<p>Platelets are the smallest cells in the blood. They are formed in the bone-marrow and are important for the blood coagulation. Platelet tranfusions are given to patients propyhlactically before an operation but also in therapeutical purpose in connection with bleeding. It’s importent that the quality controls of the platelet concentrates are reliable.</p><p>ADVIA®60 (Bayer HealthCare) is a fully automated cell counter which uses impedance principle to count platelets in blood samples. The purpose of the study was to evaluate this new instrument for use in the blood bank of Akademiska Sjukhuset in Uppsala. The instrument was bought to be used for quality control of platelet concentrates. 30 samples from platelet concentrates, from both apheresis and from buffy coats, were analyzed 10 times each on ADVIA®60 and the coefficient of variation (CV) was calculated for each sample. CV variated from 0,8 % to 2,9 % which is good considering that according to Bayer HealthCare the CV should be < 5 % for thrombocytes on ADVIA®60. The instrument was newly calibrated when the study was performed. Platelet count can also be performed by immunological or optical principles.</p>

Myocardial infarction (MI) is often caused by a disruption of an atherosclerotic plaque with activation of coagulation, platelets and inflammation. The aims were; to investigate whether the oral direct thrombin inhibitor, ximelagatran affected markers for coagulation, platelet and inflammation in a patient cohort with recent MI and if the coagulation markers could identify patients with increased risk of new ischemic events; to evaluate some of the mechanisms involved in formation of platelet-monocyte aggregates (PMAs). In a biomarker substudy patients with recent MI were randomized to 24-60 mg of ximelagatran or placebo for six months. There was a persistent dose-independent reduction of coagulation markers (F1+2, D-dimer) by ximelagatran treatment. 60 % reduced their D-dimer levels after one week and that group had less ischemic events during treatment. There was an early increase of the platelet activation marker and ximelagatran in higher doses attenuated these increased levels. Both in vivo and in vitro the direct thrombin inhibitor diminished procoagulant activity and tissue factor (TF) presenting microparticles. In contrast, the inflammatory markers increased after six months of ximelagatran treatment. The PMA-levels were elevated for long-term after MI. In vitro thrombin inhibition diminished formation of PMAs. Formation of PMAs in stimulated whole blood was P-selectin dependent and induced TF expression through phosphorylation of the Src-family member Lyn in monocytes. Addition of an oral direct thrombin inhibitor reduces coagulation and platelet activation markers for long-term after a MI together with reduced procoagulant activity which may contribute to the clinical benefit of the drug. Early reduction of D-dimer levels seems to be suitable to identify patients with reduced risk of new ischemic events independent of antithrombotic treatment. Circulating PMAs persist after a MI connecting coagulation to inflammation. Within these aggregates P-selectin induces TF, the main initiator of coagulation, partly through phosphorylation of Lyn.

The subjects were 130 male prisoners in Swedish jails were examined by SCID and the diagnoses given in terms of DSM-IV. The most common mental disorder was drug abuse. On Axis II several personality disorders were diagnosed. Personality assessments were made by KSP. High scores were mainly found for e.g. impulsiveness, sensation seeking, aggression and low scores in socialisation. MAO assays were performed in 99 male criminal offenders and in 60 non-criminal volunteers. Offenders had lower MAO activity than controls also with the confounding factor smoking under control. It is proposed that platelet MAO is linked to personality traits, which can predispose for criminality. For testing the existence of combinations of vulnerability factors, a configuration frequency analysis (CFA) was used. The criteria which formed the basis for the subgrouping were; MAO activity below or above –0.5 SD of the mean (L and H), the presence or absence of an Axis I disorder (= drug abuse) (Y/N), the presence or absence of an Axis II disorder (Y/N), or the presence or absence of an Axis I and II disorder (Y/N). In this way eight subgroups were formed. Two significant "types" were found among the criminals: One was characterised by low platelet MAO activity, Cluster B personality diagnosis as well as Drug Abuse Disorder diagnosis (LYY); and the other by a pattern of normal platelet MAO activity, no Cluster B personality disorder, and no Drug Disorder diagnosis (HNN). Also two "antitypes", occurring less frequent than expected, were identified; LYN and LNY. Thus, the aggregation of certain risk factors in the same individual has been shown to contribute to the development of criminal behaviour. The subgroups HNN, LYN, LNY and LYY were then analysed for a variety of criminological factors. There was a difference in mean age between the subgroups, the HNN being lowest. Economical crimes were more common at an early criminal debut and crimes involving violence at an adult debut. The HNN subgroup had a lower number of crimes and times spent in jail than the other subgroups. More than 50% of the clients in all groups had previously been sentenced to Reformatory.

