Changes of plasma fibronectin and fibronectin-fibrin complexes in dams of stillborn dairy calves.

BACKGROUND: Fibronectin (FN) is a large (450-500 kDa), multidomain and multifunctional glycoprotein existing in mammalian tissues. Some fibronectin (FN) molecular forms might be involved in biological processes occurring within the perinatal period, such as tissue remodeling, coagulation, and repair. RESULTS: In this study fibronectin (FN) and fibrinogen (Fb) concentrations and FN-fibrin complexes occurrence and its relative amounts with increasing high molecular masses were respectively determined by ELISA, heat precipitation, and SDS-agarose-immunoblotting methods. Plasma samples from three groups of dams with: 1) singleton stillborn calf without or with negligible autolytic changes in internal organs (DSBn), 2) singleton stillborn calf with advanced autolytic changes in internal organs (DSBa), 3) singleton live-born control calf (DC), and 4) a group of cows during mid to late lactation (LC) were analyzed. Maternal plasma FN concentration in the DSBn and DSBa groups was significantly lower than in the LC group. The plasma samples of DSBa showed a significantly lower FN concentration than in the DC group. Plasma Fb concentration was significantly higher in the DSBa and DSBn, than in the LC group. FN immunoblotting of the cow plasma samples revealed, besides an FN-dimer band, the presence of supramolecular FN-fibrin bands corresponding to FN-fibrin complexes with increasing molecular masses: up to 5 bands from 750 kDa to 1900 kDa in the DSBn and DSBa plasma samples, two bands of 750 and 1000 kDa in the DC group, and only the smallest one of 750 kDa in the LC group. CONCLUSIONS: The observed low FN concentration and occurrence of supramolecular FN-fibrin complexes (1000 kDa and more) in the maternal plasma comparing to cows in lactation might have been associated with periparturient changes in tissues. The presence in maternal plasma of high-molecular FN-fibrin complexes (1300-1900 kDa) arouse the question if this is the consequence of calf perinatal mortality.

Delivery-associated presence of supramolecular fibronectin-fibrin complexes in puerperal and cord plasma.

Objective: The variable fibronectin (FN) molecular forms are known to be engaged in coagulation and fibrinolysis pathways as well as tissue remodeling and repair processes. Some of them seem to be indispensable molecules within intensive biological processes associated with delivery. The aim of the study was to evaluate the FN molecular status in maternal and cord plasma after vaginal birth and cesarean section (C-section). Materials and methods: The study included nonpregnant women's plasma samples (n = 31) and puerperal and cord plasma samples collected from 49 mothers who delivered healthy newborns at term by vaginal birth (n = 25) and C-section (n = 24). The maternal and cord plasma FN concentrations and presence and relative ratios of different FN-fibrin complexes were determined by ELISA and sodium dodecyl sulfate (SDS) -agarose immunoblotting, respectively. Results: FN concentration in puerperal plasma after vaginal birth (232.08 ± 71.8 mg/L) and C-section (228.17 ± 71.2 mg/L) was significantly higher than in the plasma of nonpregnant women (190.00 ± 48.75 mg/L). In contrast, FN concentration in cord plasma of the C-section group (101.95 ± 30.3 mg/L) was significantly lower than that of the vaginal birth group (121.80 ± 22.2 mg/L). Immunoblotting of puerperal and cord plasma distinguished the most abundant dimeric plasma FN form, the 220-280-kDa FN degradation products and 750-1900-kDa FN-fibrin complexes, which occurred more frequently and in higher amounts in puerperal and cord plasma groups than the nonpregnant women group, although independently of the mode of delivery. Conclusions: Occurrence and relative amount of delivery-associated FN-fibrin complexes in both puerperal and cord plasmas might be bound with the physiological adaptive mechanisms reducing the risk of hemorrhage and intensive remodeling and repair processes after delivery.

Potential plasma biomarkers of bladder cancer identified by proteomic analysis: A pilot study.

