RESUMEN
Abstract Objective The present study aimed to compare the effects of intraarticular infiltration of platelet-rich plasma with those of hyaluronic acid infiltration in the treatment of patients with primary knee osteoarthritis. Methods A randomized clinical trial was conducted with 29 patients who received an intraarticular infiltration with hyaluronic acid (control group) or platelet-rich plasma. Clinical outcomes were assessed using the visual analog scale for pain and the Western Ontario and McMaster Universities Arthritis Index (WOMAC) questionnaire before and after the intervention. In addition, the posttreatment adverse effects were recorded. Categorical variables were analyzed using the chi-square and Fisher exact tests, whereas continuous variables were analyzed using the Student t test, analysis of variance, and the Wilcoxon test; all calculations were performed with the Stats package of the R software. Results An independent analysis of each group revealed a statistical difference within the first months, with improvement in the pain and function scores, but worsening on the 6th month after the procedure. There was no difference in the outcomes between the groups receiving hyaluronic acid or platelet-rich plasma. There was no serious adverse effect or allergic reaction during the entire follow-up period. Conclusion Intraarticular infiltration with hyaluronic acid or platelet-rich plasma in patients with primary knee gonarthrosis resulted in temporary improvement of functional symptoms and pain. There was no difference between interventions.
Resumo Objetivo Comparar o efeito da infiltração intraarticular do plasma rico em plaqueta com a do ácido hialurônico no tratamento de pacientes com osteoartrose primária de joelho. Métodos Realizou-se um ensaio clínico randomizado com 29 pacientes, sendo um grupo submetido à infiltração com ácido hialurônico (controle) e o outro com plasma rico em plaquetas. Os desfechos clínicos avaliados foram a escala visual analógica da dor; o questionário Western Ontario and McMaster Universities Arthritis Index (WOMAC), antes e depois da intervenção; e os efeitos adversos após as aplicações. Utilizou-se os testes do qui-quadrado e exato de Fisher para as variáveis categóricas, e o teste t de Student, análise de variância, e Wilcoxon para as variáveis contínuas, através do software R. Resultados A análise independente de cada grupo revelou uma diferença estatística nos meses iniciais, com melhora dos escores de dor e função; porém, com piora no 6° mês após o procedimento. Não houve diferença dos desfechos avaliados entre os grupos que foram submetidos à infiltração com ácido hialurônico ou com plasma rico em plaquetas. Não houve efeito adverso grave ou reação alérgica durante todo o seguimento. Conclusão A infiltração intraarticular com ácido hialurônico ou plasma rico em plaquetas nos joelhos dos pacientes com gonartrose primária apresentou melhora temporária dos sintomas de função e dor. Não houve diferença entre as duas intervenções.
Asunto(s)
Humanos , Osteoartritis/terapia , Benchmarking , Plasma Rico en Plaquetas/efectos de los fármacos , Ácido Hialurónico/uso terapéutico , Anestesia Local , Rodilla/patologíaRESUMEN
BACKGROUND: Platelet-rich plasma (PRP) has the potential to be used for bone regeneration. However, its effect on osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) and its effect on cell autophagy of hPDLSCs remain unknown. In this study, we investigated the effects of PRP on cell viability and osteogenic differentiation of hPDLSCs and the underlying molecular mechanisms. METHODS: hPDLSCs were isolated and identified by morphology and flow cytometry analysis. Next, thrombin-activated PRP was used to stimulate hPDLSCs. The MTT assay was used to analyze cell viability. Osteogenic differentiation was investigated using alkaline phosphatase (ALP) activity assay, alizarin red S (ARS) staining, and gene expression analysis of osteogenic markers. Expression of the autophagic proteins was determined using western blotting. RESULTS: Thrombin-activated PRP significantly enhanced cell viability, ALP activity, osteogenic-related mRNA levels and alizarin red-mineralization activity in hPDLSCs in a dose-dependent manner. Furthermore, activated PRP dose-dependently increased LC3-II/I ratio and the expression of SIRT1 and Beclin-1. PRP treatment also enhanced the autophagic flux. It was also demonstrated that the inhibition of SIRT1 using sirtinol or suppression of autophagy by 3-methyladenine (3-MA) abrogated PRP-induced viability and osteogenic differentiation of hPDLSCs. CONCLUSION: Our study suggested that thrombin-activated PRP accelerated the viability and osteogenic differentiation of hPDLSCs via SIRT1-mediated autophagy induction.
