Characterization Of Pcl And Chitosan Nanoparticles As Carriers Of Enoxaparin And Its Antithrombotic Effect In Animal Models Of Venous Thrombosis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)This study was based on the preparation, characterization, and animal in vivo experiments performed to evaluate nanoparticles of poly(epsilon-caprolactone) (PCL) and chitosan as carriers of enoxaparin. Thenanoparticles were characterized and presented satisfactory results in terms of size, polydispersity, and encapsulation efficiency. Anticoagulant activity of the nanoparticles was maintained for 14 hours when the administration was subcutaneous; however no activity was observed after oral administration. There was a significant reduction in thrombus size, in vivo, for both free and encapsulated enoxaparin in comparison with the control group after subcutaneous administration. Oral administration results however were indifferent. In conclusion, the double emulsion method w/o/w was efficient for enoxaparin encapsulation, producing spherical nanoparticles with high encapsulation efficiency. For in vivo studies, the encapsulated enoxaparin showed a sustained anticoagulant activity for a higher period of time compared to free enoxaparin, with an antithrombotic effect when administered subcutaneously.FAPESPCNPqBrazilian Network on Nanocosmetics (MCT/CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Angiopoietin2 is associated with coagulation activation and tissue factor expression in extracellular vesicles in COVID-19

Coagulation activation in immunothrombosis involves various pathways distinct from classical hemostasis, offering potential therapeutic targets to control inflammation-induced hypercoagulability while potentially sparing hemostasis. The Angiopoietin/Tie2 pathway, previously linked to embryonic angiogenesis and sepsis-related endothelial barrier regulation, was recently associated with coagulation activation in sepsis and COVID-19. This study explores the connection between key mediators of the Angiopoietin/Tie2 pathway and coagulation activation. The study included COVID-19 patients with hypoxia and healthy controls. Blood samples were processed to obtain platelet-free plasma, and frozen until analysis. Extracellular vesicles (EVs) in plasma were characterized and quantified using flow cytometry, and their tissue factor (TF) procoagulant activity was measured using a kinetic chromogenic method. Several markers of hemostasis were assessed. Levels of ANGPT1, ANGPT2, and soluble Tie2 correlated with markers of coagulation and platelet activation. EVs from platelets and endothelial cells were increased in COVID-19 patients, and a significant increase in TF+ EVs derived from endothelial cells was observed. In addition, ANGPT2 levels were associated with TF expression and activity in EVs. In conclusion, we provide further evidence for the involvement of the Angiopoietin/Tie2 pathway in the coagulopathy of COVID-19 mediated in part by release of EVs as a potential source of TF activity

Characterization of autologous platelet rich plasma and its biological effects in patients with Behçet Disease