BACKGROUND: Bladder cancer diagnosis and surveillance includes cystoscopy and cytology. New methods for the detection of bladder cancer are needed, because cystoscopy is invasive and expensive, and because urine cytology is not sensitive enough. OBJECTIVES: The aim of the study was to select potential plasma protein markers for bladder cancer which could be useful in developing a specific laboratory test to improve diagnosis and to establish treatment strategies in order to prevent the recurrence of the disease. MATERIAL AND METHODS: Plasma proteome maps were prepared based on 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), combined with image gel analysis and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry of plasma samples from patients with urothelial bladder cancer, and they were compared to normal samples. RESULTS: The analyses of bladder cancer plasma samples allowed us to distinguish 3 groups of proteins whose relative abundance differed from that in normal samples. The 1st one comprised modified forms of plasma transferrin, fibrinogen gamma and complement C3b, which were absent in normal plasma. The 2nd group comprised haptoglobin, alpha-2-macroglobulin, vitamin D-binding protein, and pigment epithelium-derived factor, which occurred in the cancerous samples in large quantities. The 3rd group consisted of 3 molecular forms of immunoglobulin M (IgM), the relative abundance of which was significantly lower in the cancerous plasma samples. CONCLUSIONS: The data indicated potential plasma biomarkers associated with inflammation, immunity and coagulation processes accompanying bladder cancer. They could be used for the development of a laboratory test(s) useful in clinical practice.

Time-dependent changes in extra-domain A-fibronectin concentration and relative amounts of fibronectin-fibrin complexes in plasma of patients with peripheral arterial disease after endovascular revascularisation.

Fibronectin (FN) may be involved in time- and stage-dependent and inter-related controlled processes of inflammation, coagulation, and wound healing accompanying peripheral arterial disease (PAD). In the present study, FN and FN-containing extra-domain A (EDA-FN), macromolecular FN-fibrin complexes, and FN monomer were analysed in the plasma of 142 PAD patients, including 37 patients with restenosis, for 37 months after revascularisation. FN concentration increased significantly in the plasma of PAD patients within 7 to 12 months after revascularisation, whereas the high concentration of EDA-FN was maintained up to 24 months, significantly higher in the group 7 to 12 months after revascularisation with recurrence of stenosis and lower in the PAD groups 1 to 3 months and 4 to 6 months after revascularisation with comorbid diabetes and ulceration, respectively. The relative amounts of FN-fibrin complexes up to 1600 kDa and FN monomer were significantly higher, within intervals of 4 to 24 months and 4 to 6 months after revascularisation, respectively. Moreover, the relative amounts of 750 to 1600 kDa FN-fibrin complexes within 13 to 24 months after revascularisation were higher in comparison with those in the group without restenosis. In conclusion, high levels of EDA-FN and FN-fibrin complexes could have potential diagnostic value in the management of PAD patients after revascularisation, predicting restenosis risk.

Analysis of Soluble Molecular Fibronectin-Fibrin Complexes and EDA-Fibronectin Concentration in Plasma of Patients with Atherosclerosis.

Atherosclerosis, a chronic vascular disease, leads to molecular events bound with interplaying processes of inflammation and coagulation. In the present study, fibronectin (FN), FN containing extra domain A (EDA-FN), frequency of occurrence, and relative amounts of soluble plasma FN-fibrin complexes were analyzed in 80 plasma samples of patients suspected of coronary artery disease based on clinical evaluation and changes in arteries found by computed tomographic coronary angiography. The study showed that in the plasma of the patients' group with high risk of coronary artery disease EDA-FN concentration was significantly higher (3.5 ± 2.5 mg/L; P < 0.025) and the molecular FN-fibrin complexes of 1000 kDa and higher occurred more often than in the groups of patients with mild risk of coronary artery disease and the normal age-matched. The increased level of EDA-FN and occurrence of FN-fibrin complexes could have a potential diagnostic value in the diagnosis and management of patients with coronary artery disease.

Occurrence of fibronectin-fibrin complexes in plasma of patients with multimorbidity due to the inflamm-aging phenomenon.