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Diferenciación Celular , Osteogénesis , Ligamento Periodontal/citología , Plasma Rico en Plaquetas/fisiología , Sirtuina 1/metabolismo , Células Madre/citología , Trombina/farmacología , Adulto , Autofagia , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Femenino , Hemostáticos/farmacología , Humanos , Masculino , Ligamento Periodontal/efectos de los fármacos , Ligamento Periodontal/metabolismo , Plasma Rico en Plaquetas/efectos de los fármacos , Sirtuina 1/genética , Células Madre/efectos de los fármacos , Células Madre/metabolismoRESUMEN
La enfermedad de la superficie ocular incluye a un grupo de patologías con diversas etiologías, síntomas y hallazgos clínicos que comparten la producción de reacción inflamatoria y daño de esta superficie. El uso de derivados hemáticos para el tratamiento de patologías de la superficie ocular se ha incrementado en el área de la oftalmología, ya que su composición es análoga a la de la lágrima natural. Con el objetivo de mostrar la terapia celular como una nueva disciplina científica a aplicar en nuestro medio, se realizó una búsqueda automatizada sobre el tema, teniendo en cuenta las publicaciones de los últimos 5 años. Se utilizó la plataforma Infomed, cuya información fue resumida para la elaboración del informe final, donde se expone que los colirios de hemoderivados proveen estrategias de tratamiento eficaces y seguras para pacientes con afecciones oftálmicas. El colirio de plasma rico en plaquetas ofrece una opción exitosa de tratamiento en numerosas afecciones de la superficie ocular. Sin embargo, estudios adicionales son necesarios para establecer la seguridad y la eficacia de este tipo de terapias(AU)
Ocular surface diseases are a group of conditions of different etiologies, symptoms and clinical findings with the common features of developing an inflammatory reaction and damaging the ocular surface. Use of blood-derived products for the treatment of ocular surface disorders has increased in ophthalmic care, since their composition is similar to that of natural tears. With the purpose of presenting cell therapy as a new scientific discipline that could be used in our environment, an automated search was conducted about the topic which included publications from the last five years. The search was performed on the Infomed platform, and the information obtained was summarized into a final report stating that blood-derived eye drops provide effective and safe treatment strategies for patients with ophthalmic conditions. Platelet-rich plasma eye drops are a potentially successful treatment option for many ocular surface disorders. However, further studies are required to establish the safety and effectiveness of this type of therapy(AU)
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Humanos , Soluciones Oftálmicas/efectos adversos , Medicamentos Hemoderivados , Plasma Rico en Plaquetas/efectos de los fármacos , Literatura de Revisión como Asunto , Bases de Datos Bibliográficas , Informe de InvestigaciónRESUMEN
PRP-Exos are nanoscale cup-shaped vesicles that carry a variety of proteins, mRNAs, microRNAs, and other bioactive substances. PRP-Exos can be formed through several induction pathways, which determine their molecular profiles and facilitate their tailormade participation in intercellular communication. Currently, little is known on how the PRP-Exos activation method influences the quality and quantity of PRP-Exos. The present study aims to observe and analyze the number, profile, and growth factors of PRP-Exos through TEM, Nanoflow, and WB after PRP activation and compare the difference in function of PRP-Exos on HUVECs, with different stimuli (calcium gluconate, thrombin, or both). We found that PRP activated with both thrombin and calcium gluconate harvested the highest concentration of exosomes [(7.16 ± 0.46) × 1010 particles/ml], compared to thrombin group [(4.87 ± 0.15) × 1010 particles/ml], calcium gluconate group [(5.85 ± 0.43) × 1010 particles/ml], or saline group [(7.52 ± 0.19) × 109 particles/ml], respectively (P < 0.05) via Nanoflow analysis. The WB analysis showed that cytokines (VEGF, PDGFBB, bFGF, TGF-ß) are differentially encapsulated in PRP-Exos, depending on the PRP stimulus, in which the mixture-PRP-Exos yielded the highest concentration of cytokines. In the function assay of PRP-Exos on HUVECs, the mixture-PRP-Exos promoted HUVECs proliferation, increased HUVECs migration, promoted the formation of vessel-like by HUVECs via the AKT ERK signal pathway more dramatically, compared with other groups. In summary, our studies showed that PRP activated by the mixture of calcium gluconate and thrombin harvested the best quality of exosomes which had the top biological functions. This study provides a protocol for selecting appropriate PRP activators to obtain high-quality exosomes for future applications.
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Exosomas/metabolismo , Plasma Rico en Plaquetas/efectos de los fármacos , Células Cultivadas , HumanosRESUMEN
Platelets mediate circulating endothelial progenitor cell (EPC) recruitment and maturation, participating in vascular repair, however the underlying mechanism(s) remain unclear. We investigated the effect of platelet-rich plasma (PRP) on the functionality of CD34+-derived late-outgrowth endothelial cells (OECs) in culture. Confluent OECs were coincubated with PRP under platelet aggregation (with adenosine diphosphate; ADP) and nonaggregation conditions, in the presence/absence of the reversible P2Y12 platelet receptor antagonist ticagrelor. Outgrowth endothelial cell activation was evaluated by determining prostacyclin (PGI2) and monocyte chemoattractant protein-1 (MCP-1) release and intercellular adhesion molecule-1 (ICAM-1) membrane expression. Similar experiments were performed using human umbilical vein endothelial cells (HUVECs). Platelet-rich plasma increased ICAM-1 expression and PGI2 and MCP-1 secretion compared with autologous platelet-poor plasma, whereas ADP-aggregated platelets in PRP did not exhibit any effect. Platelet-rich plasma pretreated with ticagrelor prior to activation with ADP increased all markers to a similar extent as PRP. Similar results were obtained using HUVECs. In conclusion, PRP induces OEC activation, a phenomenon not observed when platelets are aggregated with ADP. Platelet inhibition with ticagrelor restores the PRP capability to activate OECs. Since EPC activation is important for endothelial regeneration and angiogenesis, we suggest that agents inhibiting platelet aggregation, such as ticagrelor, may promote platelet-EPC interaction and EPC function.
