Platelet adherence to cancer cells promotes escape from innate immune surveillance in cancer metastasis.

The impacts of post­operative abdominal infectious complications increase hematogenous distant metastasis and result in poor long­term survival after curative resection. Even if curative resection can be performed, the presence of circulating tumor cells is affected. The liver, the most common site of metastases, is an important organ in innate immune surveillance. However, the molecular mechanisms of distant hematogenous metastasis are not yet fully known. Platelets are crucial components in the tumor microenvironment that function to promote tumor progression and metastasis. The purpose of this study was to identify the effect of platelets on escape from innate immune surveillance in post­operative abdominal infectious complications. Platelet adherence was assessed by co­culturing human pancreatic cancer cells including transforming growth factor (TGF­ß)­treated BxPC­3. CD44 isoform, transcription factors and epithelial­mesenchymal transition markers were examined using western blotting. We also assessed whether cancer cells surrounded by activated platelets could escape from innate immune surveillance, using infectious and non­infectious mouse models injected intraperitoneally with LPS. Platelets were found to preferentially adhere to mesenchymal cells rather than epithelial cells. BxPC­3 epithelial cells showed upregulation of CD44­variant and epithelial splicing regulatory protein 1 (ESRP­1) expression. However, Panc­1 mesenchymal cells and TGF­ß­treated BxPC­3 cells showed upregulation of CD44­standard and zinc finger E­box­binding homeobox 1 (ZEB­1) expression and a reduction in ESRP­1. In the non­infectious model, cancer cells were not found in the liver. In the infectious model, although epithelial cells without platelet adhesion were in an apoptotic state, mesenchymal cells showed many viable cancer cells surrounded by activated platelets. Cancer cells were suggested to have phenotypic plasticity through the switching of CD44 isoforms. Mesenchymal cells, which express CD44­standard, could escape from immune surveillance by becoming surrounded by adhered activated platelets. Therefore, it may be necessary to administer antiplatelet agents to prevent distant hematogenous metastasis when post­operative abdominal infectious complications occur.

Fungal Chitin Reduces Platelet Activation Mediated via TLR8 Stimulation.

Platelets play an important role in the innate immune response. During candidaemia, circulating fungal polysaccharides, including chitin, are released into the bloodstream and can interact with platelets and induce modulation of platelet activities. However, the role of circulating chitin in platelet modulation has not been investigated. The aims of the present study were to assess the effect of fungal chitin on activation, adhesion, aggregation and receptor expression of platelets and their impact on the host defense against Candida albicans. Platelets pre-treated with different concentrations of chitin (10-400 µg/mL) extracted from C. albicans were analyzed in terms of activation, Toll-like receptor (TLR) expression, aggregation and adhesion to C. albicans. Chitin treatment reduced platelet adhesion to C. albicans and neutrophils. P-selectin expression was significantly decreased in platelets challenged with chitin. Aggregation and intracellular Ca2+ influx were also decreased in platelets. TLR8 mRNA and proteins were expressed in platelets pre-treated with chitin when compared to untreated platelets. Overall, chitin purified from C. albicans reduced the adhesion, activation and aggregation of platelets mediated via TLR8 stimulation by decreasing intracellular Ca2+ influx and P-selectin expression.

Matrix metalloproteinase12 facilitated platelet activation by shedding carcinoembryonic antigen related cell adhesion molecule1.

