[Molecular markers of ovarian carcinoma]. / Molekularne markery raka jajnika.

Definition of a tumor marker and significance of proper choice of ovarian cancer marker in diagnosis, prognosis and monitoring effects of chemotherapy, are pointed out. Markers of malignant ovarian tumors were classified into oncogenes and oncoproteins, antigens, hormones, enzymes and their inhibitors, acute phase proteins (cytokines, growth factors, haptoglobin), hemostasis and some others. Methods of quantitative determinations of cancer markers are presented.

Serum haptoglobin, CA 125 and interleukin 6 levels in malignant and non-malignant tumors of the ovary.

In sera of women with malignant and non-malignant ovarian tumors, concentrations of the following proteins were determined: haptoglobin, measured by the reactions either with hemoglobin (Hp-Hb) or with concanavalin A (Hp-Con A), CA 125 antigen and interleukin 6 (IL-6). Besides preoperative data, obtained from all the patients and the group of healthy women, in the patients with ovarian cancer (III/IV FIGO stages) effects of administration of cytostatics were measured by monitoring changes in the levels of the examined parameters through the course of the therapy. All the results were submitted to statistical evaluation which showed differences and correlations among definite groups (normal/non-malignant, normal/cancers, non-malignant/cancers). Similar results were obtained for Hp-Hb and CA 125 determination. Moreover, a correlation between haptoglobin level and age was found. Patterns of monitoring revealed a surprising absence of Hp-Con A interaction in some cases.

[Pathophysiologic programming of cell death]. / Patofizjologia programowanej smierci komórki.

In multicellular organisms homeostasis depends on a balance between cell proliferation and cell death. In this review principles of the physiology of programmed cell death (apoptosis), i.e. biochemical features, involved genes and proteolytic enzymes, are described. Alterations in apoptosis contribute to the pathogenesis of a number of human diseases, including cancer, viral infections, inflammation, hematopoietic and immunological system defects (e.g. AIDS), neurodegenerative disorders. Specific effect on regulation of apoptosis might lead to new possibilities for treatment. Methods of quantitative determinations of apoptosis are discussed.

Biological functions of haptoglobin--new pieces to an old puzzle.

Haptoglobin, an "acute phase" protein, has different functions, which display genetic polymorphism. The complex of haptoglobin with haemoglobin is metabolized in the heptic reticuloendothelial system. Biosynthesis of haptoglobin occurs not only in the liver, but also in adipose tissue and in lung; providing antioxidant and antimicrobial activity. Changes in the measured concentrations of haptoglobin in serum may help to assess the disease status of patients with inflammations, infections, malignancy etc. (increases) as well as in haemolytic conditions (decreases). Haptoglobin plays a role in stimulation of angiogenesis and has highly potent cholesterolcrystallization-promoting activity. Probably the most important biological function of haptoglobin consists in the host defence responses to infection and inflammation, acting as a natural antagonist for receptor-ligand activation of the immune system.

Measurements of haptoglobin by the reaction with concanavalin A in sera of patients with ovarian tumours.

The concentration of haptoglobin in sera of healthy women and patients with non-malignant and malignant ovarian tumours was measured by two methods, i.e. complex formation with haemoglobin and complex formation with concanavalin A. High correlation (r = 0.74) between both the methods was found in the group of healthy women, but correlation coefficients were much lower in the group of non-malignant and malignant tumours (r = 0.42 and 0.37, respectively). Direct determinations of haptoglobin, and calculation of the ratio of haptoglobin bound to concanavalin A to haptoglobin bound to haemoglobin revealed statistically significant differences among the examined groups. Comparison of two methods of haptoglobin quantitation suggest that processes connected with ovarian disorders may alter the glycosylation of haptoglobin oligosaccharide chains.

Quantitation of human haptoglobin by ELISA system based on streptococcal haptoglobin receptors.

Streptococcus pyogenes cells with binding properties for human haptoglobin were used for quantitative determination of the acute phase protein, haptoglobin in various biological fluids. The S. pyogenes cells with protein surface antigen T4 served as solid phase in a microtitre ELISA system. After binding to the bacteria the amount of haptoglobin could be quantified by polyclonal or monoclonal antibodies. The constructed ELISA proved to be sensitive and correlated well with a conventional peroxidase method and with an immunoassay based on hemoglobin binding to haptoglobin.