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Plaquetas/metabolismo , Comunicación Celular , Células Progenitoras Endoteliales/metabolismo , Plasma Rico en Plaquetas/metabolismo , Antígenos CD34/metabolismo , Biomarcadores/metabolismo , Plaquetas/efectos de los fármacos , Comunicación Celular/efectos de los fármacos , Células Cultivadas , Quimiocina CCL2/metabolismo , Células Progenitoras Endoteliales/efectos de los fármacos , Epoprostenol/metabolismo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Molécula 1 de Adhesión Intercelular/metabolismo , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/farmacología , Plasma Rico en Plaquetas/efectos de los fármacos , Ticagrelor/farmacologíaRESUMEN
To be faced with an infected bone defect and the need to accelerate bone union while controlling infection is a welcome challenge for orthopedists. Platelet-rich plasma (PRP) has been applied in tissue defects given their composition of growth factors however the weak antibacterial effects have limited the use of PRP in the clinical setting. Therefore, the aim of this study was to explore the feasibility of using PRP in a local antibiotic delivery system (PADS) with the characteristics of promoting wound healing of bone infection. PADS was prepared with the addition of antibiotics or no antibiotics as control after PRP was prepared by a two-step centrifugation procedure. Antibacterial tests showed zones of inhibition produced by antibiotics were not significantly different with antibiotics combined with PRP. HPLC analysis demonstrated that about 60% of the total vancomycin (VAN) and ceftazidime (CAZ) dose were released within 10 min, then the release rate gradually decreased. However, 90% clindamycin was released within 10 min. Interestingly, above 10 times the minimum inhibitory concentration was presented after 72 h. Additionally, ELISA and morphology studies of PADS indicated that loaded antibiotics could reduce the PRP-released growth factor concentration and disturb the structure of platelet-fibrin beams and fibrin network in a dose-dependent manner. Fortunately, the lower dose of antibiotics maintained their anti-microbial effect, meanwhile growth factors released from PADS, the structure of platelet-fibrin beams, fibrin network remained unaffected. In addition, a patient experiencing infected bone defect receiving this PADS treatment achieved union within the 15-month follow-up. Therefore, this novel PADS approach might represent a potential therapy for patients who have sustained infected bone defects.
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Antibacterianos/uso terapéutico , Plasma Rico en Plaquetas/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacos , Adulto , Antibacterianos/farmacología , Humanos , MasculinoRESUMEN
Mesenchymal stem cells (MSCs) have been recently introduced in veterinary medicine as a potential therapeutic tool for several pathologies. The large-scale in vitro expansion needed to ensure the preparation of a suitable number of MSCs for clinical application usually requires the use of xenogeneic supplements like the fetal bovine serum (FBS). The substitution of FBS with species-specific supplements would improve the safety of implanted cells, reducing the risk of undesired immune responses following cell therapy. We have evaluated the effectiveness of canine adipose tissue-derived stromal vascular fraction (SVF) and MSCs (ADMSCs) expansion in the presence of canine blood-derived supplements. Cells were cultured on traditional plastic surface and inside a 3D environment derived from the jellification of different blood-derived products, i.e., platelet-poor plasma (PPP), platelet-rich plasma (PRP), or platelet lysate (PL). PPP, PRP, and PL can contribute to canine ADMSCs in vitro expansion. Both allogeneic and autologous PPP and PL can replace FBS for ADMSCs culture on a plastic surface, exhibiting either a similar (PPP) or a more effective (PL) stimulus to cell replication. Furthermore, the 3D environment based on homospecific blood-derived products polymerization provides a strong stimulus to ADMSCs replication, producing a higher number of cells in comparison to the plastic surface environment. Allogeneic or autologous blood products behave similarly. The work suggests that canine ADMSCs can be expanded in the absence of xenogeneic supplements, thus increasing the safety of cellular preparations. Furthermore, the 3D fibrin-based matrices could represent a simple, readily available environments for effective in vitro expansion of ADMSCs using allogeneic or autologous blood-products.
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Tejido Adiposo/metabolismo , Medios de Cultivo/metabolismo , Fibrina/metabolismo , Células Madre Mesenquimatosas/metabolismo , Plásticos/metabolismo , Xenobióticos/farmacología , Tejido Adiposo/efectos de los fármacos , Animales , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Técnicas de Cultivo de Célula/métodos , Perros , Células Madre Mesenquimatosas/efectos de los fármacos , Plasma Rico en Plaquetas/efectos de los fármacos , Plasma Rico en Plaquetas/metabolismo , Suero/metabolismoRESUMEN
The expansion of nanotechnology for drug delivery applications has raised questions regarding the safety of nanoparticles (NPs) due to their potential for interacting at molecular and cellular levels. Although polymeric NPs for drug delivery are formulated using FDA-approved polymers such as lactide- and glycolide-based polymers, their interactions with blood constituents, remain to be identified. The aim of this study was to determine the impact of size-selected Poly-lactide-co-glycolide-polyethylene glycol (PLGA-PEG) NPs on platelet activity. The NPs of 113, 321, and 585 nm sizes, were formulated and their effects at concentrations of 0-2.2 mg/mL on the activation and aggregation of platelet-rich plasma (PRP) were investigated. The results showed that NPs of 113 nm did not affect adenosine diphosphate (ADP)-induced platelet aggregation at any NP concentration studied. The NPs of 321 and 585 nm, at concentrations ≥0.25 mg/mL, reduced ADP-activated platelet aggregation. The platelet activation profile remained unchanged in the presence of investigated NPs. Confocal microscopy revealed that NPs were attached to or internalised by platelets in both resting and activated states, with no influence on platelet reactivity. The results indicate minimal risks of interference with platelet function for PLGA-PEG NPs and that these NPs can be explored as nanocarriers for targeted drug delivery to platelets.