Platelets express several MMPs that modulate their activation, which in turn regulates thrombosis, but the exact mechanism is unclear. This study evaluated the platelet expression of MMP12 and platelet activation by shedding CEACAM1 mediated by MMP12. Expression of MMP12 was measured by RT-PCR, Western blot (WB), and casein zymography in platelet from whole blood by gel filtration over plateletpheresis. The site of CEACAM1 cleavage by MMP12 was determined by high performance liquid chromatography (HPLC), mass spectrometry, WB and flow cytometry (FCM). Furthermore, the regulation of platelet aggregation, release and adhesion by MMP12-dependent shedding of platelet CEACAM1 was analyzed. We have observed that human platelets express MMP12. In addition, CEACAM1 as enzymatic substrates of MMP12 have also been found in this study. MMP12 can cleave the CEACAM1 exodomain at several sites and generated several short peptides. Among these fragments, one peptide, WYKG was identified, whose cutting sits were S66/W67 and A83/I84. We also found that MMP12 facilitated type I collagen induced platelet aggregation, adhesion and alpha granule secretion. Similarly, one short peptide, WYKG, facilitated type I collagen induced platelet alpha granule secretion. We conclude that platelet express MMP12 may facilitate platelet activation through shedding of CEACAM1.

Directed transport of neutrophil-derived extracellular vesicles enables platelet-mediated innate immune response.

The innate immune response to bacterial infections requires the interaction of neutrophils and platelets. Here, we show that a multistep reciprocal crosstalk exists between these two cell types, ultimately facilitating neutrophil influx into the lung to eliminate infections. Activated platelets adhere to intravascular neutrophils through P-selectin/P-selectin glycoprotein ligand-1 (PSGL-1)-mediated binding, a primary interaction that allows platelets glycoprotein Ibα (GPIbα)-induced generation of neutrophil-derived extracellular vesicles (EV). EV production is directed by exocytosis and allows shuttling of arachidonic acid into platelets. EVs are then specifically internalized into platelets in a Mac1-dependent fashion, and relocated into intracellular compartments enriched in cyclooxygenase1 (Cox1), an enzyme processing arachidonic acid to synthesize thromboxane A2 (TxA2). Finally, platelet-derived-TxA2 elicits a full neutrophil response by inducing the endothelial expression of ICAM-1, intravascular crawling, and extravasation. We conclude that critical substrate-enzyme pairs are compartmentalized in neutrophils and platelets during steady state limiting non-specific inflammation, but bacterial infection triggers regulated EV shuttling resulting in robust inflammation and pathogen clearance.

Platelets and vascular inflammation of the brain.

UNLABELLED: There is emerging evidence that platelets have an important role in inflammation beyond their involvement in hemostasis. Platelets can contribute to inflammatory reactions via crosstalk both with immune cells and endothelial cells. Inflamed vessels are characterized by the presence of activated endothelial cells. These activated endothelial cells upregulate receptors necessary for leukocyte recruitment, but also for the adhesion of platelets. Subsequently, immune cells can bind to platelets through adhesion receptors presented on the platelet surface, thus supporting leukocyte recruitment to the vessel wall. There are several neurological diseases associated with vascular inflammation including multiple sclerosis (MS) and stroke. Increased markers of platelet activation could be demonstrated in patients suffering from MS compared to healthy individuals. Reports from murine models indicate that platelets may be of importance for disease progression and severity by mediating leukocyte recruitment as one potential underlying mechanism. Blocking platelet function disease severity was considerably ameliorated. Moreover, processes of tissue remodelling may be influenced by platelet derived mediators. Whether a role of platelets for vascular inflammation can be extrapolated to further neurological diseases will have to be investigated in further in depth experimental and clinical trials. CONCLUSION: Platelets and platelet associated mechanisms may offer novel starting points to understand neurovascular diseases from a different point of view and to develop novel approaches to access the disease.

Interaction of von Willebrand factor with platelets and the vessel wall.

The initiation of thrombus formation at sites of vascular injury to secure haemostasis after tissue trauma requires the interaction of surface-exposed von Willebrand factor (VWF) with its primary platelet receptor, the glycoprotein (GP) Ib-IX-V complex. As an insoluble component of the extracellular matrix (ECM) of endothelial cells, VWF can directly initiate platelet adhesion. Circulating plasma VWF en-hances matrix VWF activity by binding to structures that become exposed to flowing blood, notably collagen type I and III in deeper layers of the vessel along with microfibrillar collagen type VI in the subendothelium. Moreover, plasma VWF is required to support platelet-to-platelet adhesion - i. e. aggregation - which promotes thrombus growth and consolidation. For these reasons, understanding how plasma VWF interaction with platelet receptors is regulated, particularly any distinctive features of GPIb binding to soluble as opposed to immobilized VWF, is of paramount importance in vascular biology. This brief review will highlight knowledge acquired and key problems that remain to be solved to elucidate fully the role of VWF in normal haemostasis and pathological thrombosis.

