Generous breastfeeding: an observational retrospective study of milk donor's characteristics in the province of Siena, Italy.

BACKGROUND: Breastfeeding is definitely one of the measures capable of improving not only children's but also mother's health. Human milk banks are institutions providing human milk to babies with limited access to breastfeeding for various reasons. STUDY DESIGN: The aim of this observational retrospective study was to examine principal characteristics of breast milk donors in the province of Siena and to identify variables influencing Milk Quantity (MQ) and Length of donation period (LD). METHODS: Information was extracted from medical records of Human Milk Bank of Siena, all women that donated their breast milk during the period from January 2010 until August 2017 were included. Examined variables were: age, place of birth (Italy/outside Italy), residence (Siena city/Siena province), education, profession, type of labor (preterm/in-term) and type of delivery (vaginal/cesarean section), gestational age, number of children, previous donations (blood, milk), quantity of donated milk and length of donation. RESULTS: A total of 304 donors were included: 75.7% of Italian nationality. The mean age was 32.4±5.2, Italian donors were older (33.5±5.0 vs. 28.7±4.2; Mann-Whitney; p<0.001). Socio-economic situation of Italian donors was better compared to non-Italian donors (chi-squared; p<0.005). Non-Italian donors had more probability to go through preterm labor (OR=3.68; p<0.001). Average length of donation was 2.7±1.8 months. Mean quantity of donated milk was 4.8±7.6l. From multiple linear regression, preterm birth (p<0.005) and length of donation (p<0.001) emerged as a variable that can predict higher quantity of donated milk. CONCLUSIONS: Preterm babies are usually recovering in a neonatology intensive therapy unit, with mothers staying close to them, which facilitates the whole donation process. Mothers of premature babies have a higher perception of the meaningfulness of donation and a need of adequate nutrition for fragile infants.

Midwives in England and in Italy: two realities compared.

BACKGROUND: Midwives have the responsibility to provide assistance to pregnant women for non-complicated deliveries. The aim of this study was to identify the distinctive features of midwives in Italy and in England in order to provide interesting comparisons. METHODS: From September to October 2015 we administered an online questionnaire to groups of Italian and English midwives. The questionnaire was composed by a part trying to collect the sociodemographic information and a second part to define the attitudes of care and the type of approach to birth and degree of attained professional autonomy. The questions were used to create a score for 5 macro areas: "Midwifery: who is the midwife?", "Education: how much does the midwife know?", "Responsibility: what does the midwife do?", "Ethics: how does the midwife act?", "Social acknowledgment: the midwife and the other people". Results were loaded in a database and analyzed using the using STATA. RESULTS: 183 Midwives (100 Italian and 83 English) completed the questionnaire. 92% of Italian respondents said to have a "Bachelor", 6% a "University Diploma" and only 2% a "Diploma". 80% of British midwives had a "Midwifery Degree" and 8% attended a "Midwifery Short Program", a degree course in Nursing with an additional specialization of a year and a half in Midwifery. 34% of the Italian claimed to have also a "1st level Master", 16% a "Full Degree" and the remaining 50% attended post degree training courses. Only 23% of the English sample had a "Master of Science" and the remaining 67% reported to have "other" general qualifications. 72% of Italian midwives declared to work and, in particular, in a "III level point of birth" (35%), "II level point of birth" (25%), "I level point of birth" (17%), or in "private health facilities" (4%), in "professional offices" (6%) or "counseling ambulatories" (5%). 46% of the English midwives who participated in the study worked in hospitals; 21% as employees of the NHS in territorial structures. 12% worked in birth centers and 11% had a private profession. There was a statistically significant difference between Italian and English Midwives, for all the macro areas (except for the first), with better results for English Midwives. CONCLUSION: English Midwives tend to consider their professional role and their professional skills more acknowledged and appreciated than Italian Midwives. Actually Italians tend to be not very satisfied. However, the Midwives of both countries feel very close to women and have the same conception of themselves and of their profession.

Differential effects of citrate versus PPACK anticoagulation on measured platelet inhibition by abciximab, eptifibatide and tirofiban.

BACKGROUND: High levels of glycoprotein (GP) IIb/IIIa receptor inhibition are required to prevent arterial thrombosis following percutaneous coronary intervention. Ex-vivo turbidometric platelet aggregation in citrate anticoagulated blood samples has been the primary method previously utilized to derive dose regimens for administering platelet GP IIb/IIIa inhibitors. Enhanced GP IIb/IIIa binding and inhibition of platelet aggregation for eptifibatide secondary to citrate induced reduction of ionized plasma calcium concentrations has been reported. METHODS/RESULTS: We evaluated the differential effects of citrate versus PPACK anticoagulation on turbidometric platelet inhibition in normal volunteers by eptifibatide, tirofiban or abciximab. The decrease in ionized calcium afforded by citrate was associated with enhanced in vitro platelet inhibition for all three GP IIb/IIIa inhibitors, including abciximab. The magnitude of citrate effect was greatest for eptifibatide. Both tirofiban and abciximab have similar citrate calcium chelation associated enhancement of measured platelet inhibition. CONCLUSION: Accurate assessment and comparison of platelet inhibition by GP IIb/IIIa inhibitors may require avoidance of calcium chelating anticoagulants.