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Plaquetas/efectos de los fármacos , Sistemas de Liberación de Medicamentos , Nanopartículas/administración & dosificación , Agregación Plaquetaria/efectos de los fármacos , Plasma Rico en Plaquetas/efectos de los fármacos , Polietilenglicoles/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , Plaquetas/fisiología , Portadores de Fármacos/química , Humanos , Nanopartículas/química , Agregación Plaquetaria/fisiología , Plasma Rico en Plaquetas/fisiologíaRESUMEN
Our objective in this study was to determine the biocompatibility and hemocompatibility of thin film metallic glass (TFMG) and its potential use in hypodermic needles for intramuscular or intravenous injection. Mouse and rabbit models were employed under approval from the Institutional Animal Care and Use Committee (n = 5/group, two groups in total for both animal models). Platelet-rich plasma (PRP) was collected from the whole blood of rabbits (ear vein) without anti-coagulant for use in in vitro coagulation tests. Histological analysis and optical microscopy were used to assess the endothelial structure of the inner lining of veins after being punctured with needles and detained for 3 days. Histological analysis of ear vein sections revealed that the extent of endothelial damage after puncturing with a TFMG-coated needle was 33% less than that produced by bare needles. Our results confirm that the deposition of a thin TFMG layer (e.g., Zr53Cu33Al9Ta5) on the surface of hypodermic needle can have remarkably clinical benefits, including anti-adhesion, reduced invasion, and minimal endothelial damage. Our results also confirm the good biocompatibility and hemocompatibility of the TFMG coatings.
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Materiales Biocompatibles Revestidos/efectos adversos , Ensayo de Materiales , Nanopartículas del Metal/efectos adversos , Agujas/efectos adversos , Células 3T3 , Animales , Coagulación Sanguínea/efectos de los fármacos , Adhesión Celular/efectos de los fármacos , Materiales Biocompatibles Revestidos/química , Endotelio Vascular/efectos de los fármacos , Diseño de Equipo , Vidrio/química , Inyecciones Intramusculares/instrumentación , Inyecciones Intravenosas/instrumentación , Masculino , Nanopartículas del Metal/química , Ratones , Modelos Animales , Plasma Rico en Plaquetas/efectos de los fármacos , Conejos , Propiedades de SuperficieRESUMEN
A não união óssea é uma complicação ortopédica que ocorre normalmente devido à instabilidade da fratura em decorrência de uma escolha de fixação inadequada ou inapropriada, suprimento sanguíneo deficiente, osteomielite e afastamento excessivo dos fragmentos; sendo mais comumente em cães de raças pequenas e miniatura; e de maior ocorrência em regiões distais de rádio, ulna, tíbia e fíbula. Este trabalho relata a utilização da associação entre Hidroxiapatita e Plasma Rico em Plaqueta no tratamento de uma não união óssea de rádio e ulna e esclarece os benefícios desses biomateriais no processo de regeneração do tecido ósseo. O presente trabalho tem por objetivo relatar o caso de um canino com não união óssea de rádio/ulna devido ao alinhamento inadequado dos fragmentos ósseos no tratamento conservador com bandagem, proporcionando instabilidade do foco da fratura. O tratamento cirúrgico consistiu na colocação de uma placa óssea e enxertia com Hidroxiapatita e Plasma Rico em Plaqueta para melhor regeneração óssea. A utilização de tais biomateriais no tratamento da não união foi benéfica para a formação do calo ósseo primário, não produzindo efeitos adversos para o paciente. A partir desse resultado pode-se concluir que, a utilização desses biomateriais e enxertia precisa ser mais bem estudada e aprimorada na reparação óssea de uma não união, visto que, a aplicabilidade dessa associação mostrou-se um método eficiente, não apresentando sinais de infecção e nem evidência de rejeição.(AU)
Bone nonunion is an orthopedic complication that usually occurs due to fracture instability as a result of an inadequate or inappropriate choice of fixation, deficient blood supply, osteomyelitis, and excessive removal of fragments; which is more commonly seen in small and miniature breeds; and more frequent in the distal regions of radius, ulna, tibia, and fibula. This paper reports on the use of the association between Hydroxyapatite and Platelet-Rich Plasma in the treatment of a radius and ulna nonunion and clarifies the benefits of these biomaterials in the bone tissue regeneration process. This study reports the case of a dog presenting nonunion of radius and ulna bone due to inadequate alignment of bone fragments in a conservative treatment with bandage, providing instability of the fracture focus. The surgical treatment consisted of placing a bone plate and grafting with Hydroxyapatite and Platelet-Rich Plasma for better bone regeneration. The use of such biomaterials in the treatment of nonunion injuries was beneficial for the formation of the primary bone callus, without producing adverse effects for the patient. From this result, it can be concluded that the use of these biomaterials and grafting needs to be further studied and improved for use in bone repair of nonunion cases, since the applicability of this association proved to be an efficient method, with no signs of infection or evidence of rejection.(AU)
