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I Politis

Publications and source records attributed to I Politis.

At least 37 records · Page 2Linked to original sources
PubMed1995

Effect of oxygen tension on killing of Escherichia coli by bovine polymorphonuclear neutrophil leucocytes in vitro.

A batch fermenter modified to simulate the physical conditions of an inflamed v. uninflamed mammary gland was utilized to evaluate the effect of oxygen tension on killing of Escherichia coli P4 by bovine polymorphonuclear neutrophil leucocytes. Leucocytosis was simulated in vitro 4 h after inoculation with Escherichia coli by adding bovine neutrophils isolated from peripheral blood to the culture medium. Experiments were conducted at 39 degrees C in UHT treated milk. At micro-aerophilic oxygen tension (oxygen partial pressure, 3.11 kPa), bacterial numbers declined during the first hour following addition of the neutrophils. Oxygen tension declined rapidly following PMN addition. Once oxygen was depleted, neutrophil activity was presumably diminished and Esch. coli numbers began to increase. Under anaerobic conditions (oxygen partial pressure, 0.17 kPa), no reduction in population was observed. Photomicrographs taken at the time of neutrophil addition and at subsequent time intervals demonstrated a specific association between neutrophils and the pathogen. Subsequent lysis of neutrophils associated with Esch. coli growth was seen coincident with oxygen depletion.

Animals↗
PubMed1995

Effects of Staphylococcus aureus products on growth and function of bovine mammary myoepithelial cells in vitro.

The effects of culture supernatants conditioned by the growth of Staphylococcus aureus M60 on in vitro growth and functional properties of bovine mammary myoepithelial cells were examined. Myoepithelial cell proliferation was reduced by Staph. aureus M60 culture supernatants. Exposure of myoepithelial cells to culture supernatants of isogeneic mutants of Staph. aureus M60 that produced either alpha or beta toxins reduced proliferation, but to a lesser extent than supernatants from the wild type strain. Thus, alpha and beta toxins may play some role in affecting myoepithelial cell proliferation. Of the cells tested, 42% contracted following addition of oxytocin (10(-7) M) in the culture medium. Treatment of myoepithelial cells for 15 min with Staph. aureus M60 supernatants, prior to addition of oxytocin in the culture medium, increased the number of cells that contracted to 92%. Exposure of cells for 3 h to the same supernatant, prior to addition of oxytocin in the culture medium, abolished oxytocin responsiveness, had no effect on immunolocalization of actin and vimentin, but affected the localization of alpha-actinin within myoepithelial cells. Treatment of myoepithelial cells for 3 h with a combination of purified staphylococcal proteinases XVI and XVII-B abolished oxytocin responsiveness and mimicked the effect of the Staph. aureus culture supernatant. We conclude that Staph. aureus M60 culture supernatant affected proliferation and functional properties of myoepithelial cells.

Actinin↗
PubMed1995

Distribution of plasminogen activator in different fractions of bovine milk.

The type and relative amounts of plasminogen activator (PA) in different fractions of bovine milk obtained from 15 Holstein cows were examined. Raw milk was centrifuged to separate skim milk and a somatic cell pellet. PA was mainly localized within the casein fraction, being 42 times that in the serum, and in association with somatic cells. The predominant form of PA in milk casein was isolated from SDS-PAGE gel extracts and had a molecular mass of approximately 75 kDa. Its activity was increased 4.1-fold (P < 0.01) in the presence of fibrin but was unaffected by the presence of amiloride, indicating that it was due to tissue-PA. The predominant forms of PA associated with milk somatic cells were isolated from SDS-PAGE gel extracts and had molecular masses of approximately 30 and approximately 50 kDa. The activity of both proteins was unaffected by the presence of fibrin but was dramatically reduced by the presence of amiloride, indicating that they represented urokinase-PA.

Amiloride↗
PubMed1995

Effects of Staphylococcus aureus toxins on the growth of bovine mammary epithelial cells (MAC-T) in culture.