We have recently established that aberrant alkylacylglycerophosphocholine metabolism results in the increased tissue concentration of platelet activating factors (PAFs) in the temporal cortex of Alzheimer Disease (AD) patients and in TgCRND8 mice over-expressing mutant human amyloid precursor protein. PAF lipids activate a G-protein coupled receptor (PAFR) reported to be expressed by microglia and subsets of neurons in rat. It is not known whether this same expression pattern is recapitulated in mice however, as the expression has only been inferred by use of pharmacological PAFR antagonists, many of which impact on both PAFR-dependent and PAFR-independent signalling pathways. PAFR plays a role in long term potentiation (LTP) induction in rats. PAFR has also been implicated in behavioural indices of spatial learning and memory in rats. Contradictory reports using mice provide ambiguity regarding the role of PAFR in LTP induction in mice. To assess whether PAFR is expressed in murine neurons, I localized PAFR mRNA in wild-type C57BL/6 mice using PAFR KO mice as a negative control. I further showed that the loss of PAFR did not impair learning and memory although this assessment must be considered preliminary as the behavioural test employed was not optimized to detect changes in learning and memory of C57BL/6 mice over time adequately.Finally, I showed that the loss of PAFR in TgCRND8 mouse model of AD had no impact upon Aβ plaque number. My observations suggest that PAFR is restricted to microglial-like cells in mouse hippocampus and as such, it may not play a role in learning and memory.

Platelets are the smallest cells in the blood. They are formed in the bone-marrow and are important for the blood coagulation. Platelet tranfusions are given to patients propyhlactically before an operation but also in therapeutical purpose in connection with bleeding. It’s importent that the quality controls of the platelet concentrates are reliable. ADVIA®60 (Bayer HealthCare) is a fully automated cell counter which uses impedance principle to count platelets in blood samples. The purpose of the study was to evaluate this new instrument for use in the blood bank of Akademiska Sjukhuset in Uppsala. The instrument was bought to be used for quality control of platelet concentrates. 30 samples from platelet concentrates, from both apheresis and from buffy coats, were analyzed 10 times each on ADVIA®60 and the coefficient of variation (CV) was calculated for each sample. CV variated from 0,8 % to 2,9 % which is good considering that according to Bayer HealthCare the CV should be &lt; 5 % for thrombocytes on ADVIA®60. The instrument was newly calibrated when the study was performed. Platelet count can also be performed by immunological or optical principles.

We have recently established that aberrant alkylacylglycerophosphocholine metabolism results in the increased tissue concentration of platelet activating factors (PAFs) in the temporal cortex of Alzheimer Disease (AD) patients and in TgCRND8 mice over-expressing mutant human amyloid precursor protein. PAF lipids activate a G-protein coupled receptor (PAFR) reported to be expressed by microglia and subsets of neurons in rat. It is not known whether this same expression pattern is recapitulated in mice however, as the expression has only been inferred by use of pharmacological PAFR antagonists, many of which impact on both PAFR-dependent and PAFR-independent signalling pathways. PAFR plays a role in long term potentiation (LTP) induction in rats. PAFR has also been implicated in behavioural indices of spatial learning and memory in rats. Contradictory reports using mice provide ambiguity regarding the role of PAFR in LTP induction in mice. To assess whether PAFR is expressed in murine neurons, I localized PAFR mRNA in wild-type C57BL/6 mice using PAFR KO mice as a negative control. I further showed that the loss of PAFR did not impair learning and memory although this assessment must be considered preliminary as the behavioural test employed was not optimized to detect changes in learning and memory of C57BL/6 mice over time adequately.Finally, I showed that the loss of PAFR in TgCRND8 mouse model of AD had no impact upon Aβ plaque number. My observations suggest that PAFR is restricted to microglial-like cells in mouse hippocampus and as such, it may not play a role in learning and memory.