Ausencia de unión ósea es uma complicación ortopédica que normalmente ocurre debido a la instabilidade de la fractura como resultado de uma elección de fijación inadecuada o inapropriada, aporte sanguíneo deficiente, osteomielitis y remoción excessiva de fragmentos; que se observa com mayor frecuencia en perros de razas pequeñas y miniaturas; y más frecuente em regiones distales de radio, cúbito, tíbia y peroné. Este artículo informa sobre el uso de la asociación de Hidroxiapatita y Plasma rico en plaqueta en el tratamiento de ausencia de unión del radio e el cúbito, y aclara los benefícios de esos biomateriales en el processo de regeneración del tejido ósseo. Esa investigación ha tenido como objetivo reportar el caso de un perro sin unión de radio / cúbito por alineación inadecuada de fragmentos óseos en tratamiento conservador con vendaje, proporcionando inestabilidad del foco de la fractura. El tratamiento quirúrgico consistió en la colocación de una placa ósea e injerto con Hidroxiapatita y Plasma rico en plaqueta para uma mejor regeneración ósea. El uso de tales biomateriales en el tratamiento de ausencia de unión ha sido beneficioso para la formación del callo óseo primario, sin producir efectos adversos para el paciente. A partir de ese resultado se puede concluir que, el uso de esos biomateriales e injertos necesitan ser mejor estudiado y mejorado en la reparación de ausencia ósea, ya que la aplicabilidad de esa asociación demostró ser um método eficaz, sin presentar signos de infección y evidencia de rechazo.(AU)
Asunto(s)
Animales , Regeneración Ósea/efectos de los fármacos , Perros/lesiones , Fracturas Óseas/veterinaria , Plasma Rico en Plaquetas/efectos de los fármacos , Fracturas de la Muñeca/tratamiento farmacológico , Durapatita/químicaRESUMEN
Rhizoma Bletillae, the tubes of Bletilla striata, has been traditionally used in China as a hemostatic agent. In preliminary studies, the major active fraction responsible for its hemostatic effect have been confirmed to be Rhizoma Bletillae polysaccharide (RBp), but the hemostatic mechanism of action of RBp is still unknown.The main aim of this study was to clarify its mechanism of hemostatic effect. RBp was prepared by 80 % ethanol precipitation of the water extract of Rhizoma Bletillae followed by the Sevag method to remove proteins. The average molecular weight (Mw) of the crude RBp maintained at a range of 30.06-200 KDa. The hemostatic effects of RBp were evaluated by testing its effect on the platelet aggregation of rat platelet-rich plasma (PRP). PRP was dealt with different concentrations of RBp and platelet aggregation was measured by the turbidimetric method. The hemostatic mechanism of RBp was investigated by examining its effect on platelet shape, platelet secretion, and activation of related receptors (P2Y1, P2Y12 and TXA2) by electron microscopy and the turbidimetric method. RBp significantly enhanced the platelet aggregations at concentrations of 50-200 mg/L in a concentration-dependent manner. The inhibitory rate of platelet aggregation was significantly increased by apyrase and Ro31-8220 in a concentration-dependent manner, while RBp-induced platelet aggregation was completely inhibited by P2Y1, P2Y12 and the PKC receptor antagonists. However, the aggregation was not sensitive to TXA2. RBp, the active ingredients of Rhizoma Bletillae responsible for its hemostatic effect, could significantly accelerate the platelet aggregation and shape change. The hemostatic mechanism may involve activation of the P2Y1, P2Y12, and PKC receptors in the adenosine diphosphate (ADP) receptor signaling pathway.
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Hemostáticos/farmacología , Plasma Rico en Plaquetas/efectos de los fármacos , Polisacáridos/farmacología , Receptores Purinérgicos P2/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Animales , Relación Dosis-Respuesta a Droga , Técnicas In Vitro , Peso Molecular , Extractos Vegetales/farmacología , Tubérculos de la Planta/química , Agregación Plaquetaria/efectos de los fármacos , Proteína Quinasa C/efectos de los fármacos , Ratas , Receptores Purinérgicos P2Y1/efectos de los fármacos , Receptores Purinérgicos P2Y12/efectos de los fármacosRESUMEN
BACKGROUND: This study evaluated blood components processed by the platelet rich plasma (PRP) method from fresh whole blood (FWB) treated with a pathogen reduction technology (PRT). The effects of storage temperature on PRT treated platelet concentrates (PCs) were also examined. STUDY DESIGN AND METHODS: PRT was performed using riboflavin and ultraviolet light on FWB in citrate phosphate dextrose anticoagulant. Following PRT, red blood cells (RBCs), PCs, and plasma for fresh frozen plasma (FFP), were isolated by sequential centrifugation. RBCs were stored at 4°C, FFP at -80°C, and PC at 22°C or at 4°C. Components were assayed throughout their storage times for blood gases, chemistry and CBC, hemostatic function as well as platelet (PLT) and RBC integrity. RESULTS: Component processing following PRT resulted in a significant drop in platelet recovery. Most PRT-PC bags fell below AABB guidelines for platelet count. PRT-PC also showed a decrease in clot strength and decreased aggregometry response. Platelet caspases were activated by PRT. Storage at 4°C improved platelet function. In PRT-FFP, prothrombin time and partial thromboplastin time (PT and aPTT) were prolonged; factors V, VII, VIII, and XI, protein C, and fibrinogen were significantly decreased. Free hemoglobin was elevated two-fold in PRT-RBC. CONCLUSION: Blood components isolated by the PRP method from PRT-treated WB result in a high percentage of PC that fail to meet AABB guidelines. FFP also shows diminished coagulation capacity. However, PRT-RBC are comparable to control-RBC. PRT-WB retains acceptable hemostatic function but alternatives to the PRP method of component separation may be more suitable.