BACKGROUND: Multimorbidity is the co-occurrence of chronic diseases associated with low-grade chronic inflammation of connective tissue. AIM OF STUDY: Frequency of occurrence and relative amounts of fibronectin (FN) complexes with fibrin (FN-fibrin) and FN monomer were analyzed in 130 plasma samples of 18 to 94-year-old multimorbid patients in relation to concentrations of FN and extra domain A (EDA)-FN, and C-reactive protein (CRP) as well as to age, number of coexisting chronic diseases and presence of specified diseases. RESULTS: Immunoblotting revealed, besides FN dimer, the presence of FN monomer, and 750-, 1000-, and 1300-kDa FN-fibrin complexes in the multimorbid plasmas. The FN-fibrin complexes appeared more frequently and in higher relative amounts, but FN monomer less frequently and in a lower relative amount in the groups of elderly multimorbid patients, with a higher number of coexisting diseases and with dominance of cardiovascular diseases and osteoarthrosis, and with CRP concentration of 3-5mg/l. In contrast, the normal plasma contained only the FN-fibrin complex of 750 kDa in a lower relative amount, but with an increasing amount with normal aging. Moreover, FN concentration increased and EDA-FN decreased with the number of co-existing diseases and aging of patients, although both concentration values were lower than in the age-matched normal groups. FN concentration was the lowest in the exacerbation of a chronic disease and EDA-FN in the stable chronic disease groups. CONCLUSION: The alterations in plasma FN molecular status were associated with micro-inflammation and micro-coagulation, as well as multimorbidity of subjects and their physiological aging.

Lectin-based analysis of fucosylated glycoproteins of human skim milk during 47 days of lactation.

Glycoproteins of human milk are multifunctional molecules, and their fucosylated variants are potentially active molecules in immunological events ensuring breastfed infants optimal development and protection against infection diseases. The expression of fucosylated glycotopes may correspond to milk maturation stages. The relative amounts of fucosylated glycotopes of human skim milk glycoproteins over the course of lactation from the 2(nd) day to the 47(th) day were analyzed in colostrums, transitional and mature milk samples of 43 healthy mothers by lectin-blotting using α1-2-, α1-6-, and α1-3-fucose specific biotinylated Ulex europaeus (UEA), Lens culinaris (LCA), and Lotus tetragonolobus (LTA) lectins, respectively. The reactivities of UEA and LCA with the milk glycoproteins showed the highest expression of α1-2- and α1-6-fucosylated glycotopes on colostrum glycoproteins. The level of UEA-reactive glycoproteins from the beginning of lactation to the 14(th) day was high and relatively stable in contrast to LCA-reactive glycoproteins, the level of which significantly decreased from 2-3 to 7-8 days then remained almost unchanged until the 12(th)-14(th) days. Next, during the progression of lactation the reactivities with both lectins declined significantly. Eighty percent of α1-2- and/or α1-6-fucosylated glycoproteins showed a high negative correlation with milk maturation. In contrast, most of the analyzed milk glycoproteins were not recognized or weakly recognized by LTA and remained at a low unchanged level over lactation. Only a 30-kDa milk glycoprotein was evidently LTA-reactive, showing a negative correlation with milk maturation. The gradual decline of high expression of α1-2- and α1-6-, but not α1-3-, fucoses on human milk glycoproteins of healthy mothers over lactation was associated with milk maturation.

O-glycosylation of α-1-acid glycoprotein of human milk is lactation stage related.

BACKGROUND: Human milk provides a multitude of glycoproteins, including highly glycosylated α-1-acid glycoprotein (AGP), which elicits anti-inflammatory and immunomodulatory properties. The milk AGP glycoforms may provide the breastfed infant with a wide range of biological benefits. Here, we analyzed the reactivity of O-linked sugar-specific lectins with human milk AGP over the process of lactation and compared the results with those of the lactating mother's plasma. MATERIALS AND METHODS: Relative amounts of human skim milk AGP O-glycans were analyzed in early colostrum, colostrum, and transitional and mature milk samples of 127 healthy mothers by lectin-AGP enzyme-linked immunosorbent assay using sialyl T (sialyl-α2,3/α2,6 Galß1,3GalNAc-), asialyl T (Galß1,3GalNAc-), and Tn (GalNAc-) antigen-specific biotinylated Artocarpus integrifolia (Jacalin), Arachis hypogaea (PNA), and Vicia villosa (VVA) lectins, respectively. RESULTS: Milk AGP elicited high expression of Jacalin- and PNA-reactive glycotopes and low expression of VVA-reactive glycotopes, which were absent on plasma AGP of lactating mothers and healthy individuals. The expression of sialyl, asialyl T, and Tn glycotopes of human milk AGP was lactation stage related. The relative amount of Jacalin-reactive AGP glycotope was highest in the colostrum samples and then decreased starting from Day 8 of lactation. In contrast, an increase of the relative amount of PNA-reactive glycotope with milk maturation was observed. The relative amount of VVA-reactive glycotope remained almost constant over the development of lactation. CONCLUSIONS: Milk AGP differs from mother's plasma AGP by the presence of O-linked sialylated and asialylated T as well as Tn antigens. The variation of the expression of sialylated and asialylated T and Tn antigens on AGP is associated with milk maturation.

