both t–PA and PAI–2 increases with experimental gingival inflammation in the dog, and furthermore, the two techniques demonstrate a strong correlation between the topographical distribution of the site of protein synthesis and the tissue location of the antigens for both t–PA and PAI–2. The distribution correlates well with previous findings in humans.

Influence of PAI-1 on adipose tissue growth and metabolic parameters in a murine model of diet-induced obesity.

An increased plasma plasminogen activator inhibitor-1 (PAI-1) level is a risk factor for myocardial infarction, particularly when associated with visceral obesity. Although the link between PAI-1 and obesity is well documented, little is known about the physiological relevance of PAI-1 production by adipose tissue. Therefore, we have compared adipose tissue development and insulin resistance plasma parameters in PAI-1-deficient mice (PAI-1(-/-)) and wild-type littermates (PAI-1(+/+)) in a model of nutritionally induced obesity. After 17 weeks of consuming a high-fat diet (HFD), PAI-1(+/+) mice showed marked obesity, with a 52% increase in body weight compared with mice that were kept on a standard fat diet (P<0.0001). This weight gain was accompanied by adipocyte hypertrophy and an increase in the number of stroma cells in the gonadal fat pad, expressed as stroma cells/adipocytes (0.67+/-0.05 versus 0.43+/-0. 02; P<0.001). In plasma, the HFD induced a marked increase in PAI-1 antigen (5.1+/-0.56 versus 2+/-0.22 ng/mL; P<0.001), fasting insulinemia (1.1+/-0.21 versus 0.21+/-0.04 ng/mL; P<0.001), and glycemia (7.4+/-0.5 versus 5+/-0.3 mmol/L; P<0.001), whereas plasma triglyceride levels were not affected. When we compared PAI-1(-/-) and PAI-1(+/+) mice on the HFD, PAI-1(-/-) mice gained weight faster than did PAI-1(+/+) mice, with a significant difference in body weight between 3 and 8 weeks of the diet (32+/-1.7 versus 26+/-1.6 g at 6 weeks; P<0.05). After 17 weeks of the HFD, its effect on weight gain and the number and size of adipocytes was similar in PAI-1(+/+) and PAI-1(-/-) mice. By contrast, the increase in the number of stroma cells presented by PAI-1(+/+) mice was not observed in PAI-1(-/-) mice. In obese PAI-1(-/-) mice, tissue-type PA activity and antigen levels in the gonadal fat pad were significantly higher than in obese PAI-1(+/+) mice (230+/-50 versus 47+/-20 arbitrary units/g, P<0.01; 40+/-13 versus 17+/-13 ng/g, P<0.05, respectively), whereas urokinase-type PA activity and antigen levels were similar in both groups. In plasma, nonobese PAI-1(-/-) mice displayed 62% higher insulin levels (P<0.05) than did PAI-1(+/+) mice. Obese PAI-1(-/-) mice displayed 68% higher triglyceride levels (P<0.01) and 21% lower glucose levels (P<0.05) than did PAI-1(+/+) mice. These data support an effect of PAI-1 on weight gain and adipose tissue cellularity in the induction of obesity in mice. Moreover, PAI-1 influences glucidolipidic metabolism. The elevated expression of PAI-1 observed in human obesity could be involved in mechanisms that control adipose tissue development.

Establishment of a novel B-lymphoma cell line, CTB-1, with strong Pas antigen expression having chromosomal translocation (14;22)

We established a new lymphoma cell line, designated CTB-1, from pericardial effusion of a patient with diffuse large B-cell lymphoma. This cell line showing vigorous growth ability has undergone 260 passages over a period of 34 months in suspension culture, and is heterotransplantable to nude mice. The cultured cells were positive for CD10, CD19, CD20, CD21, HLA-DR, and surface IgG kappa, and negative for T cell antigens. Chromosomal analysis revealed a t(14;22)(q32;q11) that is consistent with original lymphoma cells. CTB-1 cells show the high cell surface expression level of Fas antigen/APO-1. However, ligation of Fas antigen with anti-Fas monoclonal antibody (clone CH-11) did not induce apoptosis of CTB-1 cells. This suggests that Fas itself or the downstream signaling pathways of Fas may be impaired in this cell line. This new cell line may provide a useful in vitro system to study the biology and pathogenesis of B-cell lymphoma which is independent of Fas-mediated apoptosis.

A Comparison of 4 Ultrasensitive Prostate Specific Antigen Assays for Early Detection of Residual Cancer after Radical Prostatectomy

The Yang Pros-Check, Abbott IMx, Tosoh AIA-PACK PA and Nichols Institute reference laboratory prostate specific antigen (PSA) assays were compared in 30 patients (138 sera) known to have no residual prostate cells. The mean + 3 standard deviations of these sera was used to define a level of PSA that would indicate residual cancer. This residual cancer detection limit for each of the 4 assays is 0.06, 0.01, 0.07 and 0.05 ng./ml., respectively, but the 0.01 level for the IMx assay is an artifact caused by setting the zero calibrator too high.All 4 assays were then used to compare the number of days from radical prostatectomy to the detection of residual cancer in 23 cases (211 sera) that ultimately failed radical prostatectomy. The Yang, Tosoh and Nichols Institute assays were all similar, with an average of 569 to 589 days. The Abbott IMx assay was relatively insensitive in detecting the first appearance of PSA after radical prostatectomy (average 821 days) and it showed the earliest detection among the 4 assays in only 1 of 23 patients.