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Eritrocitos/metabolismo , Plasma/metabolismo , Plasma Rico en Plaquetas/metabolismo , Anticoagulantes/farmacología , Factores de Coagulación Sanguínea/metabolismo , Análisis de los Gases de la Sangre , Conservación de la Sangre , Eritrocitos/efectos de los fármacos , Eritrocitos/efectos de la radiación , Hemoglobinas/análisis , Humanos , Tiempo de Tromboplastina Parcial , Recuento de Plaquetas , Pruebas de Función Plaquetaria , Plasma Rico en Plaquetas/efectos de los fármacos , Plasma Rico en Plaquetas/efectos de la radiación , Tiempo de Protrombina , Riboflavina/farmacología , Rayos UltravioletaRESUMEN
Background. Studies of platelet aggregation (PA) in essential thrombocythemia (ET) reported contrasting results, likely due to differences in analytical conditions.Objective. We investigated platelet aggregation using different techniques and analytical conditions.Patients and Methods. PA was studied by light-transmission aggregometry (LTA) in platelet-rich plasma (PRP) and impedance aggregometry in PRP and whole blood (WB). ADP, collagen, thrombin receptor activating peptide (TRAP-14) and adrenaline were used as agonists. Since ET patients (n = 41) were on treatment with aspirin (100 mg/d), healthy controls (n = 29) were given aspirin (100 mg/d) for 5 days before testing: therefore, thromboxane A2-independent PA was tested in all subjects. Blood samples were collected in citrate (C) [low Ca2+] or lepirudin (L) [physiological Ca2+]; platelet count was adjusted to 250 x 109/L in a set of C-PRP (adjusted C-PRP) and left unmodified in the other samples.Results. Results of PA in 17 ET patients who were poor responders to aspirin (high serum thromboxane B2 levels) were not included in the analysis. With LTA, PA in ET was lower than in controls in adjusted C-PRP and normal in native C-PRP and L-PRP. With impedance aggregometry, PA in L-PRP and L-WB tended to be higher in ET than in controls. Platelet serotonin and ADP contents were reduced in ET. The percentages of circulating platelets expressing P-selectin and platelet-leukocyte hetero-aggregates were higher in ET.Conclusions. Analytical conditions dramatically affect in vitro PA of ET patients, which appears defective under the least physiological conditions and normal/supranormal under conditions that are closer to the physiological.
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Plaquetas/fisiología , Pruebas de Función Plaquetaria/métodos , Plasma Rico en Plaquetas , Trombocitemia Esencial/sangre , Nucleótidos de Adenina/sangre , Adulto , Anciano , Anciano de 80 o más Años , Aspirina , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Ácido Cítrico/farmacología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Selectina-P/sangre , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/farmacología , Recuento de Plaquetas , Plasma Rico en Plaquetas/efectos de los fármacos , Serotonina/sangre , Trombocitemia Esencial/tratamiento farmacológico , Trombocitemia Esencial/patología , Adulto JovenRESUMEN
Losartan and honokiol are small molecules which have been described to inhibit aggregation of platelets by collagen. Losartan has been proposed to block clustering of GPVI but not to affect binding of collagen. Honokiol has been reported to bind directly to GPVI but only at a concentration that is three orders of magnitude higher than that needed for inhibition of aggregation. The mechanism of action of both inhibitors is so far unclear. In the present study, we confirm the inhibitory effects of both agents on platelet aggregation by collagen and show that both also block the aggregation induced by the activation of CLEC-2 or the low affinity immune receptor FcγRIIa at similar concentrations. For GPVI and CLEC-2, this inhibition is associated with a reduction in protein tyrosine phosphorylation of multiple proteins including Syk. In contrast, on a collagen surface, spreading of platelets and clustering of GPVI (measured by single molecule localisation microscopy) was not altered by losartan or honokiol. Furthermore, in flow whole-blood, both inhibitors suppressed the formation of multi-layered platelet thrombi at arteriolar shear rates at concentrations that hardly affect collagen-induced platelet aggregation in platelet rich plasma. Together, these results demonstrate that losartan and honokiol have multiple effects on platelets which should be considered in the use of these compounds as anti-platelet agents.
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Compuestos de Bifenilo/farmacología , Lignanos/farmacología , Losartán/farmacología , Inhibidores de Agregación Plaquetaria/farmacología , Agregación Plaquetaria/efectos de los fármacos , Glicoproteínas de Membrana Plaquetaria/antagonistas & inhibidores , Plaquetas/metabolismo , Colágeno/farmacología , Humanos , Lectinas Tipo C/metabolismo , Glicoproteínas de Membrana/metabolismo , Fosforilación , Glicoproteínas de Membrana Plaquetaria/metabolismo , Plasma Rico en Plaquetas/efectos de los fármacos , Plasma Rico en Plaquetas/enzimología , Plasma Rico en Plaquetas/metabolismo , Receptores de IgG/metabolismo , Quinasa Syk/metabolismo , TrombosisRESUMEN
Previous in vitro studies suggest a direct relevance for the peptide-free lipid fraction (LF) of platelet-rich plasma (PRP) in biological mechanisms related to wound healing. However, there are no scientific reports to date on the wound healing activities of this lipid component in vivo. Thus, the present study provides a scientific evaluation for the wound healing potential of the lipid portion of the activated PRP. For the wound healing activity assessment, in vivo full-thickness excisional wounds were created on the dorsal skin of Sprague-Dawley rats. Lipid extract from pooled PRP was applied topically to the wounds on 0, 3, and 7 days after injury. Histological assessment of epidermal and dermal regeneration, granulation tissue thickness and angiogenesis by Sirius red and Masson's trichrome staining, in addition to immunohistochemical staining for transforming growth factor beta-1 (TGF-ß1), collagen type I (COL I), and collagen type III (COL III) were performed on skin biopsies at 3, 7 and 14 days. The total histological scores of the LF group were significantly higher than the 25% dimethylsulfoxide-control group. According to the immunohistochemical staining, the observed expression changes for TGF-ß1, COL I and III at 3, 7, and 14 days after wounding were significantly better in the study group than the control group. Furthermore, COL I/III ratio in the lipid extract-treated group at day 14 was much higher than that of the control group. Meanwhile, analysis of the data also indicated that the LF has less positive effects on all evaluated parameters than PRP. From the present data, it could be concluded that the peptide-free LF of PRP has potent wound healing capacity in vivo for cutaneous wounds, although not as much as that of PRP. Strengthening our understanding of the wound healing potential of lipid components of PRP and platelet-derived lipid factors may provide new avenues for improving the healing process of a wound with elevated protease activity.