The effects of Staphylococcus aureus M60 culture supernatant on growth of mammary epithelial cells were tested in vitro. Exposure of MAC-T cells to S. aureus culture supernatant reduced cell proliferation and colony-forming ability because of reduced ability to adhere to plastic. Growth-inhibiting effects of S. aureus culture supernatant were abolished by pretreatment with trypsin. Lysis of S. aureus cells with lysostaphin demonstrated that the inhibition was also present in cell lysates. Treatment of MAC-T cells with culture supernatant from isogeneic mutants of S. aureus M60 that produced either alpha or beta toxins implicated alpha toxin as the main factor inhibiting cell proliferation.

Animals↗
PubMed1995

Effect of individual caseins on plasminogen activation by bovine urokinase-type and tissue-type plasminogen activators.

The effect was examined of individual caseins on the rate of plasminogen activation by bovine urokinase-type and tissue-type plasminogen activators. All individual caseins (alpha-CN, beta-CN, and kappa-CN) enhanced the activity of both types of plasminogen activators. Optimal concentrations for alpha-CN and beta-CN were 5 and 25 micrograms/ml, respectively. The enhancement of enzymatic activity declined when concentrations of alpha-CN and beta-CN were higher. In contrast, increasing concentrations of kappa-CN from 0 to 200 micrograms/ml resulted in corresponding increases in activity of both types of plasminogen activators. On a weight basis, alpha-CN was the most effective enhancer of plasminogen activator activity. Indirect evidence obtained with experiments utilizing alpha-CN immobilized on agarose suggested that the effect is related to extensive binding of plasminogen and both types of plasminogen activators to casein.

Amino Acid Sequence↗
PubMed1995

Subcloning the MAC-T bovine mammary epithelial cell line: morphology, growth properties, and cytogenetic analysis of clonal cells.

The objectives of the present study were 1) to determine the heterogeneity of the MAC-T cell line; 2) to examine whether homogeneous clones could be derived from MAC-T cells; and 3) to examine cell morphology, cytoskeletal characteristics, size, colony-forming ability, growth characteristics, beta-casein production, response to oxytocin, and cytogenetic properties of the clones. Three clonal cells, designated CU-1, CU-2, and CU-3, were derived from MAC-T cells. CU-1 and CU-2 cells were morphologically homogeneous. CU-3 cells were heterogeneous and contained two distinct subtypes. All clones contained cytokeratin 14 and 18. CU-2 and CU-3 cells were 30 and 18% larger, respectively, than CU-1 cells. CU-1 cells did not grow in serum-free medium. Doubling times for MAC-T, CU-2, and CU-3 were 46, 48, and 78 h, respectively, in serum-free medium. MAC-T cells and clones constitutively expressed beta-casein in culture ranging from .1 to .3 micrograms/ml per 24 h. Cytogenetic analyses revealed Robertsonian translocations and isochromosomes in the clonal lines. We conclude that parental MAC-T cells are heterogeneous in morphology, growth, and cytogenetic characteristics.

Animals↗
PubMed1995

Effects of vitamin E on immune function of dairy cows.

The effect of vitamin E supplementation on the immune function of dairy cows was studied. Twelve cows were assigned to 1 of the 2 experimental groups: control (no vitamin E supplementation), and vitamin E-supplemented. Supplementation of vitamin E started 4 weeks before and continued up to 8 weeks after parturition and included oral supplementation of vitamin E at the rate of 3,000 IU/cow/d. In addition, the same group of cows received 1 injection of vitamin E (5,000 IU), 1 week prior to the expected date of parturition. Data indicated that blood neutrophils isolated from control cows produced twofold less (P < 0.05) superoxide anion after parturition, compared with the corresponding value before parturition. Furthermore, blood macrophages isolated from control cows produced 15 and 35% (P < 0.05) less interleukin 1 (IL-1) and major histocompatibility (MHC) class-II antigens, respectively, after parturition, compared with the corresponding values before parturition. These data, collectively, indicate that functions of blood macrophages and neutrophils are depressed during the early postpartum period in control cows. In contrast, there were no differences in superoxide anion production by blood neutrophils, or in IL-1 production, and MHC class-II antigen expression by blood macrophages before and after parturition in cows supplemented with vitamin E. There were no differences in lymphocyte proliferation, or IL-1 production and MHC class-II antigen expression by mammary macrophages when control and vitamin E-supplemented cows were compared. We conclude that vitamin E prevented suppression of blood neutrophil and macrophage function during the early postpatum period.