Frecuencia del síndrome de plaquetas pegajosas en pacientes con pérdidas fetales recurrentes / Frequency of sticky platelet syndrome in patients with recurrent fetal loss

El síndrome de las plaquetas pegajosas es un trastorno plaquetario autosómico dominante considerado como una de las causas más frecuentes de eventos trombóticos. Se cree que el defecto específico puede estar localizado en los receptores de la superficie plaquetaria y está caracterizado por un incremento anormal en la agregabilidad de las plaquetas con difosfato de adenosina, epinefrina, o ambos. En estudios realizados acerca de esta entidad se ha descrito una elevada incidencia, pero esta se ha calculado en pacientes que han tenido episodios trombóticos previos (enfermedad tromboembólica, infarto agudo del miocardio y accidente cerebrovascular), antecedentes familiares o tienen factores de riesgo para este tipo de eventos, mientras que su incidencia en una población de mujeres en edad fértil, con dos o más pérdidas de embarazo y sin antecedentes es desconocida. En el presente trabajo se estudiaron un total de 126 pacientes femeninas con al menos dos pérdidas de embarazo; de ellas, 27 resultaron positivas al estudio de hipersensibilidad plaquetaria con difosfato de adenosina y epinefrina mediante pruebas de agregometría, lo que representa el 21 por ciento de positividad en la población estudiada. Es significativo que la clase de síndrome de plaqueta pegajosa predominante fue de tipo II (hipersensibilidad con epinefrina). Finalmente, destacar que el seguimiento y tratamiento oportuno durante la gestación en las pacientes incluidas en el estudio ha permitido el nacimiento de 4 niños saludables de madres con síndrome de plaquetas pegajosas(AU)

The sticky platelet syndrome is an autosomal dominant platelet disorder considered one of the most frequent causes of thrombotic events. It is supposed that the specific fault can be localized to the platelet surface receptor and is characterized by an abnormal increase in aggregability adenosine diphosphate and/or epinephrine. In studies on this entity a high incidence has been described, but this was calculated in patients who have had previous thrombotic events (thromboembolic disease, myocardial infarction and stroke), family history or risk factors for this kind of events, while its incidence in a population of women of childbearing age with two or more pregnancy losses and no previous history is unknown.During our research a total of 126 female patients with at least two miscarriages were studied; 27 of them showed platelet hypersensitivity to adenosine diphosphate and epinephrine by testing aggregometry, which represents 21 percent positivity in the population studied. It is significant that the sticky platelet syndrome was predominantly of type II (hypersensitivity with epinephrine). In conclusion, we consider important to remark that with the monitoring and opportune treatment during pregnancy, four of the patients included in our study gave birth four healthy children despite the sticky platelet syndrome(AU)

Frecuencia del síndrome de plaquetas pegajosas en pacientes con pérdidas fetales recurrentes / Frequency of sticky platelet syndrome in patients with recurrent fetal loss