Orientador: Joyce Maria Annichino-BizzacchiTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: A doença de Behçet (DB) é uma vasculite crônica sistêmica imunomediada, caracterizada por manifestações clínicas que incluem: úlceras muco-cutâneas, comprometimento ocular, alterações imunológicas, envolvimento vascular e neurológico. Sua etiologia ainda é desconhecida, mas acredita-se que está relacionada com a interação de fatores genéticos, ambientais e imunológicos que caracterizam o curso da doença. Os tratamentos disponíveis apresentam limitações como alto custo e efeitos colaterais, sendo que a busca por um tratamento biológico de baixo custo com potencial imunomodulador, torna-se de grande valia. O plasma rico em plaquetas (PRP) apresenta algumas características que poderiam indicar o seu uso como imunomodulador devido a vasta gama de citocinas secretadas, especialmente pela participação do TGF-? na diferenciação das células Treg. Diante do exposto, o objetivo deste estudo foi: caracterizar o PRP de pacientes com DB e úlceras em atividade, e avaliar seus efeitos como imunomodulador através de aplicação subcutânea. Para isto, foram selecionados pacientes com diagnóstico de DB segundo os critérios do ISGBD, acompanhados no Ambulatório de Vasculites do Hospital das Clínicas da Unicamp, no período de fevereiro de 2014 a agosto de 2015. Os critérios de inclusão foram o uso de dose máxima de 10 mg de prednisona ao dia, e presença de úlceras orais. Os critérios de exclusão foram atividade neurológica ou ocular, uso de anticoagulante ou antiagregante plaquetário, e neoplasia nos últimos 5 anos. O P-PRP foi caracterizado através da contagem de plaquetas e leucócitos, e pela quantificação de citocinas. Os efeitos do uso do P-PRP foram avaliados pela frequência de células TCD4+, TCD8+, Treg, NK e NK ativada, além do perfil de citocinas no plasma dos pacientes, e também pelas manifestações clínicas através do escore BR-BDCAF, do questionário SF-36, e da avaliação do número e do tempo de fechamento das úlceras orais. Os resultados mostraram uma mediana do número de plaquetas de 1.130.000 células/µL e leucócitos de 1500/µl, apresentando valores adequados e sem variações siginificativas inter e intra-individual. Os níveis de citocinas presentes no P-PRP foram relativamente homogêneos, com exceção do TGF-?1, o qual apresentou um aumento siginificativo (p<0,0001) nos 6 meses (última aplicação). As avaliações sistêmicas durante o uso do P-PRP mostraram alterações significativas, com resultados relevantes para o aumento da frequência de células Treg (p=0,0416) e diminuição de células NK ativadas (p=0,0010), durante o período de seguimento dos pacientes. Contudo, não se observou uma correlação clínica com as dosagens sistêmicas. O dado clínico mais relevante foi a diminuição no tempo de fechamento das úlceras durante todo o período de aplicação. Apesar de resultados preliminares e ao pequeno número de pacientes, nossos resultados indicam que a avaliação da função celular de Treg seria interessante para implementar o conhecimento sobre o efeito do P-PRP. Além disso, critérios clínicos mais objetivos para análise dos resultados seriam importantes para definição dos efeitosAbstract: Behçet's disease (DB) is an immune-mediated chronic systemic vasculitis, characterized by clinical manifestations that include: mucocutaneous ulcers, ocular involvement, immunological alterations, vascular and neurological involvement. Its etiology is still unknown, but it is believed to be related to the interaction of genetic, environmental and immunological factors that characterize the course of the disease. The available treatments present limitations such as high cost and side effects, and the search for a low cost biological treatment with immunomodulatory potential, becomes of great value. Platelet rich plasma (PRP) has some characteristics that could indicate its use as an immunomodulator due to the wide range of secreted cytokines, especially by the participation of TGF-? in the differentiation of Treg cells. In view of the above, the objective of this study was: to characterize the PRP of patients with DB and active ulcers, and to evaluate its effects as an immunomodulator through subcutaneous application. For this, we selected patients with a diagnosis of DB according to the criteria of the ISGBD, followed in the Vasculitis Outpatient Clinic of the Hospital das Clínicas of Unicamp, from February 2014 to August 2015. The inclusion criteria were the use of a maximum dose of 10 mg prednisone per day, and presence of oral ulcers. Exclusion criteria were neurological or ocular activity, use of anticoagulant or antiplatelet agent, and neoplasia in the last 5 years. P-PRP was characterized by platelet and leukocyte counts, and quantification of cytokines. The effects of the use of P-PRP were evaluated by the cell frequency of TCD4 +, TCD8 +, Treg, NK and NK activated, as well as the cytokine profile in the patients' plasma, and the clinical manifestations through the BR-BDCAF score, SF-36 questionnaire, and the assessment of the number and timing of oral ulcer closure. The results showed a median platelet count of 1130,000 cells/?L and leukocytes of 1500/?L, presenting adequate values and without significant inter- and intra-individual variations. The levels of cytokines present in P-PRP were relatively homogeneous, with the exception of TGF-?1, which presented a significant increase (p <0.0001) at 6 months (last application). The systemic evaluations during the use of P-PRP showed significant alterations, with relevant results for the increase in Treg cell frequency (p = 0.0416) and decrease in activated NK cells (p = 0.0010) during the follow-up. However, no clinical correlation was observed with systemic dosages. The most relevant clinical data was the decrease in the closing time of ulcers throughout the application period. Despite preliminary results and small numbers of patients, our results indicate that the evaluation of Treg cell function would be interesting to implement knowledge about the effect of P-PRP. In addition, more objective clinical criteria for outcome analysis would be important in defining effectsDoutoradoFisiopatologia MédicaDoutora em CiênciasCAPE

Evaluation of the action of topical use of encapsulated heparin in polymeric nanoparticles recovered by chitosan in rat skin ulcer model

Orientador: Joyce Maria Anicchino-BizzacchiDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo poderá ser visualizado no texto completo da tese digitalAbstract is available with the full electronic documentMestradoClinica MedicaMestra em Clínica Médic

Leukocyte-rich PRP for knee osteoarthritis: current concepts

Knee osteoarthritis is a major painful and debilitating orthopaedic disease affecting a large number of adult individuals on a global scale. Over the years, this severe condition has been widely studied and while many alternatives have been utilized, platelet-rich plasma (PRP) remains one of the most popular solutions among researchers and clinicians alike. While there are different formulations and techniques involved in the preparation of PRP, produced either manually or via the use of commercial kits, the presence of leukocytes in a PRP mixture is a factor that raises concern due to their well-known pro-inflammatory activity. Although it is reasonable to worry about this, it should be taken into consideration that in order for the healing process to occur, the inflammatory phase is necessary. Leukocytes present in the inflammatory phase release both pro and anti-inflammatory molecules and, when combined with activated platelets, their potential increases. Additionally, due to the macrophage's plasticity to switch from the subtype 1 to subtype 2, it is suggested that the inclusion of the components from the buffy coat layer in a PRP mixture, classifying it as leukocyte-rich platelet-rich plasma or L-PRP, may provide benefits instead of detriments, from a standpoint of the regenerative potential of PRP101S179S182sem informaçãosem informaçã

Characterization of PCL and Chitosan Nanoparticles as Carriers of Enoxaparin and Its Antithrombotic Effect in Animal Models of Venous Thrombosis