[Uroplakins as markers of diseases of the urinary system]. / Uroplakiny jako markery chorób ukladu moczowego

An unique element of bladder urothelium is a multilayer membrane, which extends from the renal pelvis to the urethra. Urotelial membrane covers more than 90% of the inner portion of the bladder and is in direct contact with urine. Urothelium is composed of characteristic two-dimensional, asymmetric plaques, composed of uroplakins (UP), differentiated, hexagonally arranged proteins. The unique structure of the urothelial plaques determines the tightness, integrity and strength of the urothelium, prevent rupture of the walls of the bladder during the build-up of urine in the bladder and protects against the toxic ingredients. Uroplakins are tissue-specific, heterogeneous glycoproteins whose oligosaccharide part plays a specific role in the structure and function of urothelium. Disorders of normal expression of uroplakins are highly associated with the pathogenesis in infection and urinary tract malignancies, primary vesico-urinary reflux, hydronephrosis and renal impairment. The emergence of uroplakins in urine and / or plasma may have a potential role in the early detection of bladder tumors. In this paper, the structure and function of uroplakins types Ia, Ib, II, IIIa, their natural oligomerization into heterodimers, tetramers and hexamers, and the role in the construction of asymmetric and flexible urothelial epithelium is presented. We discuss the potential role of uroplakins in laboratory diagnosis of umbrella cell differentiation and in the screening analysis of urinary bladder disorders. The possibilities of using the knowledge of uroplakins in clinical settings as well as in modern strategies for treatment of infectious diseases and cancer of the urinary tract are highlighted.

Terminal glycotope expression on milk fibronectin differs from plasma fibronectin and changes over lactation.

OBJECTIVES: Fibronectin (FN) is a multifunctional glycoprotein appearing in various glycovariants with potential biological activities. Using lectins we analyzed the expression of terminal glycotopes on human milk fibronectin over lactation and compared it with that of the mother's plasma. DESIGN AND METHODS: FN concentration and relative amounts of its fucosylated and sialylated glycovariants as well as O-glycans were analyzed in early colostrum, colostrum, transitional and mature milk samples of 132 healthy mothers by lectin-FN-ELISA using α2,3- and α2,6-sialic acid, α1,2-, α1,3-, and α1,6-fucose, and sialyl-T, asialyl-T and Tn antigen specific biotinylated Maackia amurensis, Sambucus nigra, Ulex europaeus, Tetragonolobus purpureus, Lens culinaris, Artocarpus integrifolia, Arachis hypogaea, and Vicia villosa lectins, respectively. RESULTS: FN concentration was almost unchanged during human milk maturation and was about 150 times lower than in plasma of lactating mothers. Milk FN elicited significantly higher expression of sialylated glycotopes including sialyl-T antigen than plasma FN, and contained fucose-linked glycans, as well as T and Tn antigens absent in plasma FN. With milk maturation the expression of α2,6-sialylated, sialyl-T, α1,6- and α1,2-fucosylated epitopes decreased in transitional milk compared with colostrum, whereas that of asialyl-T antigen increased. The expression levels of α2,3-sialyl- and α1,3-fucosyl-glycotopes and Tn antigen on FN were low and did not change over lactation. CONCLUSION: The expression of terminal sugars on milk FN is different from that of plasma FN of the lactating mother and is associated with milk maturation. The analysis of degree of milk sialylation and fucosylation should be considered during control of biochemical quality of milk collected in milk banks.

Occurrence of soluble supra-molecular FN-fibrin complexes in the plasma of children with recurrent respiratory infection.