[Cytokines, their receptors and inhibitors. Importance in laboratory diagnosis of rheumatoid arthritis]. / Cytokiny, ich receptory i inhibitory. Znaczenie w diagnostyce laboratoryjnej reumatoidalnego zapalenia stawów.

The present paper reviews the literature on biological activities of human cytokines in homeostasis, as modulated by their secretion, expression of membrane-bound and soluble receptors as well as by means of other cytokines and specific inhibitors, respectively. The role of structure and properties of interleukins 1, 2, 6 and their receptors in pathophysiology and laboratory diagnostics of rheumatoid arthritis, is discussed. Methods of quantitative determination of cytokines are described.

Development of concanavalin A-enzyme immunosorbent assay for glycated haptoglobin using polyclonal and monoclonal antibodies.

Two-site lectin-haptoglobin-enzyme immunosorbent assay (L-Hp-ELISA), is described. Haptoglobin binding to Concanavalin A, immobilized to polystyrene microtiter plate, was estimated by anti-haptoglobin polyclonal and monoclonal antibodies conjugated with horse-radish peroxidase. The range of haptoglobin binding to Concanavalin A, measured by the L-Hp-ELISA was 25 to 300 ng/ml using polyclonal, and 50 to 600 ng/ml using monoclonal anti-haptoglobin antibodies, respectively.

Acute phase reactants and circulating immune complexes in patients with ovarian carcinoma.

Serum levels of haptoglobin (HP), sialic acid total (NAN) and lipid-bound (NAL), seromucoid (SER), its content in total protein (%SER), as well as circulating immune complexes (CIC), were measured in sera of women with ovarian carcinoma, prior to their treatment and through the course of chemotherapy, remission and recurrence of malignancy, respectively. Control groups consisted of healthy women and patients with benign tumors (ovarian cysts and uterine myomas). Pretreatment measurements of acute phase reactants discriminated cancers (FIGO stages I+II, III, IV) from healthy group, however differences between benign tumors and stages of ovarian cancer were not so distinct. Changes in the examined parameters (acute phase reactants) indicated satisfactorily a response to the administered chemotherapy and early signs of the progression of the disease. Because of great variations in serum CIC concentrations, they were found to be of no value either in diagnosis or in the surveillance of the disease status.

Characterization of haptoglobin-binding properties of streptococci of serological group G.

Two group G streptococcal cultures (G 10187, G 11122) with surface antigen T4 possess surface receptors for human haptoglobin (Hp). G 10187 additionally interacted with immunoglobulin G and albumin, G 11122 with fibrinogen and fibronectin. Binding of 125I-Hp 2-1 was time-dependent, saturable, reversible in the presence of unlabelled Hp and could be inhibited by unlabelled human-Hp 2-1, -Hp 2-2, -Hp 1-1, Hp-hemoglobin complexes and by Hp preparations from pigs, horses and rabbits. The Hp binding sites could be destroyed by heat treatment (95 degrees C) and by proteolytic treatment of the bacteria. Hp binding sites were solubilized from group G streptococcal surface by heat treatment of the bacteria at acid pH and subsequently isolated by affinity chromatography on Hp 2-1 sepharose. SDS-PAGE and Western blotting of the Hp binding proteins revealed numerous protein bands with 125I-Hp 2-1 binding activity. Specific antibodies against G streptococcal binding proteins prepared in chickens inhibited binding of 125I-Hp to group G and group A streptococci, but not to Actinomyces pyogenes.

Enzyme immunoassay to measure low levels of haptoglobin in biological fluids.