El síndrome de las plaquetas pegajosas es un trastorno plaquetario autosómico dominante considerado como una de las causas más frecuentes de eventos trombóticos. Se cree que el defecto específico puede estar localizado en los receptores de la superficie plaquetaria y está caracterizado por un incremento anormal en la agregabilidad de las plaquetas con difosfato de adenosina, epinefrina, o ambos. En estudios realizados acerca de esta entidad se ha descrito una elevada incidencia, pero esta se ha calculado en pacientes que han tenido episodios trombóticos previos (enfermedad tromboembólica, infarto agudo del miocardio y accidente cerebrovascular), antecedentes familiares o tienen factores de riesgo para este tipo de eventos, mientras que su incidencia en una población de mujeres en edad fértil, con dos o más pérdidas de embarazo y sin antecedentes es desconocida. En el presente trabajo se estudiaron un total de 126 pacientes femeninas con al menos dos pérdidas de embarazo; de ellas, 27 resultaron positivas al estudio de hipersensibilidad plaquetaria con difosfato de adenosina y epinefrina mediante pruebas de agregometría, lo que representa el 21 por ciento de positividad en la población estudiada. Es significativo que la clase de síndrome de plaqueta pegajosa predominante fue de tipo II (hipersensibilidad con epinefrina). Finalmente, destacar que el seguimiento y tratamiento oportuno durante la gestación en las pacientes incluidas en el estudio ha permitido el nacimiento de 4 niños saludables de madres con síndrome de plaquetas pegajosas...

The sticky platelet syndrome is an autosomal dominant platelet disorder considered one of the most frequent causes of thrombotic events. It is supposed that the specific fault can be localized to the platelet surface receptor and is characterized by an abnormal increase in aggregability adenosine diphosphate and/or epinephrine. In studies on this entity a high incidence has been described, but this was calculated in patients who have had previous thrombotic events (thromboembolic disease, myocardial infarction and stroke), family history or risk factors for this kind of events, while its incidence in a population of women of childbearing age with two or more pregnancy losses and no previous history is unknown.During our research a total of 126 female patients with at least two miscarriages were studied; 27 of them showed platelet hypersensitivity to adenosine diphosphate and epinephrine by testing aggregometry, which represents 21 percent positivity in the population studied. It is significant that the sticky platelet syndrome was predominantly of type II (hypersensitivity with epinephrine). In conclusion, we consider important to remark that with the monitoring and opportune treatment during pregnancy, four of the patients included in our study gave birth four healthy children despite the sticky platelet syndrome...

Nucleation of platelets with blood-borne pathogens on Kupffer cells precedes other innate immunity and contributes to bacterial clearance.

Through the use of intravital imaging of the liver, we demonstrate a collaborative role for platelets with Kupffer cells (KCs) in eradicating blood-borne bacterial infection. Under basal conditions, platelets, via the platelet-adhesion receptor GPIb, formed transient 'touch-and-go' interactions with von Willebrand factor (vWF) constitutively expressed on KCs. Bacteria such as Bacillus cereus and methicillin-resistant Staphylococcus aureus (MRSA) were rapidly caught by KCs and triggered platelets to switch from 'touch-and-go' adhesion to sustained GPIIb-mediated adhesion on the KC surface to encase the bacterium. Infected GPIbα-deficient mice had more endothelial and KC damage than did their wild-type counterparts, which led to more fluid leakage, substantial polycythemia and rapid mortality. Our study identifies a previously unknown surveillance mechanism by which platelets survey macrophages that rapidly converts to a critical host response to blood-borne bacteria.

The inflammatory chemokine CXC motif ligand 16 triggers platelet activation and adhesion via CXC motif receptor 6-dependent phosphatidylinositide 3-kinase/Akt signaling.