Integrin tyrosine phosphorylation in platelet signaling.

The beta 3 integrin cytoplasmic tyrosine (ICY) motif of alpha IIb beta 3 becomes tyrosine phosphorylated during platelet aggregation, causing Shc and myosin to interact with the beta-integrin cytoplasmic domain. Platelets from mice lacking beta 3 ICY motif tyrosines formed defective aggregates and poorly retracted clots, establishing integrin tyrosine phosphorylation as a key mediator of beta 3-integrin signals.

Beta3 tyrosine phosphorylation in alphaIIbbeta3 (platelet membrane GP IIb-IIIa) outside-in integrin signaling.

The platelet integrin alphaIIbbeta3 not only binds fibrinogen and von Willebrand factor to mediate platelet aggregation and adhesion, it also serves as a signaling receptor. Platelet agonists such as ADP, thrombin and collagen induce "inside-out" signaling which activates the receptor function of alphaIIbbeta3 for soluble fibrinogen. Subsequent platelet aggregation leads to "outside-in" signaling, inducing platelet aggregate stabilization and triggering a variety of functions important to platelet physiology. This review focuses on the role of beta3 tyrosine phosphorylation in alphaIIbbeta3 outside-in signaling. Tyrosine phosphorylation of beta3 in platelets is a dynamic process which is initiated upon platelet aggregation and also by adhesion of platelets to immobilized fibrinogen. Tyrosine phosphorylation occurs on the beta3 integrin cytoplasmic tyrosine (ICY) domain, a conserved motif found in the beta subunits of several integrins. Beta3 ICY domain tyrosine phosphorylation induces the recruitment of two proteins to the cytoplasmic domains of alphaIIbbeta3: the cytoskeletal protein myosin, important to clot retraction; and the signaling adapter protein Shc, important to platelet stimulation, The critical role of beta3 tyrosine phosphorylation to platelet function was established by the diYF mouse, a novel strain which expresses an alphaIIbbeta3 in which the two beta3 ICY domain tyrosines have been mutated to phenylalanine. These mice are selectively impaired in outside-in alphaIIbbeta3 signaling, with defective aggregation and clot-retraction responses in vitro, and an in vivo bleeding defect which is characterized by a pronounced tendency to rebleed. Taken together, the data suggest that the beta3 tyrosine phosphorylation signaling mechanism is important to alphaIIbbeta3 function and might be applicable to a wide variety of integrin-mediated events.

Genetic and pharmacological analyses of Syk function in alphaIIbbeta3 signaling in platelets.

Agonists induce inside-out alphaIIbbeta3 signaling resulting in fibrinogen binding and platelet aggregation. These in turn trigger outside-in signaling resulting in further platelet stimulation. Because the Syk tyrosine kinase is activated during both phases of integrin signaling, we evaluated its role in alphaIIbbeta3 function in murine platelets rendered null for Syk by gene targeting and in human platelets incubated with piceatannol, a tyrosine kinase inhibitor reportedly selective for Syk. Both Syk null murine platelets and piceatannol-treated human platelets exhibited a partial, but statistically significant defect in activation of alphaIIbbeta3 by adenine diphosphate (ADP) +/- epinephrine as assessed by fibrinogen binding. Syk null platelets adhered normally to immobilized fibrinogen, and mice with these platelets exhibited normal tail bleeding times. In contrast, piceatannol treatment of human platelets completely inhibited platelet adhesion to immobilized fibrinogen. The discrepancy in extent of integrin dysfunction between murine and human platelet models may be due to lack of specificity of piceatannol, because this compound inhibited the activity of Src and FAK as well as Syk and also reduced tyrosine phosphorylation of multiple platelet proteins. These results provide genetic evidence that Syk plays a role in alphaIIbbeta3 signaling in platelets and pharmacological evidence that, although piceatannol also inhibits alphaIIbbeta3 signaling, it does so by inhibtion of multiple protein tyrosine kinases.

Tyrosine phosphorylation of the beta3 cytoplasmic domain mediates integrin-cytoskeletal interactions.