A novel immunoenzymatic assay is described for the quantitation of human haptoglobin (Hp). Two binding sites on the Hp molecule, namely for hemoglobin (Hb) and for the specific antibody, are involved in the reaction. Hb adsorbed onto the polystyrene microplate binds Hp present in any biological fluid. The formed Hp-Hb complex is detected with horse-radish peroxidase conjugated with anti-Hp antibody. By means of this ELISA, Hp may be measured in the range of 5 to 150 micrograms/L. Comparison of the Hp-ELISA with two other methods of Hp determination resulted in correlation coefficients of 0.97 to 0.99. Intra- and inter-assay coefficients of variation ranged from 4.7 to 6.7%. Hp levels were measured in urine, cord serum, cerebrospinal fluid, amniotic fluid and saliva.

Monoclonal antibodies against human haptoglobin.

Three monoclonal antibodies: 2.36.71.41, 7.60.66.55, and 18.4.40. 80 to human haptoglobin 2-1 were produced, purified and characterized. The affinity constants ranged within 0.3-2.4 x 10(8) M-1. The monoclonal antibodies 7.60.66.55 and 18.4.40.80 reacted with beta subunit of haptoglobin, showed similar epitope affinities and epitope densities on main haptoglobin types. However, the epitope on the haptoglobin molecule for the monoclonal antibody 18.4.40.80 occupied somewhat more surface than that for the antibody 7.60.66.55. The monoclonal antibody 2.36.71.41 was able to bind both alpha and beta chains of haptoglobin. In ELISA affinity reactions this antibody achieved with haptoglobin 2-2 the plateau phase at absorbance values 15% higher than with haptoglobin 2-1, and 60% higher than with haptoglobin 1-1. End-point titration of the monoclonal antibody 2.36.71.41 against three haptoglobin types showed differences in titer, indicating distinct epitope densities.

The model of allergic encephalomyelitis of nonacute, phase course.

Allergic encephalomyelitis characterized by relapses and remissions was induced in chickens. The contents of M and G immunoglobulins in the serum was observed to increase at disease attacks and to decrease at its remissions. Similar correlation--though not so clear cut was noted with antitrypsin level. The usefulness of this model in pharmacotherapeutic investigations is discussed.

Further characterization of haptoglobin binding to streptococci of serological group A.

Certain group A streptococci with surface antigen T 4 possess surface receptors for human haptoglobin (Hp). Binding of 125I Hp 2-1 to two representative group A streptococcal cultures could be inhibited by unlabelled Hp 2-1, Hp 2-2 and Hp 1-1 but not by the alpha 1, alpha 2 or beta chains of Hp. Hp complexes formed with equine hemoglobin and asialo-Hp also reduced 125I-Hp 2-1 binding to group A streptococci. Hp binding proteins could be solubilized from streptococcal surface by hot acid treatment of the bacteria and purified by subsequent affinity chromatography on human Hp 2-1 sepharose. The isolated Hp binding proteins specifically inhibited 125I-Hp 2-1 binding to group A streptococci and retained their 125I-Hp 2-1 binding activity in a dot binding assay on nitrocellulose membranes. SDS-PAGE and protein blots of Hp binding proteins developed with 125I-labeled Hp 2-1 revealed numerous high molecular weight proteins with 125I-Hp 2-1 binding activity.

Azimexone and cysteine hydrazide in the treatment of allergic encephalomyelitis.

In the course of allergic encephalomyelitis (EAE) in chickens, azimexone suppressed the production of the specific IgM immunoglobulins. Moreover, it decreased the level of haptoglobin and sialic acid but stimulated the activity of antitrypsin. It was also observed to reduce the production of the plasmatic cells in the spleen. L-cysteine hydrazide hydrochloride decreased the level of sialic acid in EAE; intensified the activity of trypsin inhibitor and exerted no effect upon the level of antimyelin antibodies.

Microheterogeneity of alpha 1-acid glycoprotein in the sera of patients with cancer or inflammatory states of the ovaries.

alpha 1-Acid glycoprotein (AGP) and its concanavalin A-dependent four microheterogeneous fractions were measured in sera from women with ovarian carcinoma and those with an inflammatory ovarian disease. Total concentrations of AGP in inflammatory and cancerous sera were significantly higher than in control group, but did not differ from each other. In the fourth stage cancer the distribution of Con A-dependent fractions 1, 3 and 4 was significantly different as compared with control, second stage cancer and inflammatory groups.