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INTRODUCTION: Haemophilia B (HB) is an X-linked hereditary bleeding disorder characterized by coagulation factor IX (FIX) deficiency. To improve the quality of life of patients and adherence to treatment, recombinant factor concentrates modified to extend their half-life have been developed, called extended half-life factors (EHL: extended half-life). Nonacog beta pegol (N9-GP) is a glycopegylated recombinant human FIX molecule that has a half-life of 93 h with a single dose and has shown a higher recovery percentage than other molecules. To diagnose and monitor the treatment of haemophiliac patients, FIX activity is determined with the one-stage clotting assay (OSA) and/or the chromogenic assay. The objective of this work, carried out in three centres, was to measure the recovery of N9-PG with 10 different activated partial thromboplastin time (APTT) reagents on three platforms, in samples spiked in vitro with N9-GP, at four different concentration levels. METHODS: It was measured the recovery of N9-GP with 10 different APTT reagents (polyphenol, ellagic acid, silice dioxide, colloidal silica as APTT activator on three platforms, in sample spiked in vitro with N9-GP. RESULTS: The results show heterogeneity in the activity of N9-GP measured by OSA with the different APTT reagents when the calibrations were performed with the specific calibrator of each coagulometer. A recovery percentage between 87% and 108% was obtained only with polyphenol and ellagic acid as activator in the three platforms evaluated. The other reagents studied overestimate or underestimate, with no clear profile. When a calibration curve was performed with a calibrator prepared from the N9-GP vial, all APTT reagents met the established recovery requirement. CONCLUSION: APTT reagents with polyphenol or ellagic acid as activator would be the only ones appropriate when using the commercially available OSA with specific calibrator to monitor patients treated with N9-GP.

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Attenuated Total Reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy is an emerging technology in the medical field. Blood D-dimer was initially studied as a marker of the activation of coagulation and fibrinolysis. It is mainly used as a potential diagnosis screening test for pulmonary embolism or deep vein thrombosis but was recently associated with COVID-19 severity. This study aimed to evaluate the use of ATR-FTIR spectroscopy with machine learning to classify plasma D-dimer concentrations. The plasma ATR-FTIR spectra from 100 patients were studied through principal component analysis (PCA) and two supervised approaches: genetic algorithm with linear discriminant analysis (GA-LDA) and partial least squares with linear discriminant (PLS-DA). The spectra were truncated to the fingerprint region (1800-1000 cm-1). The GA-LDA method effectively classified patients according to D-dimer cutoff (≤0.5 µg/mL and >0.5 µg/mL) with 87.5 % specificity and 100 % sensitivity on the training set, and 85.7 % specificity, and 95.6 % sensitivity on the test set. Thus, we demonstrate that ATR-FTIR spectroscopy might be an important additional tool for classifying patients according to D-dimer values. ATR-FTIR spectral analyses associated with clinical evidence can contribute to a faster and more accurate medical diagnosis, reduce patient morbidity, and save resources and demand for professionals.

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BACKGROUND: Scaffold (SCA) functionalization with aptamers (APT) provides adsorption of specific bioactive molecules on biomaterial surfaces. The aim of this study was to observe if SCA enriched with anti-fibronectin APT can favor coagulum (PhC) and osteoblasts (OSB) differentiation. METHODS: 20 µg of APT was functionalized on SCA by simple adsorption. For PhC formation, SCAs were inserted into rat calvaria defects for 17 h. Following proper transportation (buffer solution PB), OSBs (UMR-106 lineage) were seeded over PhC + SCAs with and without APT. Cells and PhC morphology, PhC cell population, protein labeling and gene expression were observed in different time points. RESULTS: The APT induced higher alkaline phosphatase and bone sialoprotein immunolabeling in OSB. Mesenchymal stem cells, leukocytes and lymphocytes cells were detected more in the APT group than when scaffolds were not functionalized. Additionally, an enriched and dense fibrin network and different cell types were observed, with more OSB and white blood cells in PhC formed on SCA with APT. The gene expression showed higher transforming growth factor beta 1 (TGF-b1) detection in SCA with APT. CONCLUSIONS: The SCA functionalization with fibronectin aptamers may alter key morphological and functional features of blood clot formation, and provides a selective expression of proteins related to osteo differentiation. Additionally, aptamers increase TGF-b1 gene expression, which is highly associated with improvements in regenerative therapies.