Analysis of Variance↗
PubMed1994

Distribution of plasminogen activator forms in fractions of goat milk.

Distribution of plasminogen activator forms in fractions of goat milk was examined. Raw milk was centrifuged to separate skim milk, cream, and a somatic cell pellet. Somatic cell extracts were obtained by sonication. Skim milk was centrifuged to separate milk serum and casein micelles. Activity of plasminogen activator was detected in casein, serum fractions, and in association with somatic cells. Plasminogen activator forms in milk casein had approximate molecular weights of 75,000, 50,000, and 30,000. The predominant forms of plasminogen activator in milk serum and in association with milk somatic cells had molecular weights of 30,000 and 50,000. Based on fibrin dependency and inhibition of activity in the presence of amiloride, the forms at 30,000 and 50,000 represent urokinase-plasminogen activator, and the form at 75,000 represents tissue-plasminogen activator.

Amiloride↗
PubMed1994

Production of various forms of plasminogen activator and plasminogen activator inhibitor by cultured mammary epithelial cells.

We examined amounts and types of plasminogen activator and plasminogen activator inhibitor produced by cultured bovine mammary epithelial cells. The MAC-T and two other mammary epithelial cell lines, MACT-UV1 and MACT-UV2 derived from the parental MAC-T cells by subcloning, were used as model systems. Cells were cultured in a medium free of serum and protein. Data showed that MACT-UV2 cells produced 6.2 and 17.2% more plasminogen activator than MACT-UV1 and parental MAC-T cells, respectively. Addition of amiloride, a specific urokinase-plasminogen activator inhibitor, dramatically decreased the activity in the culture medium of parental and subclonal lines, indicating that urokinase-plasminogen activator was present. Zymography revealed the presence of urokinase-plasminogen activator with an approximate molecular mass of 50,000 kDa in the culture medium of parental MAC-T cells. The culture medium of the subclonal lines contained urokinase-plasminogen activator and tissue-plasminogen activator with approximate molecular masses of 50,000 and 75,000 kDa, respectively. Complexes of both types of plasminogen activators with plasminogen activator-inhibitor-1 were detected in the culture medium of subclonal lines.

Animals↗
PubMed1994

Effect of once daily milking and concurrent somatotropin on mammary tight junction permeability and yield of cows.

Six pairs of monozygous Friesian twin cows during late lactation were used to assess the effect of once daily milking and bST treatment on yields and tight junction permeability of mammary epithelial cells. During the first 7 d, all cows were milked twice daily; on d 8 through d 21, all cows were milked once daily, but one cow of each twin pair was treated daily with 20 mg of bST on d 13 through 21; and, finally, during d 22 through 28, all cows were again milked twice daily. Once daily milking, a common management practice in New Zealand, resulted in a small (7%) but significant decrease in milk yield. Treatment with bST increased milk yield by 19%, thereby exceeding the milk yield loss from once daily milking. The integrity of mammary tight junctions was assessed indirectly by measuring concentrations of plasma lactose and milk BSA. Once daily milking temporarily disrupted tight junction integrity, based on a 4- to 5-fold increase in plasma lactose and a 42 to 55% increase in the concentrations of milk BSA. In the present study, bST did not affect the permeability of mammary tight junctions.

Animals↗
PubMed1994

Effect of milking frequency and somatotropin on the activity of plasminogen activator, plasminogen, and plasmin in bovine milk.