OBJECTIVES: Fibronectin (FN) is able to bind fibrin and FN-fibrin complexes and is found in the plasma of some patients suffering from inflammatory disease. The present study was undertaken to determine whether soluble supra-molecular FN-fibrin complexes were present in the plasma of children with recurrent respiratory infections (RRI). DESIGN AND METHODS: The frequency of occurrence and relative amounts of the supra-molecular FN-fibrin forms, concentrations of immunoglobulins and numbers of natural killer cells (NK) were determined in the plasma of children with recurrent respiratory infections. The frequencies of these parameters were compared with their frequencies in the plasma of children with acute respiratory infections and plasma from healthy children. RESULTS: SDS-agarose immunoblotting of patients' plasma revealed the presence of several additional FN-fibrin bands, with decreasing electrophoretic mobilities and increasing molecular masses of 750 kDa, 1000 kDa, 1300 kDa, 1600 kDa and 1900 kDa. Such FN-fibrin complexes occurred with higher frequency and in larger amounts in the plasma of children with RRI and acute infection than they did in plasma from normal children. Moreover, bands above 1000 kDa were absent in most young healthy individuals. The occurrence of FN-fibrin complexes did not correlate with either immunoglobulin concentrations, or with the number of NK cells. CONCLUSIONS: The occurrence of plasma supra-molecular FN-fibrin complexes is associated with acute and recurrent respiratory infections of children.

Glycosylation of uroplakins. Implications for bladder physiopathology.

Urothelium, a specialized epithelium, covers the urinary tract and act not only as a barrier separating its light from the surrounding tissues, but fulfills an important role in maintaining the homeostasis of the urothelial tract and well-being of the whole organism. Proper function of urothelium is dependent on the precise assemble of highly specialized glycoproteins called uroplakins, the end products and differentiation markers of the urothelial cells. Glycosylation changes in uroplakins correlate with and might reflect progressive stages of pathological conditions of the urothelium such as cancer, urinary tract infections, interstitial cystitis and others. In this review we focus on sugar components of uroplakins, their emerging role in urothelial biology and disease implications. The advances in our understanding of uroplakins changes in glycan moieties composition, structure, assembly and expression of their glycovariants could potentially lead to the development of targeted therapies and discoveries of novel urine and plasma markers for the benefit of patients with urinary tract diseases.

Lactation stage-related expression of sialylated and fucosylated glycotopes of human milk α-1-acid glycoprotein.

BACKGROUND: Because terminal sugars of α-1-acid glycoprotein (AGP) are reported to be involved in anti-inflammatory and immunomodulatory processes, their expressions might have an influence on the proper function of immune system of newborns. Here, relative amounts of sialylated and fucosylated glycotopes on human milk AGP over normal lactation were investigated. MATERIALS AND METHODS: AGP concentration and relative amounts of its sialylated and fucosylated glycovariants were analyzed in early colostrum, colostrum, and transitional and mature milk samples of 127 healthy mothers by lectin-AGP enzyme-linked immunosorbent assay using α2,3- and α2,6-sialic acid and α1,2-, α1,3-, and α1,6-fucose specific biotinylated Maackia amurensis, Sambucus nigra, Ulex europaeus, Tetragonolobus purpureus, and Lens culinaris lectins, respectively. RESULTS: AGP concentration in human milk was about 30 times lower than in plasma of lactating mothers and decreased gradually over lactation. Milk AGP showed significantly higher expression of sialylated and fucosylated glycotopes in comparison with those of plasma AGP. Milk AGP glycovariants containing α2,6-sialylated and α1,6- and α1,2-fucosylated glycotopes showed the highest relative amounts in early colostrums. With progression of lactation, the expressions of glycotopes α1,2-fucosylated decreased starting from Day 4 and those of α2,6-sialylated and α1,6-fucosylated from Day 8 of lactation, whereas the level of α2,3-sialyl-glycotope was almost constant over 45 days of lactation. In contrast, the expression of α1,3-linked fucose on AGP was low in colostrums and significantly higher in transitional and mature milk. CONCLUSIONS: The relative amounts of sialylated and fucosylated glycovariants of human hindmilk AGP significantly varied between Days 2 and 45 of normal lactation.

Identification of soluble supramolecular FN-fibrin complexes in human plasma.

SDS-agarose FN immunoblotting of 257 normal and pathological human plasma samples revealed the ladder pattern of multiple plasma FN bands which corresponded to FN monomer and dimer, and 5 FN-fibrin bands with increasing molecular masses. The FN-fibrin bands of about 750 kDa, 1000 kDa, 1300 kDa, 1600 kDa, and 1900 kDa appeared more frequently and in significantly higher relative amounts in the pathological samples (P < 0.000) than in relatively healthy individuals. The revealing of high-molecular FN-fibrin complexes by SDS-agarose FN immunobloting might have the potential to become a laboratory biomarker of some diseases in which the coagulation system is triggered.