Tyrosine phosphorylation of the beta3 subunit of the major platelet integrin alphaIIb beta3 has been shown to occur during thrombin-induced platelet aggregation (1). We now show that a wide variety of platelet stimuli induced beta3 tyrosine phosphorylation, but that this phosphorylation occurred only following platelet aggregation. Several lines of evidence suggest that the beta3 cytoplasmic domain tyrosine residues and/or their phosphorylation function to mediate interactions between beta3 integrins and cytoskeletal proteins. First, phospho-beta3 was retained preferentially in a Triton X-100 insoluble cytoskeletal fraction of thrombin-aggregated platelets. Second, in vitro experiments show that the cytoskeletal protein, myosin, associated in a phosphotyrosine-dependent manner with a diphosphorylated peptide corresponding to residues 740-762 of beta3. Third, mutation of both tyrosines in the beta3 cytoplasmic domain to phenylalanines markedly reduced beta3-dependent fibrin clot retraction. Thus, our data indicate that platelet aggregation is both necessary and sufficient for beta3 tyrosine phosphorylation, and this phosphorylation results in the physical linkage of alphaIIb beta3 to the cytoskeleton. We hypothesize that this linkage may involve direct binding of the phosphorylated integrin to the contractile protein myosin in order to mediate transmission of force to the fibrin clot during the process of clot retraction.

Effect of Ca2+ on GP IIb-IIIa interactions with integrilin: enhanced GP IIb-IIIa binding and inhibition of platelet aggregation by reductions in the concentration of ionized calcium in plasma anticoagulated with citrate.

BACKGROUND: Integrilin (eptifibatide), a potent inhibitor of the fibrinogen binding function of GP IIb-llla, has been shown to reduce the thrombotic complications of angioplasty and of acute coronary syndromes. The present study was designed to determine whether the reduced Ca2+ concentrations in plasma anticoagulated with citrate affect Integrilin binding to GP IIb-IIIa and the ex vivo pharmacodynamic measurements for this drug. METHODS AND RESULTS: Lower concentrations of Integrilin were found to inhibit platelet aggregation in plasma anticoagulated with citrate (for ADP, mean+/-SD IC(50)=140+/-40 nmol/L, n=6; Ca2+ =40 to 50 micromol/L) than with PPACK (IC(50)=570+/-70 nmol/L, P<.0001, n=6; Ca2+ approximately 1 mmol/L). Chelation of Ca2+ with EDTA or citrate caused a similar degree of enhancement in the inhibitory activity of Integrilin. Measurements of D3 LIBS epitope expression showed that the enhanced inhibitory activity was caused by enhanced GP IIb-IIIa occupancy by Integrilin. Citrate anticoagulation decreased the amounts of Integrilin required to inhibit the binding of PAC1, a monoclonal antibody that mimics the GP IIb-IIIa binding activity of fibrinogen. Reduced Ca2+ also increased Integrilin inhibition of the binding of biotinylated fibrinogen to purified, immobilized GP IIb-IIIa. CONCLUSIONS: These data suggest that citrate anticoagulation removes Ca2+ from GP IIb-IIIa and enhances the apparent inhibitory activity of Integrilin. This finding indicates that the inhibitory activity of Integrilin is overestimated in blood samples collected with citrate, suggesting that it may be possible to achieve greater antithrombotic efficacy beyond that observed in clinical trials to date with Integrilin.

Outside-in integrin signal transduction. Alpha IIb beta 3-(GP IIb IIIa) tyrosine phosphorylation induced by platelet aggregation.

alpha IIb beta 3-(GP IIb IIIa) is the most abundant integrin expressed on platelets and plays a critical role in platelet aggregation and normal hemostasis. In response to platelet stimulation by agonists such as thrombin, alpha IIb beta 3 becomes a receptor for the adhesive proteins fibrinogen, von Willebrand factor, vitronectin, and fibronectin. Binding of extracellular matrix ligands allows the integrin to transmit a signal to the inside of the cell, but the exact mechanisms whereby integrins transduce these signals remains unclear. In this paper we demonstrate that the beta 3 subunit of alpha IIb beta 3 was phosphorylated on tyrosine residues in response to thrombin-induced platelet aggregation. However, tyrosine phosphorylation was not observed when platelets were stimulated by thrombin in the presence of an inhibitor of aggregation. Phosphotyrosine was only detected when platelets were solubilized under protein-denaturing conditions. A peptide corresponding to residues 740-762 of the beta 3 cytoplasmic domain was capable of binding the signaling proteins SHC and GRB2. GRB2 binding occurred only when both tyrosine residues (Tyr-747 and Tyr-759) were phosphorylated. SHC binding also occurred to a peptide monophosphorylated at Tyr-759. The data suggest that tyrosine phosphorylation of an integrin beta subunit may be important in initiating outside-in signaling cascades by inducing association of signaling components directly with the integrin.

[Comparison of Immunobead-Binding, TAT and SIT in the diagnosis of male antisperm autoimmunization]. / Confronto tra l'Immunobead-Binding, il TAT e il SIT nella diagnosi di autoimmunizzazione antinemasperma maschile.

The results obtained in this study show that the difference between local and general autoimmunization is less remarkable with the use of Immunobead-Binding tests, than with classical tests (TAT and SIT).