Sex-related differences of plasminogen activator activity and plasminogen activator inhibition in the gastric wall after chronic ethanol consumption: Effect of orchidectomy or ovariectomy

Plasminogen activator activity (PAA), tissue-type PA(t-PA) antigen level, plasminogen activator inhibition (PAI), and plasmin inhibition (PI) showed no sex-dependent differences in the stomach of the rat. Chronic ethanol consumption induced an increase of PAA and t-PA antigen and a decrease of PAI (urokinase-type PA [u-PAI]) and PI in the body and pyloric region (PI only in the pyloric region). PAA increase and u-PAI decrease in the pyloric region were greater in the male than in the female. Orchidectomy, but not ovariectomy, induced effects similar to those of ethanol. The effects of orchidectomy and chronic ethanol consumption on gastric PAA, t-PA antigen, and PAI were additive. An increased PAA and decreased PAI or PI in the stomach in cases of hypogonadism, particularly if this deficiency is combined with chronic ethanol consumption, might influence the susceptibility to pathological conditions related to or affected by the local fibrinolytic activity (PAA, PAI, or PI).

Stimulation of tissue type plasminogen activator by leukaemic cell homogenates.

In patients with disseminated intravascular coagulation (DIC), hyperfibrinolysis was observed in patients with leukaemia, but hypofibrinolysis was seen in those with sepsis. Although the plasma tissue plasminogen activator (t-PA) level was higher in patients with DIC than in those without DIC, there was no significant difference in t-PA level between the patients with leukaemia and sepsis. Hyperfibrinolysis might not be caused by t-PA derived from leukaemic cells, although the PA antigen level in leukaemic cell homogenates was significantly higher in patients with DIC than in those without DIC. The activation of t-PA by leukaemic cell homogenates in the absence of bromocyan fibrinogen fragments suggested that leukaemic cell homogenates had t-PA stimulator activity. The t-PA stimulator activity was high in both acute myeloblastic leukaemia (AML) and acute lymphoblastic leukaemia (ALL), especially in DIC, but this activity was not detected in chronic myelocytic leukaemia (CML) or normal cells. Since fibrinogen and soluble fibrin monomer complex levels in leukaemic cells were also high in patients with DIC, fibrinogen degradation products might be the major t-PA stimulator in leukaemic cells. This might be one of the causes of hyperfibrinolysis in leukaemia.

Influence of Heparin and a Low Molecular Weight Heparinoid on Specific Endogenous and Exogenous Fibrinolytic Factors during Rest and Exercise

The effects of heparin (5,000 IU i.v.) and the low molecular weight heparinoid Org 10172 (Orgaran) (3,250 anti-Xa units i.v.) on components of the fibrinolytic system were studied in two double-blind, randomised, placebo-controlled, cross-over trials using healthy subjects. In study A (n = 6) the effects were studied during rest and standardized exercise and in study B (n = 6) during a low dose infusion of recombinant tissue-type plasminogen activator (rt-PA; 80 micrograms over 16 min). At rest, heparin and Org 10172 did not influence the plasma concentrations of endogenous t-PA antigen and activity, urokinase-type PA (u-PA) antigen, plasmin activatable pro-urokinase (scu-PA), active urokinase (tcu-PA) and plasminogen activator inhibitor-1 (PAI-1) antigen. Recombinant t-PA antigen and activity during rt-PA infusion were also not affected. During exercise, neither heparin nor Org 10172 influenced the area under the curve (AUC) of t-PA and u-PA antigen and t-PA activity when compared with placebo. Unexpectedly, after heparin the AUC of t-PA activity was 49% larger (range +19 to +245%) than after Org 10172 (p < 0.05). The last difference was considered spurious, scu-PA, tcu-PA and PAI-1 antigen levels at 2 min after termination of exercise were unaffected by both compounds (p > 0.05). Sulphated polysaccharides do not increase fibrinolytic activity of the plasma by changing the concentrations of the components of the fibrinolytic system.

Analysis of the Expression and Immunostimulatory Capacity of Class I Major Histocompatibility Antigens on Rat Trophoblast Cell Lines

In rat strains expressing the a and other major histocompatibility complex (MHC) haplotypes, subpopulations of placental trophoblast cells synthesize the nonclassical class I Pa antigen in preference to the classical RT1.Aa antigen. In this study, a rat trophoblast cell line, R8RP.3, which was derived from midgestation placentas of PVG.R8 (RT1.Aa) rats, was shown to express class I antigens similarly to those of trophoblast cells in situ. Both unstimulated and IFN-gamma-exposed metabolically labeled R8RP.3 cells synthesized more Pa than RT1.Aa antigen. The reverse was true for labeled spleen cells from PVG.R8 rats. The R8RP.3 cells failed to stimulate allogeneic lymphocyte proliferation even when high levels of both classical and nonclassical class I MHC antigens were expressed on their membranes after incubation with IFN-gamma. These experiments thus supply the first evidence that the inductive phase of the immune response is not promoted by trophoblast cell class I MHC antigens, which could explain the failure of mothers to mount immune responses to class I MHC positive trophoblast cells.