This study was based on the preparation, characterization, and animal in vivo experiments performed to evaluate nanoparticles of poly(ɛ-caprolactone) (PCL) and chitosan as carriers of enoxaparin. The nanoparticles were characterized and presented satisfactory results in terms of size, polydispersity, and encapsulation efficiency. Anticoagulant activity of the nanoparticles was maintained for 14 hours when the administration was subcutaneous; however no activity was observed after oral administration. There was a significant reduction in thrombus size, in vivo, for both free and encapsulated enoxaparin in comparison with the control group after subcutaneous administration. Oral administration results however were indifferent. In conclusion, the double emulsion method w/o/w was efficient for enoxaparin encapsulation, producing spherical nanoparticles with high encapsulation efficiency. For in vivo studies, the encapsulated enoxaparin showed a sustained anticoagulant activity for a higher period of time compared to free enoxaparin, with an antithrombotic effect when administered subcutaneously

Characterization of PCL and chitosan nanoparticles as carriers of enoxaparin and its antithrombotic effect in animal models of venous thrombosis

This study was based on the preparation, characterization, and animal in vivo experiments performed to evaluate nanoparticles of poly(epsilon-caprolactone) (PCL) and chitosan as carriers of enoxaparin. Thenanoparticles were characterized and presented satisfactory results in terms of size, polydispersity, and encapsulation efficiency. Anticoagulant activity of the nanoparticles was maintained for 14 hours when the administration was subcutaneous; however no activity was observed after oral administration. There was a significant reduction in thrombus size, in vivo, for both free and encapsulated enoxaparin in comparison with the control group after subcutaneous administration. Oral administration results however were indifferent. In conclusion, the double emulsion method w/o/w was efficient for enoxaparin encapsulation, producing spherical nanoparticles with high encapsulation efficiency. For in vivo studies, the encapsulated enoxaparin showed a sustained anticoagulant activity for a higher period of time compared to free enoxaparin, with an antithrombotic effect when administered subcutaneously2017CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPsem informaçã

The Mechanism of Action between Pulsed Radiofrequency and Orthobiologics: Is There a Synergistic Effect?

Radiofrequency energy is a common treatment modality for chronic pain. While there are different forms of radiofrequency-based therapeutics, the common concept is the generation of an electromagnetic field in the applied area, that can result in neuromodulation (pulsed radiofrequency—PRF) or ablation. Our specific focus relates to PRF due to the possibility of modulation that is in accordance with the mechanisms of action of orthobiologics. The proposed mechanism of action of PRF pertaining to pain relief relies on a decrease in pro-inflammatory cytokines, an increase in cytosolic calcium concentration, a general effect on the immune system, and a reduction in the formation of free radical molecules. The primary known properties of orthobiologics constitute the release of growth factors, a stimulus for endogenous repair, analgesia, and improvement of the function of the injured area. In this review, we described the mechanism of action of both treatments and pertinent scientific references to the use of the combination of PRF and orthobiologics. Our hypothesis is a synergic effect with the combination of both techniques which could benefit patients and improve the life quality

Contributions for classification of platelet rich plasma – proposal of a new classification: MARSPILL

Platelet-rich plasma (PRP) has emerged as a significant therapy used in medical conditions with heterogeneous results. There are some important classifications to try to standardize the PRP procedure. The aim of this report is to describe PRP contents studying celular and molecular components, and also propose a new classification for PRP. The main focus is on mononuclear cells, which comprise progenitor cells and monocytes. In addition, there are important variables related to PRP application incorporated in this study, which are the harvest method, activation, red blood cells, number of spins, image guidance, leukocytes number and light activation. The other focus is the discussion about progenitor cells presence on peripherial blood which are interesting due to neovasculogenesis and proliferation. The function of monocytes (in tissue-macrophages) are discussed here and also its plasticity, a potential property for regenerative medicine treatments125565574sem informaçãosem informaçã

Sacral Bioneuromodulation: The Role of Bone Marrow Aspirate in Spinal Cord Injuries

Spinal cord injury (SCI) represents a severe trauma to the nervous system, leading to significant neurological damage, chronic inflammation, and persistent neuropathic pain. Current treatments, including pharmacotherapy, immobilization, physical therapy, and surgical interventions, often fall short in fully addressing the underlying pathophysiology and resultant disabilities. Emerging research in the field of regenerative medicine has introduced innovative approaches such as autologous orthobiologic therapies, with bone marrow aspirate (BMA) being particularly notable for its regenerative and anti-inflammatory properties. This review focuses on the potential of BMA to modulate inflammatory pathways, enhance tissue regeneration, and restore neurological function disrupted by SCI. We hypothesize that BMA’s bioactive components may stimulate reparative processes at the cellular level, particularly when applied at strategic sites like the sacral hiatus to influence lumbar centers and higher neurological structures. By exploring the mechanisms through which BMA influences spinal repair, this review aims to establish a foundation for its application in clinical settings, potentially offering a transformative approach to SCI management that extends beyond symptomatic relief to promoting functional recovery