RATIONALE: The recently discovered chemokine CXC motif ligand 16 (CXCL16) is highly expressed in atherosclerotic lesions and is a potential pathogenic mediator in coronary artery disease. OBJECTIVE: The aim of this study was to test the role of CXCL16 on platelet activation and vascular adhesion, as well as the underlying mechanism and signaling pathway. METHODS AND RESULTS: Reverse-transcriptase polymerase chain reaction, Western blotting, confocal microscopy, and flow cytometry revealed that CXCL16-specific receptor, CXC motif receptor 6, is highly expressed in platelets. According to flow cytometry and confocal microscopy, stimulation of platelets with CXCL16 induced platelet degranulation, integrin α(IIb)ß(3) activation, and shape change. CXCL16 increased Akt phosphorylation (Thr(308)/Ser(473)), an effect abrogated by phosphatidylinositide 3-kinase inhibitors wortmannin (100 nmol/L) and LY294002 (25 µmol/L). The phosphatidylinositide 3-kinase inhibitors and Akt inhibitor SH-6 (20 µmol/L) further diminished CXCL16-induced platelet activation. CXCL16-mediated platelet degranulation, integrin α(IIb)ß(3) activation, and Akt phosphorylation were blunted in platelets lacking CXCL16-specific receptor CXC motif receptor 6. CXCL16-induced platelet activation was abrogated in Akt1- or Akt2-deficient platelets. CXCL16 enhanced platelet adhesion to endothelium in vitro after high arterial shear stress (2000(-s)) and to injured vascular wall in vivo after carotid ligation. CXCL16-induced stimulation of platelet adhesion again was prevented by phosphatidylinositide 3-kinase and Akt inhibitors. Apyrase and antagonists of platelet purinergic receptors P(2)Y(1) (MRS2179, 100 µmol/L) and especially P(2)Y(12) (Cangrelor, 10 µmol/L) blunted CXCL16-triggered platelet activation as well as CXCL16-induced platelet adhesion under high arterial shear stress in vitro and after carotid ligation in vivo. CONCLUSIONS: The inflammatory chemokine CXCL16 triggers platelet activation and adhesion via CXC motif receptor 6-dependent phosphatidylinositide 3-kinase/Akt signaling and paracrine activation, suggesting a decisive role for CXCL16 in linking vascular inflammation and thrombo-occlusive diseases.

Adiponectin-deficiency exaggerates sepsis-induced microvascular dysfunction in the mouse brain.

Obesity increases circulating cell-endothelial cell interactions; an early marker of inflammation in laboratory model of sepsis, but little is known about the effect of different adipokines. Adiponectin is an anti-inflammatory adipokine secreted by adipocytes. Adiponectin deficiency is implicated in exaggerated proinflammatory phenotype in both obesity and sepsis via increased proinflammatory cytokine expression. However the effect of adiponectin deficiency on circulating cell-endothelial cell interactions in polymicrobial sepsis is unknown. Furthermore although brain dysfunction in septic patients is a known predictor of death, the pathophysiology involved is unknown. In the current study, we examined the effects of adiponectin deficiency on leukocyte (LA) and platelet adhesion (PA) in cerebral microcirculation of septic mice. Adiponectin deficient (Adipoq(-/-): Adko) and background strain C57Bl/6 (wild type (WT)) mice were used. Sepsis was induced using cecal ligation and puncture (CLP). We studied LA and PA in the cerebral microcirculation using intravital fluorescent video microscopy (IVM), blood brain barrier (BBB) dysfunction using Evans Blue (EB) leakage method and E-selectin expression using dual radiolabeling technique in different WT and Adko mice with CLP. Adiponectin deficiency significantly exaggerated LA (WT-CLP:201 ± 17; Adko-CLP: ± 53 cells/mm(2); P < 0.05) and PA (WT-CLP:125 ± 17; Adko-CLP:188 ± 20 cells/mm(2); P < 0.05) in cerebral microcirculation, EB leakage (WT-CLP:10 ± 3.7; Adko-CLP:24 ± 4.3 ng/g × µl plasma; P < 0.05) and E-selectin expression (WT-CLP:0.06 ± 0.11; Adko-CLP:0.44 ± 0.053 ng/g; P < 0.05) in the brain tissue of the mice with CLP. Furthermore, E-selectin monoclonal antibody (mAb) treatment attenuated cell adhesion and BBB dysfunction of Adko-CLP mice. Adiponectin deficiency is associated with exaggerated leukocyte and PA in cerebral microcirculation of mice with CLP via modulation of E-selectin expression.

Antithrombotic drug candidate ALX-0081 shows superior preclinical efficacy and safety compared with currently marketed antiplatelet drugs.