Changes in fucosylation of human seminal IgG and secretory component of IgA in leukocytospermic patients.

Our study compares the status of human seminal plasma immunoglobulin G (IgG) and IgA secretory component (SC) fucosylation between infertile leukocytospermic and normal, fertile normozoospermic patients. The seminal IgG and SC are decorated with AAL-reactive core fucose, and antennary UEA- and LTA-reactive fucose of Lewis(y) and Lewis(x) structures, respectively. However, a correlation between IgG core fucosylation and IgG concentration (r = -0.52; p < 0.0003) was observed. The IgG present in leukocytospermic samples is characterized by lower expression of core fucose than in the normal group (0.82 ± 0.3 AU and 1.2 ± 0.3 AU, respectively; p < 0.002). In seminal plasma the SC is present in two forms: 78-kDa and 63-kDa. The present study has also shown a higher AAL and LTA specific reactivity of glycans expressed in 63-kDa SC, in comparison to 78-kDa SC, in the normal group. In leukocytospermia, the values of specific lectin reactivity for core fucose, fucose α(1-2)- and α(1-3)- linked, were similar for both SC bands. Moreover, the present study has shown that in leukocytospermic samples the mean concentrations of IgG and S-IgA are twice as high (131.68 ± 102.6 mg/l and 36 ± 27 mg/l, respectively) as in the normal group (67.68 ± 29.2 mg/l; p < 0.02, and 19 ± 18 mg/l, p < 0.019, respectively). The analysis of IgG and SC fucosylation status and the determination of IgG and S-IgA concentrations in seminal plasma might constitute a valuable diagnosis tools for the evaluation of male infertility associated with leukocytospermia with accompanying inflammation.

Human seminal fibronectin fragmentation patterns and their domain immunoreactivities in leucocytospermic patients.

The aim of the work was to analyse fibronectin (FN) domain immunoreactivities and profiles of FN fragmentation in seminal plasmas of fertile normozoospermic and infertile leucocytospermic male patients. ELISA with domain-specific monoclonal antibodies and immunoblotting were used in these measurements. Immunoblotting of normal and leucocytospermic seminal plasmas revealed the presence of twelve FN bands of ~70-196kDa with nearly identical FN profiles under reducing and non-reducing conditions. The epitopes of the cell-, fibrin-, collagen-binding FN domains and the extra domain A (EDA) FN segment retained the ability to bind their specific monoclonal antibodies, whereas the fibrin-heparin domain (N-terminal end) and the area around the disulfide bridges (C-terminal end) of the FN polypeptide did not show any reactivities with their respective specific antibodies. The mean values of cell- (338.4±138.4 and 398.3±310mgL(-1)), fibrin- (79.1±38.5 and 145.2±188.8mgL(-1)) and collagen-binding (19±19.8 and 50.9±73.4mgL(-1)) FN domain immunoreactivities and the relative amount of (EDA)FN did not show any significant differences between the normal and leucocytospermic groups. The high values of standard deviations for the FN domain immunoreactivities in the leucocytospermic group probably results from different aetiology of leucocytospermia. The profile of FN fragmentation and alterations of FN domain immunoreactivities in seminal plasma may influence their engagement in the fertilisation process. The analysis of seminal FN molecular status would be helpful for selecting the highest quality spermatozoa for use in assisted reproduction techniques.

EDA-containing fibronectin levels in the cerebrospinal fluid of children with meningitis.

Fibronectin containing an alternatively spliced extra domain A (EDA-FN) participates in diverse biological cell functions, being also directly or indirectly engaged during an inflammatory response to brain injury and/or neuron regeneration. We analyzed FN and EDA-FN isoform levels by ELISA in 85 cerebrospinal fluid samples and 67 plasma samples obtained from children suffering from bacterial or viral meningitis and non-meningitis peripheral inflammation. We have found that the cerebrospinal level of EDA-FN was significantly lower in the bacterial meningitis group than in the viral- and non-meningitis groups. In the patients' plasma, EDA-FN was almost undetectable. The determination of fibronectin containing the EDA segment might be considered as an additional diagnostic marker of bacterial meningitis in children.

[Proteins of human milk involved in immunological processes]. / Bialka mleka ludzkiego zaangazowane w procesy immunologiczne.