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Lípidos/farmacología , Plasma Rico en Plaquetas/metabolismo , Piel/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacos , Animales , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Femenino , Lípidos/sangre , Lípidos/aislamiento & purificación , Plasma Rico en Plaquetas/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Piel/citología , Piel/lesiones , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
BACKGROUND: Light transmission aggregometry (LTA) is the gold standard for platelet function assessment. The automated coagulation analyzer from Sysmex that performs LTA offers the advantage of being a walk-away technology. Recently, a new parameter "ADP-induced platelet aggregation level (APAL)" was developed to support the interpretation of results. APAL is calculated as a score from 0.0 to 10.0 based on platelet aggregation patterns with 1 and 10 µM adenosine diphosphate (ADP). Here, the basic performance of the newly developed APAL system and comparison with the maximum aggregation rate of ADP (ADP-MA) was evaluated. METHODS: The within-run precision was calculated by conducting five replicate analyses of the platelet-rich plasma (PRP) from healthy volunteers and 0.05 µM of cangrelor-spiked PRP. Cangrelor is a P2Y12 inhibitor that does not require liver CYP activation. The reference interval was calculated from the results of 67 healthy volunteers. The effect of the antiplatelet P2Y12 agent was evaluated using several concentrations of cangrelor. A comparative study was performed using 103 PRP samples with different levels of aggregation. Each test was analyzed with both APAL and ADP-MA. RESULTS: The percentage coefficient of variation in within-run precision was within 7% for APAL and 10 µM ADP-MA. Reference interval of APAL and 10 µM ADP-MA was 7.1 - 10.0 and 80.0 - 99.2%, respectively. APAL signifi-cantly decreased with the addition of 0.02 µM cangrelor, while 10 µM ADP-MA was barely affected. A significant correlation was observed between APAL and 10 µM ADP-MA (r = 0.94; p < 0.0001). CONCLUSIONS: The newly developed APAL system exhibited an acceptable performance. APAL score showed a good correlation with ADP-MA and was adequate to detect the weak effect of P2Y12 inhibitors. APAL is a new platelet aggregation scoring system with the potential to monitor the effects of P2Y12 inhibitor over a wide range.
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Adenosina Difosfato/farmacología , Pruebas de Coagulación Sanguínea/instrumentación , Agregación Plaquetaria/efectos de los fármacos , Pruebas de Función Plaquetaria/instrumentación , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/farmacología , Pruebas de Coagulación Sanguínea/métodos , Humanos , Inhibidores de Agregación Plaquetaria/farmacología , Pruebas de Función Plaquetaria/métodos , Plasma Rico en Plaquetas/efectos de los fármacos , Reproducibilidad de los ResultadosRESUMEN
BACKGROUND: Heparin-induced thrombocytopenia (HIT) is caused by platelet-activating antibodies that recognize platelet factor 4/heparin (PF4/hep)-complexes. The in vitro demonstration of PF4/hep antibodies using functional assays is essential for an optimal management of patients suspected to have HIT. However, conventional functional assays are technically challenging and limited to specialized laboratories. In contrast, flow cytometers are commonly used in routine laboratories. The aim of this study is to investigate the performance characteristics of a commercially available, flow cytometer based assay in the diagnosis of HIT. STUDY DESIGN: Sera of consecutive patients with suspected HIT were investigated using the Emo-test HIT Confirm® assay and compared to the standard method consisting of an IgG-specific enzyme immunoassay (EIA) for anti-PF4/hep antibodies and the heparin induced platelet aggregation (HIPA) test. RESULTS: 390 sera were included in the study, 164 sera tested IgG EIA-positive, of which 33 also tested HIPA-positive. No HIPA-positive samples were EIA-negative. In the Emo-test HIT Confirm® assay, 112 sera revealed positive results (%Heplaâ¯>â¯13); however, 51 (45.5%) were EIA-negative. Of the 33 HIPA-positive/EIA-positive HIT sera, 23 tested positive in the Emo-test HIT Confirm® assay, 2 gave ambiguous results, and 8 sera yielded false-negative results. Accordingly, the HIT Confirm® assay showed a sensitivity of 69.7% with a slightly better specificity of 75.4% compared to the EIA (sensitivity 100%, specificity 63.3%). An increase in diagnostic specificity for HIT to 85% was found when positive results were obtained in both the Emo-test HIT Confirm® assay and EIA. CONCLUSION: The Emo-Test HIT Confirm® assay may improve the specificity of laboratory investigations of HIT. However, the assay can only be recommended in combination with an immunoassay due to the high rate of false negativity. Our observation indicates a need to establish external quality assessment for functional assays to avoid such clinically relevant pitfalls.