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Abstract Background The efficacy of intravenous thrombolysis (IVT) is time-dependent. Objective To compare the door-to-needle (DTN) time of stroke neurologists (SNs) versus non-stroke neurologists (NSNs) and emergency room physicians (EPs). Additionally, we aimed to determine elements associated with DTN ≤ 20 minutes. Methods Prospective study of patients with IVT treated at Clínica Alemana between June 2016 and September 2021. Results A total of 301 patients underwent treatment for IVT. The mean DTN time was 43.3 ± 23.6 minutes. One hundred seventy-three (57.4%) patients were evaluated by SNs, 122 (40.5%) by NSNs, and 6 (2.1%) by EPs. The mean DTN times were 40.8 ± 23, 46 ± 24.7, and 58 ± 22.5 minutes, respectively. Door-to-needle time ≤ 20 minutes occurred more frequently when patients were treated by SNs compared to NSNs and EPs: 15%, 4%, and 0%, respectively (odds ratio [OR]: 4.3, 95% confidence interval [95%CI]: 1.66-11.5, p = 0.004). In univariate analysis DTN time ≤ 20 minutes was associated with treatment by a SN (p = 0.002), coronavirus disease 2019 pandemic period (p = 0.21), time to emergency room (ER) (p = 0.21), presence of diabetes (p = 0.142), hypercholesterolemia (p = 0.007), atrial fibrillation (p < 0.09), score on the National Institutes of Health Stroke Scale (NIHSS) (p = 0.001), lower systolic (p = 0.143) and diastolic (p = 0.21) blood pressures, the Alberta Stroke Program Early CT Score (ASPECTS; p = 0.09), vessel occlusion (p = 0.05), use of tenecteplase (p = 0.18), thrombectomy (p = 0.13), and years of experience of the physician (p < 0.001). After multivariate analysis, being treated by a SN (OR: 3.95; 95%CI: 1.44-10.8; p = 0.007), NIHSS (OR: 1.07; 95%CI: 1.02-1.12; p < 0.002) and lower systolic blood pressure (OR: 0.98; 95%CI: 0.96-0.99; p < 0.003) remained significant. Conclusions Treatment by a SN resulted in a higher probability of treating the patient in a DTN time within 20 minutes.

Resumen Antecedentes La respuesta a la trombólisis intravenosa (TIV) es dependiente del tiempo. Objetivo Comparar los tiempo puerta-aguja (TPAs) de neurólogos vasculares (NVs) contra los de neurólogos no vasculares (NNVs) y médicos emergencistas (MEs), y determinar los elementos asociados a un PTA ≤ 20 minutos. Métodos Análisis observacional prospectivo de pacientes con TIV tratados en Clínica Alemana entre junio de 2016 y septiembre de 2021. Resultados En total, 301 pacientes con TIV fueron tratados. El TPA promedio fue de 43,3 ± 23,6 minutos. Un total de 173 (57,4%) pacientes fueron evaluados por NVs, 122 (40,5%), por NNVs, y 6 (2,1%), por MEs; los TPAs promedios fueron de 40,8 ± 23; 46 ± 24,7 y 58 ± 22,5 minutos, respectivamente. Los TPAs ≤ 20 minutos fueron más frecuentes en pacientes tratados por NVs versus NNVs y MEs: 15%, 4% y 0%, respectivamente (odds ratio [OR]: 4,3; intervalo de confianza del 95% [IC95%]: 1,66-11,5; p = 0,004). El análisis univariado demostró que TPA ≤ 20 minutos se asoció con: tratamiento por NVs (p = 0,002), periodo de la pandemia de enfermedad por coronavirus 2019 (COVID-19; p = 0,21), tiempo a urgencia (p = 0,21), diabetes (p = 0,142), hipercolesterolemia (p = 0,007), fibrilación auricular (p < 0,09), puntaje en la National Institutes of Health Stroke Scale [NIHSS] (p = 0,001), presión arterial sistólica (p = 0,143) y diastólica menores (p = 0,21), Alberta Stroke Program Early CT Score (ASPECTS ; p = 0,09), oclusión de vasos cerebrales (p =0,05), uso de tecneteplase (p = 0,18), trombectomía (p = 0,13) y años de experiencia del médico (p < 0,001). El análisis multivariado demostró que ser tratado por NVs (OR: 3,95; IC95%: 1,44-10,8; p = 0,007), el puntaje en la NIHSS (OR: 1,07; IC95%: 1,02-1,12; p < 0,002) y la presión arterial sistólica (OR: 0,98; IC95%: 0,96-0,99; p < 0,003) se asociaron a TPA ≤ 20 minutos. Conclusões El tratamiento por NVs resultó en un TPA menor y en una mayor probabilidad de tratamiento ≤ 20 minutos.