Human milk contains a lot of components (i.e. proteins, carbohydrates, lipids, inorganic elements) which provide basic nutrients for infants during the first period of their lives. Qualitative composition of milk components of healthy mothers is similar, but their levels change during lactation stages. Colostrum is the fluid secreted during the first days postpartum by mammary epithelial cells. Colostrum is replaced by transitional milk during 5-15 days postpartum and from 15 days postpartum mature milk is produced. Human milk, apart from nutritional components, is a source of biologically active molecules, i.e. immunoglobulins, growth factors, cytokines, acute phase proteins, antiviral and antibacterial proteins. Such components of human milk are responsible for specific biological activities of human milk. This secretion plays an important role in growth and development of newborns. Bioactive molecules present in the milk support the immature immune system of the newborn and also protect against the development of infection. In this article we describe the pathways involved in the production and secretion of human milk, the state of knowledge on the proteome of human milk, and the contents of components of milk during lactation. Moreover, some growth factors and proteins involved in innate and specific immunity, intercellular communication, immunomodulation, and inflammatory processes have been characterized.

Alterations in molecular status of plasma fibronectin associated with aging of normal human individuals.

OBJECTIVES: Senescence, progressive deterioration of many bodily functions might be associated with age-dependent alterations of plasma fibronectin (FN) molecular status (i.e., domain, glycotope, and molecular form expressions). DESIGN AND METHODS: FN molecular status was analyzed in 127 plasma samples of healthy individuals in groups of newborns, and subjects aged 3-14, 15-39, 41-59, and 60-82 years by FN-ELISA, lectin-FN-ELISA, and immunoblotting using a set of domain-specific monoclonal antibodies, specific lectins, and monoclonal antibody to FN, respectively. RESULTS: During the first four decades of human life the levels of cell-binding-, carboxyl-terminal-, collagen-, heparin-, and fibrin-domains of plasma FN gradually increased. In subjects aged up to 82 years the cell-binding and carboxyl-terminal FN domain concentrations did not change, while the heparin, fibrin, and collagen domains significantly increased. The relative reactivity of plasma FN with Maackia amurensis lectin, specific to α2,3-linked sialic acid, significantly decreased after birth, reaching a stable level in the subsequent life period, whereas with Sambucus nigra lectin, specific to α2,6-linked sialic acid, it significantly decreased in the 60-82 year old group. Moreover, the appearance of 280-kDa and 320-kDa FN bands, absent in young and mature healthy individuals, was found in the groups of 41-59 and 60-82 year olds. CONCLUSIONS: The alterations of FN molecular status throughout growth, maturation and senescence might be associated not only with disturbances in the balance of FN production rate and degradation, but concomitantly with conformational rearrangements of FN and its engagement in age-related vascular remodeling processes.

Fibronectin molecular status determination useful to differentiate between rheumatoid arthritis and systemic lupus erythematosus patients.

To find whether the plasma fibronectin (FN) molecular status can be useful to differentiate between rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). The expression of plasma FN domains was determined by ELISA using monoclonal domain-specific antibodies. FN molecular forms were revealed by immunoblotting and analyzed by densitometry. The following findings were found: (1) Mean values of (Fibrin-Heparin)FN concentration were lower in SLE and RA patients than in normal plasmas. The cut off points at 31 mg/l in SLE and at 45 mg/l in RA showed a sensitivity and specificity of 54, 55 and 75%, respectively. (2) Mean values of concentrations of (CBD)FN and (Ct)FN were lower in SLE than those in normal and RA plasmas. Quantified data showed the cut off points of (CBD)FN and (Ct)FN at 200 mg/l (58% of sensitivity, 56% of specificity) and 350 mg/l (58% of sensitivity, 58% of specificity) in SLE, as well as at 295 mg/l (52% of sensitivity, 51% of specificity) and 460 mg/l in RA (70% of sensitivity, 73% of specificity). (3) The plasma FN immunopatterns, characterized by the presence of high-molecular (260-310 kDa) and/or low-molecular (158-209 kDa) FN bands, were specific only for SLE samples. The analysis of plasma FN status revealed by its Fibrin-Heparin-, CBD- and Ct-domain reactivity with monoclonal antibody and immunoblotting can be helpful to differentiate the SLE in respect to RA and normal plasmas.