Las plaquetas (PLTs) son protagonistas en la reparación de las heridas, ya que contienen factores de crecimiento (GFs), los cuales producen quimiotaxis, proliferación y diferenciación celular, neovascularización y deposición de matriz extracelular (ECM). Se ha propuesto la utilización de concentrados de plaquetas (PCs) autólogos para acelerar la reparación de las heridas y estimular la capacidad de regeneración de los tejidos lesionados. En los capitulos III y IV de esta tesis se presentan dos estudios clínicos sobre el efecto de un concentrado plaquetario (PC) autólogo en caballos con patología músculo-esquelética grave. Este PC fue preparado por medio de una nueva técnica de doble centrifugación en tubo. Una caracterización celular y molecular de este PC se presenta en el cápitulo V de ésta tesis. Se evaluó el efecto de la inyección intra-articular del PC en 7 caballos con enfermedad articular grave clasificada así según el grado de cojera (DL) y la efusión sinovial (JE). El PC produjo una mejoría estadísticamente significativa del DL y JE (p<0.05). La mejoría más notable fue observada a los dos meses de finalizado el tratamiento y permaneció hasta 8 meses después (ver cápitulo III). En el capitulo IV se evaluó el efecto del PC en 7 caballos con lesiones tendinosas (tendonitis del tendón flexor digital superficial (SDFT) y ligamentosas (desmitis del ligamento suspensorio (DSL)). Todos los caballos mejoraron significativamente su DL y la respuesta a la prueba de flexión (p<0.05). Los registros ultrasonográficos mejoraron en los caballos con tendinopatías, pero no cambiaron en los que tenían desmopatías. Dos caballos con tendinitis retornaron con exito a su nivel de competición sin recidivar. Un caballo con tendinosis crónica recidivó. El resto de pacientes con desmopatías volvieron a su nivel de competición pre-lesión. Se obtuvieron un promedio de 250 ± 71.8 x 106 plaquetas, 8.68 ± 3.78 leucocitos x 106 y 12515 ± 2443 pg de TGF- ?1/ml de PC. No se observaron signos clínicos adversos asociados con el tratamiento. En el cápitulo V se realizó un análisis celular y molecular del PC usado clínicamente (PC¬C), sangre entera y 4 fracciones de PCs adicionales (PC-A, PC-B y PC-D), los cuales son obtenidos durante la elaboración del PC-C. El objetivo fue evaluar el método de obtención de este PC, en el que son necesarios dos periodos de centrifugación. Todas las muestras fueron analizadas mediante citometría de flujo y determinación de los niveles de TGF- ?1. Las concentraciones de plaquetas para PC-A, PC-B, PC-C y PC-D fueron un 45%, 44%, 71% y 21%, respectivamente más altas en comparación con cada PC y la sangre entera. Las concentraciones de TGF- ?1 para PC-A, PC-B, PC-C y PC-D fueron un 38%, 44%, 44% and 37%, respectivamente más altas en comparación con cada PC y la sangre entera. Se concluyó que el método empleado para concentrar plaquetas es valido para producir PCs con niveles potencialmente terapéuticos de TGF-?1. Los resultados obtenidos en esta tesis abren un nuevo y prometedor campo de investigación para conocer los efectos clínicos y moleculares de los PCs en caballos con enfermedad crónica músculo-esquelética. Los resultados que se puedan obtener en caballos serán de gran valor para conocer el uso potencial de los PCs autólogos en seres humanos con similares patologías. / Platelets (PLTs) play a central role in wound healing, since they contain growth factors (GFs), which produce chemotaxis, cellular proliferation and differentiation, neovascularization, and extra¬cellular matrix (ECM) deposition. The use of autologous platelet concentrates (PCs) has been proposed to accelerate wound repair and to stimulate the regenerative capacity of injured tissues. Two clinical studies about the effect of an autologous platelet concentrate (PC) in horses with severe musculoskeletal pathology are presented in chapters III and IV of this thesis. The PC was prepared by a novel double centrifugation tube method. A cellular and molecular characterization of this PC is presented in chapter V of this thesis.The effect of the intraarticular injection of this PC in 7 horses with severe joint disease was evaluated on the basis on degree of lameness (DL) and joint effusion (JE). When PCs were injected into the joint a statistically significant improvement in both DL and JE (p<0.05) were observed. The most marked improvement was maximun 2 months after the last injection and persisted up to 8 months later (see chapter III). The clinical effect of PC injection in 7 horses with soft tissue musculoskeletal injuries namely: SDFT tendinopathy and desmopathy of the susensory ligament (DSL) was also evaluated (see the chapter IV). All the horses presented a clinical and a statistical (p<0.05) decrease of the DL and the response to flexion test. Ultrasonic appearance improved in the horses with SDFT lesions, but remained the same in the horses with DSL. Two horses with acute SDFT tendinopathy returned successfully to competition level without reinjury. One horse with chronic SDFT tendinopathy relapsed. The rest of the horses with DSL returned successfully to competition level without reinjury. A mean of 250 ± 71.8 x 106 platelets, 8.68 ± 3.78 leucocytes x 106, and 12515 ± 2443 pg TGF-?1 were obtained per ml of the PC. No adverse reactions resulted from this treatment. A cellular and a molecular study of the PC clinically used in this thesis (PC-C),of whole blood and of three additional PCs (PC-A, PC-B, and PC-D) obtained during the PC-C preparation were performed (see the chapter V) to compare the single and the double centrifugation tube methods for concentrating equine platelets. Whole blood and the 4 PCs were analyzed using flow cytometry for cellular quantification and determination of TGF- ?1 in all the samples. Platelet concentrations for PC-A, PC-B, PC-C and PC-D were 45%, 44%, 71% and 21% higher, respectively, compared to the same values for citrated whole blood samples. TGF- ?1 concentrations for PC-A, PC-B, PC-C and PC-D were 38%, 44%, 44% and 37% higher, respectively, compared to citrated whole blood sample values. In conclusion, the single and double centrifugation tube methods are reliable methods for concentrating equine platelets and obtaining potentially therapeutic TGF- ?1 levels. The results obtained in this thesis open a new encouraging research field on clinical and molecular effects of PCs in equine chronic musculoskeletal pathology. The future potential results obtained in horses can be of key value to determine the potenetial use of autologous PCs in human beings with chronic musculoskeletal pathology.