Neutralizing the interaction of the platelet receptor gpIb with VWF is an attractive strategy to treat and prevent thrombotic complications. ALX-0081 is a bivalent Nanobody which specifically targets the gpIb-binding site of VWF and interacts avidly with VWF. Nanobodies are therapeutic proteins derived from naturally occurring heavy-chain-only Abs and combine a small molecular size with a high inherent stability. ALX-0081 exerts potent activity in vitro and in vivo. Perfusion experiments with blood from patients with acute coronary syndrome on standard antithrombotics demonstrated complete inhibition of platelet adhesion after addition of ALX-0081, while in the absence of ALX-0081 residual adhesion was observed. In a baboon efficacy and safety model measuring acute thrombosis and surgical bleeding, ALX-0081 showed a superior therapeutic window compared with marketed antithrombotics. Pharmacokinetic and biodistribution experiments demonstrated target-mediated clearance of ALX-0081, which leads to a self-regulating disposition behavior. In conclusion, these preclinical data demonstrate that ALX-0081 combines a high efficacy with an improved safety profile compared with currently marketed antithrombotics. ALX-0081 has entered clinical development.

Cytomegalovirus infection leads to microvascular dysfunction and exacerbates hypercholesterolemia-induced responses.

Cytomegalovirus (CMV) persistently infects more than 60% of the worldwide population. In immunocompetent hosts, it has been implicated in several diseases, including cardiovascular disease, possibly through the induction of inflammatory pathways. Cardiovascular risk factors promote an inflammatory phenotype in the microvasculature long before clinical disease is evident. This study determined whether CMV also impairs microvascular homeostasis and synergizes with hypercholesterolemia to exaggerate these responses. Intravital microscopy was used to assess endothelium-dependent and -independent arteriolar vasodilation and venular leukocyte and platelet adhesion in mice after injection with either mock inoculum or murine CMV (mCMV). Mice were fed a normal (ND) or high-cholesterol (HC) diet beginning at 5 weeks postinfection (p.i.), or a HC diet for the final 4 weeks of infection. mCMV-ND mice exhibited impaired endothelium-dependent vasodilation versus mock-ND at 9 and 12 weeks and endothelium-independent arteriolar dysfunction by 24 weeks. Transient mild leukocyte adhesion occurred in mCMV-ND venules at 7 and 21 weeks p.i. HC alone caused temporary arteriolar dysfunction and venular leukocyte and platelet recruitment, which were exaggerated and prolonged by mCMV infection. The time of introduction of HC after mCMV infection determined whether mCMV+HC led to worse venular inflammation than either factor alone. These findings reveal a proinflammatory influence of persistent mCMV on the microvasculature, and suggest that mCMV infection enhances microvasculature susceptibility to both inflammatory and thrombogenic responses caused by hypercholesterolemia.

Platelet adhesion and intracellular calcium levels in antigen-challenged rats.

There is considerable evidence that platelet activation occurs in allergic airways diseases. In this study we aimed to investigate platelet adhesion to immobilized fibrinogen and intracellular calcium levels in a rat model of allergic inflammation. Male Wistar rats were challenged with ovalbumin (OVA). At 30 min to 24h after OVA-challenge, assays of platelet adhesion to immobilized fibrinogen and intracellular calcium levels using fura 2-AM loaded platelets were performed. The serum levels of IgE were approximately 5-fold greater in OVA-sensitized rats. A marked eosinophil influx in bronchoalveolar lavage (BAL) fluid of OVA-challenged rats at 24h after OVA-challenge was also seen. OVA-challenge resulted in a marked thrombocytopenia, as observed within 12h after OVA-challenge. The agonists ADP (0.5-50 microM) and thrombin (30-100 mU/ml) concentration-dependently increased platelet adhesion to immobilized fibrinogen. At an early time after OVA-challenge (30 min), platelets exhibited greater platelet adhesion compared with the non-sensitized group, whereas at a late time (24h) they exhibited lower platelet adhesion to both agonists. Moreover, at 30 min after OVA-challenge, intracellular calcium levels to ADP (20 microM) and thrombin (100 mU/ml)-activated platelets were greater compared with non-challenged rats. As opposed, at 24h after OVA challenge, a lower intracellular calcium level to ADP- and thrombin-activated platelets was observed. In conclusion, OVA-challenge in rats promotes a biphasic response in platelet adhesion consisting of an increased adhesion and intracellular calcium levels at an early phase (30 min), which progress to a reduction in adhesion and intracellular calcium levels at a late time (24h) after antigen challenge.