Six pairs of identical twin cows during late lactation (213 d) were used to study the effect of milking frequency (twice vs. once daily) and bST during once daily milking on the activity of plasminogen activator, plasminogen, and plasmin in milk. Less frequent milking increased the activity of plasminogen, plasmin, and plasminogen activator in milk. The ratio of plasminogen to plasmin, a measure that is independent of milk volume, decreased during less frequent milking, suggesting that at least part of the increase in activity of plasmin was due to the accelerated conversion of plasminogen to plasmin. Changes in the activity of plasminogen and plasmin in milk were positively correlated with increases in the concentrations of milk BSA and plasma lactose, both of which are indicators of disruption of tight junctions between mammary epithelial cells, indicating that paracellular leakage may have contributed to increased protease activity in milk during less frequent milking. No correlation existed between changes in plasminogen activator and indicators of tight junction disruption, suggesting that increased activity of plasminogen activator in milk was not due to leakage across the mammary epithelium, but rather to increased local production in the mammary gland. Administration of bST during once daily milking did not significantly affect milk protease activity.

Animals↗
PubMed1993

Enzymatic assay for the combined determination of plasmin plus plasminogen in milk: revisited.

The ability of beta-lactoglobulin variants A and B, alpha-lactalbumin, and BSA to inhibit plasmin plus plasminogen activity was examined. Data showed that beta-lactoglobulin A at concentrations of .2 and 1 mg/ml inhibited plasmin plus plasminogen activity by 18 and 54%. beta-Lactoglobulin B had no effect on plasmin plus plasminogen activity. At concentrations of .2 and 1 mg/ml, BSA inhibited plasmin plus plasminogen activity by 25 and 63%. alpha-Lactalbumin at concentrations of .2 and 1 mg/ml caused 1.9 and 20% inhibition of plasmin plus plasminogen activity. These data, collectively, suggest that existing methodology for measuring plasmin activity in milk serum underestimates real plasmin activity in milk. Underestimation is more pronounced in samples with high whey protein content (late lactation milk and milk obtained from mastitic quarters). To avoid this problem, we have modified the existing methodology. Our modification allows plasmin determination without interference from whey proteins and other plasmin inhibitors that are present in the serum fraction of bovine milk.

Aminocaproic Acid↗
PubMed1993

Effect of mammary-derived growth inhibitor on proliferation of MAC-T bovine mammary epithelial cells.

An established bovine mammary epithelial cell line (MAC-T) was used as a model to examine the effect of mammary-derived growth inhibitor on mammary epithelial cell proliferation. Prior to each proliferation assay, cells were synchronized in G1/G0 by culturing for 24 h without serum. Flow cytometry revealed that 90% of cells were in G1/G0, 4% in G2/M, and 6% in the S-phase after serum deprivation. Approximately 2 x 10(3) cells per well were seeded onto 24-well plates. Cells were incubated for 5 to 6 d with various amounts of mammary-derived growth inhibitor (0 to 100 ng/ml). Mammary-derived growth inhibitor and medium were changed daily. Mammary-derived growth inhibitor decreased mammary epithelial cell proliferation at .1 ng/ml. Synchronization of cells in G1/G0 was necessary for inhibition of cell proliferation. Cells not arrested by serum deprivation were not responsive to mammary derived-growth inhibitor. Inhibition of cell proliferation was transient and observed up to 96 h in culture. Mammary-derived growth inhibitor appears to act in vitro by inhibiting the resumption of stationary cell proliferation following starvation.

Animals↗
PubMed1992

Function of bovine mammary macrophages as antigen-presenting cells.

The ability of mammary macrophages treated with Staphylococcus aureus to induce antigen-specific T-cell proliferation was compared to that of the autologous blood monocytes. Induction of T-cell proliferation has been correlated with changes in major histocompatibility complex (MHC) class II antigen expression and interleukin 1 (IL-1) production by mammary macrophages and blood monocytes. The present study showed that both monocytes and mammary macrophages treated with S. aureus induced T-cell proliferation. However, there was a 3-fold decrease (P less than 0.05) in T-cell proliferation in macrophage cultures compared to those of blood monocytes, when these cells were treated with S. aureus. Mammary macrophages, the cells less effective in stimulating T-cell proliferation, expressed lower levels (2-fold) of MHC class II molecules and produced less IL-1 (3-fold) than blood monocytes. These data suggest that S. aureus may affect macrophage-T cell interaction by modulating the expression of MHC class II molecules and the synthesis of IL-1 by macrophages.