Aβ oligomer-induced neurotoxicity has become an important area of therapeutic development in treating Alzheimer’s disease. Platelet-derived growth factor (PDGF) has been shown to be able to protect neurons against several neuronal insults such as ischemia and HIV1 toxin induced cytotoxicity. These neuroprotective effects correlate well with our previous results that demonstrate the neuroprotective effects of PDGF-BB, one of the PDGF receptor ligand subtypes, against NR2B containing NMDA receptor induced excitotoxicity, a possible underlying cause of Aβ oligomer induced synaptic dysfunction and neuronal death. This project examines the neuroprotective effect of PDGF-BB against Aβ1-42 oligomer induced cytotoxicity in both SH-SY5Y cells and primary hippocampal neurons. Cell viability was monitored by MTT assay and the affected signaling pathways were examined using pharmacological methods and Western blotting. The results demonstrated that Aβ1-42 oligomer elicited a dose-dependent toxicity with a sign of saturation at higher dosages, PDGF-BB failed to protect neurons against Aβ1-42 oligomer induced cytotoxicity. In contrast, Aβ1-42 oligomers strongly inhibit PDGF-BB induced mitogenesis in both SH-SY5Y cells and primary neurons. Further investigation using Western blotting to measure PDGF receptor expression and phosphorylation in SH-SY5Y cells showed that Aβ1-42 oligomer can inhibit PDGF-BB induced phosphorylation of PDGF β-receptor on Tyr1021, a site that is crucial for PLCγ mediated mitogenesis. These findings not only explained the poor neuroprotective effect elicited by PDGF-BB against Aβ1-42 oligomers, but also led to a novel hypothesis that Aβ1-42 oligomer may interfere with neurotrophic factor induced neuronal survival, either selectively or perhaps globally. Further exploration on this hypothesis will be able to shed light on this potentially novel mechanism of pathogenesis in Alzheimer’s disease.

No description available.

Arterial thrombus forms from the capture and accumulation of circulating platelets on a stenosis. As the thrombus grows, the lumen becomes further stenotic producing very high shear rates as the blood velocities increase through the narrowed cross-section. This study explores the molecular binding conditions that may occur under these pathologic shear conditions where circulating platelets must adhere quickly and with strong bonds. Platelets binding in an arterial stenosis of >75% are subject to drag forces exceeding 10,000 pN. This force can be balanced by 100 simultaneous GPIb-vWFA1 bonds of 100 pN each. The number and density of GPIb on platelets is sufficiently high; however, platelet capture under high shear would require the density of A1 receptors to be increased to over 416 per square micron. A computational model is used to determine platelet capture as a function of shear rate, surface receptor density, surface contact and kinetic binding rate. A1 density could be increased by a combination of vWF events of: i) plasma vWF attach to the thrombus surface and elongate under shear; ii) the elongated vWF strands create a net with 3-D pockets; and iii) additional vWF is released from mural platelets by activation under shear. With all three events, A1 density matches the existing high GPIbα densities to provide sufficient multivalency for capture at 100,000 s-1 with greater than 170 bonds per platelet. If the on-rate is greater than 108 M-1s-1, then a platelet could be captured within 15 microseconds, the amount of time available to form bonds before the platelet is swept away. This mechanism of platelet capture allows for the rapid platelet accumulation in atherothombosis seen clinically and in high shear experiments.