Beyond cell capture: antibody conjugation improves hemocompatibility for vascular tissue engineering applications.

Antibody-conjugated surfaces are being studied for cardiovascular implant applications to capture endothelial progenitor cells and promote endothelialization. However, despite the large amount of literature on endothelial progenitor cell capture efficiency, little effort has been made to understand acute blood responses to the modified surfaces. We hypothesize that CD34 antibody conjugation passivates surfaces against procoagulatory events, and thus improves hemocompatibility. To test this hypothesis, we subjected the modified films to hemocompatibility tests to evaluate contact activation, platelet adhesion and activation, as well as whole blood clotting response to the films. Here, we demonstrate the alteration of blood responses due to polyacrylic acid (PAAc) engraftment and subsequent antibody conjugation on biaxially stretched polycaprolactone (PCL) films. Compared to PCL, PAAc-engrafted PCL (PCL-PAAc) and CD34-antibody-conjugated films (PCL-PAAC-CD34) resulted in a four- to ninefold (p < 0.001) reduced platelet activation. PCL-PAAc, however, resulted in an increased contact activation on thromboelastography, and a poorer blood compatibility index assay (43.4% +/- 2.3% vs. 60.9% +/- 2.5%, p < 0.05). PCL-PAAC-CD34, on the other hand, resulted in delayed clot formation (r = 19.3 +/- 1.5, k = 6.8 +/- 0.6 min) and reduced platelet adhesion and activation, and yielded the highest blood compatibility index score, indicating least thrombogenicity (69.3% +/- 3.2%). Our results suggest that CD34 antibody conjugation significantly improved the hemocompatibility of PAAc-conjugated PCL.

Staphylococcal superantigen-like 5 activates platelets and supports platelet adhesion under flow conditions, which involves glycoprotein Ibalpha and alpha IIb beta 3.

OBJECTIVES: Staphylococcal superantigen-like 5 (SSL5) is an exoprotein secreted by Staphylococcus aureus that has been shown to inhibit neutrophil rolling over activated endothelial cells via a direct interaction with P-selectin glycoprotein ligand 1 (PSGL-1). METHODS AND RESULTS: When purified recombinant SSL5 was added to washed platelets in an aggregometry set-up, complete and irreversible aggregation was observed. Proteolysis of the extracellular part of GPIb alpha or the addition of dRGDW abrogated platelet aggregation. When a mixture of isolated platelets and red cells was perfused over immobilized SSL5 at a shear rate of 300 s(-1), stable platelet aggregates were observed, and platelet deposition was substantially reduced after proteolysis of GPIb or after addition of dRGDW. SSL5 was shown to interact with glycocalicin, a soluble GPIb alpha fragment, and binding of SSL5 to platelets resulted in GPIb-mediated signal transduction as evidenced by translocation of 14-3-3 zeta. In addition, SSL5 was shown to interact with endothelial cell matrix (ECM) and this interaction enhanced aggregation of platelets from whole blood to this ECM. CONCLUSIONS: SSL5 activates and aggregates platelets in a GPIb alpha-dependent manner, which could be important in colonization of the vascular bed and evasion of the immune system by S. aureus.

[Immunity and lymphocytic-thrombocytic adhesion in community-aquired pneumonia].