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Anticoagulantes/efectos adversos , Heparina/efectos adversos , Trombocitopenia/inducido químicamente , Trombocitopenia/diagnóstico , Anciano , Reacciones Falso Negativas , Femenino , Citometría de Flujo/métodos , Humanos , Técnicas para Inmunoenzimas , Masculino , Persona de Mediana Edad , Agregación Plaquetaria/efectos de los fármacos , Plasma Rico en Plaquetas/efectos de los fármacosRESUMEN
La enfermedad de Crohn constituye una enfermedad inflamatoria crónica que puede cursar con fistulas complejas en hasta un 20% de los pacientes. A pesar de la intensificación del tratamiento, asociado o no a la cirugía, todavía es considerable el porcentaje de pacientes que no responden al tratamiento. En los últimos años se ha empezado a desarrollar nuevas terapias que permitan conseguir una mayor tasa de curación de estos pacientes, con las mínimas complicaciones posibles. Es cuando aparecen agentes que pretenden de forma directa el sellado o intervienen en la reducción local de la inflamación. Es objetivo de este artículo mostrar el papel de la Medicina Regenerativa en el tratamiento de estos pacientes.
Crohn's disease is a chronic inflammatory disease that can occur with complex fistulas in up to 20% of patients. Despite the intensification of treatment, associated with no surgery, the percentage of patients who do not respond to treatment is still considerable. In recent years, new therapies have been developed to achieve a higher cure rate for these patients, with the minimum possible complications. It is when agents appear to pretend as seal fistula tract as the local reduction of inflammation. The aim of this article is to show the role of Regenerative Medicine in the treatment of these patients.
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Humanos , Enfermedad de Crohn/terapia , Adhesivo de Tejido de Fibrina/uso terapéutico , Fístula Rectal/terapia , Medicina Regenerativa , Células Madre/efectos de los fármacos , Enfermedad de Crohn/complicaciones , Plasma Rico en Plaquetas/efectos de los fármacosRESUMEN
Lithium (Li) is used for the treatment and prophylaxis of mental disorders, associated with many serious hazards. Resveratrol (RSV) has various beneficial therapeutic effects. Platelet-rich plasma (PRP) is a new promising curative tool. This study aimed to assess the impacts of RSV versus PRP on lithium-induced thyroid follicular cell toxicity in adult male rats. Forty-nine adult male rats were divided into four groups: group I: control rats; group II: lithium-treated rats; group III: lithium- and resveratrol-treated rats; group IV: lithium and PRP-treated rats. Thyroid specimens were taken and processed for histological and immunohistochemical methods. Morphometrical studies and statistical analysis were done. Group II showed distorted thyroid follicles, significantly increased collagen fibers, and highly positive P53 immunostaining (P < 0.01). Ultrastructural examination showed dilated rough endoplasmic reticulum and damaged mitochondria. Groups III and IV exhibited significant amelioration of the histological and electron microscopic changes mentioned previously. PRP remedy was more effective than RSV for treatment of Li-induced thyroid follicular cell toxicity.
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Plasma Rico en Plaquetas/efectos de los fármacos , Resveratrol/farmacología , Células Epiteliales Tiroideas/metabolismo , Glándula Tiroides/ultraestructura , Animales , Inmunohistoquímica/métodos , Litio/farmacología , Masculino , Ratas , Células Epiteliales Tiroideas/patologíaRESUMEN
BACKGROUND: The benefits of platelet-rich plasma (PRP) are believed to be in part dependent on growth factor release after platelet activation. Platelet activation is complex and involves multiple mechanisms. One important mechanism is driven by cyclooxygenase 1 (COX-1)-mediated conversion of arachidonic acid (AA) to precursor prostaglandins that then mediate proinflammatory responses that trigger growth factor release. Acetylsalicylic acid (ASA; also known as aspirin) is known to irreversibly inhibit COX-1, thereby blocking AA-mediated signaling; however, it is unclear whether ASA use alters growth factor release from freshly isolated PRP. PURPOSE: To assess the effects of low-dose ASA use on activation of growth factor release from freshly isolated human PRP via AA and thrombin (TBN). STUDY DESIGN: Controlled laboratory study. METHODS: Twelve healthy men underwent blood collection and leukocyte-rich PRP (LR-PRP) preparation through a double-spin protocol to obtain baseline whole blood and PRP counts the same day. PRP was aliquoted into 3 groups: nonactivated, AA activated, and TBN activated. Immediately after activation, the concentrations of transforming growth factor ß1 (TGF-ß1), vascular endothelial growth factor (VEGF), and platelet-derived growth factor AB (PDGF-AB) were measured using enzyme-linked immunosorbent assays (ELISAs). The same 12 participants were then placed on an 81-mg daily dose of oral ASA for 14 days. Repeat characterization of whole blood and PRP analyses was done on day 14, followed by repeat ELISAs of growth factors under the same nonactivated and activated settings as previously stated. RESULTS: Fourteen days of daily ASA had no effect on the number of platelets and leukocytes measured in whole blood and LR-PRP. Compared with nonactivated LR-PRP, AA- and TBN-mediated activation led to significant release of VEGF and PDGF-AB. In contrast, release of TGF-ß1 from LR-PRP was observed only with activation by AA, not with TBN. Consistent with its inhibitory role in AA signaling, ASA significantly inhibited AA-mediated release of all 3 growth factors measured in this study. Although ASA had no effect on TBN-mediated release of VEGF and TGF-ß1 from LR-PRP, ASA did partially block TBN-mediated release of PDGF-AB, although the mechanism remains unclear. CONCLUSION: Daily use of low-dose ASA reduces VEGF, PDGF-AB, and TGF-ß1 expression in freshly isolated human LR-PRP when activated with AA. CLINICAL RELEVANCE: Reduction in growth factor release attributed to daily use of low-dose ASA or other COX inhibitors can be mitigated when PRP samples are activated with TBN. Clinical studies are needed to determine whether activation before PRP injection is needed in all applications where ASA is in use and to what extent ASA may inhibit growth factor release in vivo at the site of injury.