Animals↗
PubMed1992

Mammary-derived growth inhibitor in lactation and involution.

Presence of mammary-derived growth inhibitor (MDGI) in mammary tissue of lactating and involuted cows was investigated. Eighteen lactating, non-pregnant high-producing Holstein cows were randomly assigned to 3 experimental groups of 6 cows each. Cows of the first group were slaughtered while in lactation. Cows of the second group were slaughtered at 2-3 d, and the others at 4-8 d following sudden cessation of milking. Cessation of milking occurred at approximately 300 d in lactation. Western blot analysis revealed the presence of MDGI in the cytosolic and microsomal fractions of mammary tissue homogenates. High levels of MDGI were detected in mammary tissue obtained from lactating non-pregnant cows. A dramatic reduction in MDGI was observed in early involution (2-3 or 4-8 d following cessation of milking). These data suggest that a relationship exists between MDGI levels and the physiological status of the gland. Lack of MDGI may play a role during the processes of mammary involution and development prior to parturition.

Animals↗
PubMed1992

Effect of mastitis on plasminogen activator activity of milk somatic cells.

This study was conducted to examine the effects of mastitis and stage of lactation on plasminogen activator (PA) activity in milk somatic cells. An assay system, which measures the plasmin-mediated hydrolysis of the chromogenic substrate D-valyl-leucyl-lysine p-nitroanilide, was used to assess PA activity present within milk somatic cells. Milk cell associated PA activity was increased (P < 0.05) by 50% in the presence of fibrin fragments. This suggests that milk somatic cells contain tissue PA which, unlike urokinase PA, is preferentially activated in the presence of fibrin fragments. An increase of the milk somatic cell count from < 5 x 10(4) to > 10(6) cells/ml resulted in an 8-fold increase in PA activity per cell. Elevated levels of PA activity were associated with milk somatic cells isolated from mastitic quarters obtained from cows in early (< 4 months in lactation) or late lactation (> 8 months in lactation). We conclude that PA activity is increased during severe mastitic inflammation. Although the physiological function of this enzyme is as yet unclear, we propose that it may be involved in the conversion of plasminogen to plasmin, contributing to the higher levels of plasmin occurring in milk isolated from mastitic quarters.

Animals↗
PubMed1992

Receptor binding and growth-promoting activity of insulin-like growth factor-I in a bovine mammary epithelial cell line (MAC-T3).

Insulin-like growth factor-I (IGF-I) has been known to be mitogenic to a variety of cell types, although a growth-regulatory role for IGF-I on bovine mammary epithelial cells has not been fully investigated. In the present study, we examined the receptor binding of IGF-I and its effect on growth in a bovine mammary epithelial cell line (MAC-T3). Specific receptors for IGF-I were detected on cultured bovine mammary epithelial cells. Competitive binding revealed that half-maximal inhibition of 125I-labelled IGF-I binding by IGF-I was approximately 3 micrograms/l. Dissociation rate constant of the IGF-I receptor was 3.10 +/- 0.06 nmol/l (S.E.M.) with a receptor site concentration of 366 +/- 8 fmol/mg protein for the average of three experiments. IGF-I exerted a positive mitogenic effect on MAC-T3 cells according to both direct DNA assay and thymidine incorporation assay. Moreover, the mitogenic effect of IGF-I on MAC-T3 cells was enhanced by the addition of fetal calf serum in the culture media. The present results suggest that the bovine mammary epithelial cell line (MAC-T3) provides a useful model system with which to study the biological actions of insulin-like growth factors on the bovine mammary secretory tissue in vitro.

Animals↗