AIM: To investigate immunity and lymphocytic-thrombocytic adhesion in pneumonia patients. MATERIAL AND METHODS: The examination of 59 male patients with pneumonia aged 18-21 years included calculation of total leucocyte count, lymphocyte subpopulations, relative and absolute count of lymphocytic-thrombocytic aggregates (LTA), measurement of IgA, IgM, IgG concentrations. RESULTS: Pneumonia patients have high count of leukocytes and low count of total and relative count of lymphocytes, low number of main lymphocytic subpopulations (CD4+, CD8+, CD16+, CD22+). Patients with uncomplicated pneumonia demonstrated high ability of lymphocytes for forming coaggregates with platelets. Patients with severe pneumonia complications may have either very high count of LTA (mean 403 +/- 45.6) or very low (mean 42.6 +/- 8.1). Percent of lymphocytes able to adhere to platelets (CD4+ with CD16+) in pneumonia is low. No correlation was found between LTA and the level of IgA, IgG, IgG. CONCLUSION: LTA is an objective indicator of cellular immunity status in pneumonia patients.

Platelet and leukocyte adhesion in the microvasculature at the cerebral surface immediately after subarachnoid hemorrhage.

OBJECTIVE: Pathophysiology after subarachnoid hemorrhage (SAH) caused by aneurysmal rupture has not been well examined. The purpose of this study was to observe platelet-leukocyte-endothelial cell interactions as indexes of inflammatory and prothrombogenic responses in the acute phase of SAH, using an in vivo cranial window method. METHODS: Subarachnoid hemorrhage was induced in C57Bl/6J mice by using the endovascular perforation method. Intravital microscopy was used to monitor the rolling and adhesion of platelets and leukocytes that were labeled with different fluorochromes. Regional cerebral blood flow was measured with laser Doppler flowmetry. The platelet-leukocyte-endothelial cell interactions were observed 30 minutes, 2 hours, and 8 hours after SAH. The effect of P-selectin antibody and apocynin, an inhibitor of nicotinamide adenine dinucleotide phosphate oxidase, on these responses was examined at 2 hours after SAH, and compared with a different SAH model in which autologous blood was injected into the foramen magna. RESULTS: SAH was accompanied by a 60% decrease in regional cerebral blood flow, whereas no changes in regional cerebral blood flow were observed on the contralateral side. SAH elicited time- and size-dependent increases in rolling and adherent platelets and leukocytes in cerebral venules. All of these interactions were attenuated by treatment with a P-selectin antibody or apocynin. There was no significant blood cell recruitment observed in the blood-injected SAH model. CONCLUSION: SAH at the skull base induced P-selectin- and oxygen radical-mediated platelet-leukocyte-endothelial cell interactions in venules at the cerebral surface. These early inflammatory and prothrombogenic responses may cause a whole-brain injury immediately after SAH.

Platelet adhesion under flow.

Platelet-adhesive mechanisms play a well-defined role in hemostasis and thrombosis, but evidence continues to emerge for a relevant contribution to other pathophysiological processes, including inflammation, immune-mediated responses to microbial and viral pathogens, and cancer metastasis. Hemostasis and thrombosis are related aspects of the response to vascular injury, but the former protects from bleeding after trauma, while the latter is a disease mechanism. In either situation, adhesive interactions mediated by specific membrane receptors support the initial attachment of single platelets to cellular and extracellular matrix constituents of the vessel wall and tissues. In the subsequent steps of thrombus growth and stabilization, adhesive interactions mediate platelet-to-platelet cohesion (i.e., aggregation) and anchoring to the fibrin clot. A key functional aspect of platelets is their ability to circulate in a quiescent state surveying the integrity of the inner vascular surface, coupled to a prompt reaction wherever alterations are detected. In many respects, therefore, platelet adhesion to vascular wall structures, to one another, or to other blood cells are facets of the same fundamental biological process. The adaptation of platelet-adhesive functions to the effects of blood flow is the main focus of this review.