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Titanium (Ti) nanotopography modulates the osteogenic response to exogenous bone morphogenetic protein 7 (BMP-7) in vitro, supporting enhanced alkaline phosphatase mRNA expression and activity, as well as higher osteopontin (OPN) mRNA and protein levels. As the biological effects of OPN protein are modulated by its proteolytic cleavage by serum proteases, this in vitro study evaluated the effects on osteogenic cells in the presence of a physiological blood clot previously formed on a BMP-7-coated nanostructured Ti surface obtained by chemical etching (Nano-Ti). Pre-osteoblastic MC3T3-E1 cells were cultured during 5 days on recombinant mouse (rm) BMP-7-coated Nano-Ti after it was implanted in adult female C57BI/6 mouse dorsal dermal tissue for 18 h. Nano-Ti without blood clot or with blood clot at time 0 were used as the controls. The presence of blood clots tended to inhibit the expression of key osteoblast markers, except for Opn, and rmBMP-7 functionalization resulted in a tendency towards relatively greater osteoblastic differentiation, which was corroborated by runt-related transcription factor 2 (RUNX2) amounts. Undetectable levels of OPN and phosphorylated suppressor of mothers against decapentaplegic (SMAD) 1/5/9 were noted in these groups, and the cleaved form of OPN was only detected in the blood clot immediately prior to cell plating. In conclusion, the strategy to mimic in vitro the initial interfacial in vivo events by forming a blood clot on a Ti nanoporous surface resulted in the inhibition of pre-osteoblastic differentiation, which was minimally reverted with an rmBMP-7 coating.

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Mixed infections by different Trypanosoma species or genotypes are a common and puzzling phenomenon. Therefore, it is critical to refine the diagnostic techniques and to understand to what extent these methods detect trypanosomes. We aimed to develop an accessible strategy to enhance the sensitivity of the hemoculture, as well as to understand the limitations of the hemoculture and the blood clot as a source of parasitic DNA. We investigated trypanosomatid infections in 472 bats by molecular characterization (18S rDNA gene) of the DNA obtained from the blood clot and, innovatively, from three hemoculture sample types: the amplified flagellates ("isolate"), the pellet of the culture harvested in its very initial growth stage ("first aliquot"), and the pellet of non-grown cultures with failure of amplification ("sediment"). We compared (a) the characterization of the flagellates obtained by first aliquots and isolates; and (b) the performance of the hemoculture and blood clot for trypanosomatid detection. We observed: (i) a putative new species of Bodo in Artibeus lituratus; (ii) the potential of Trypanosoma cruzi selection in the hemoculture; (iii) that the first aliquots and sediments overcome the selective pressure of the hemoculture; and (iv) that the blood clot technique performs better than the hemoculture. However, combining these methods enhances the detection of single and mixed infections.

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BACKGROUND: Under certain clinical and experimental conditions hematopoiesis occurs in other site than bone marrow (BM), such as the liver. Here, we develop a 3D organoid that mimics several components of the hematopoietic niche present during liver extramedullary hematopoiesis. AIM: To evaluate the capacity of a 3D hematopoietic organoid (3D-HO) to function as a hematopoietic like-niche allowing for blood cell production outside of the BM. METHODS: The 3D-HO is constituted by liver sinusoidal endothelial cells (LSEC) as the stromal component, BM isolated from 5-FU treated mice (FU-BMCs), collagen microspheres and plasma clot as scaffolds. The ability of the 3D-HO to support the survival and functionality of FU-BMCs was investigated by using confocal microscopy, histology analysis, flow cytometry, and clonogenic assays. RESULTS: After 15 and 30 days, post-ectopic implantation, histological studies of the 3D-HO showed the presence of cells with myeloid and lymphoid lineage morphology. Flow cytometry analysis of these cells showed the presence of cells expressing hematopoietic stem progenitor cells (HSPC) (Sca-1+/c-Kit+), myeloid (Gr-1+) and lymphoid (B220+ and CD19+) markers. Clonogenic assays showed that cells from the 3D-HO formed hematopoietic colonies. Expression of the Sry gene by cells from the 3D-HO, implanted for 30 days in female mice, indicated that male donor cells persist in this model of extramedullary hematopoiesis. CONCLUSIONS: The 3D-HO constitutes an extramedullary hematopoietic-like niche which supports the survival and functionality of FU-BMCs. It may constitute an efficient model for investigating, in vitro and in vivo, those factors that control hematopoiesis outside BM.

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Introducción: La elevación de seno maxilar con implante dental simultáneo, sin el uso de substitutos óseos, ha retomado la teoría del potencial de neoformación ósea, que el coágulo de sangre alojado bajo la membrana de Schneider por sí solo presenta. Excluyendo así la necesidad de materiales exógenos. Objetivo: Exponer la efectividad de la elevación de seno maxilar con implante dental simultáneo, sin el uso de injerto, mediante la evaluación de la cantidad de ganancia ósea vertical. Presentación de casos: Caso 1: Paciente de 62 años, género femenino, que fue sometida a una elevación de seno maxilar sin injerto e implante dental simultáneo, con técnica de ventana lateral debido a altura ósea reducida de 5,24 mm. Luego del período de seguimiento tomográfico de 12 meses, después de la carga protésica, se logró una altura ósea vertical de 10,2 mm, lo que dio como resultado una ganancia ósea vertical de 4,96 mm. Caso 2: Paciente de 48 años, género femenino, que fue sometida a una elevación de seno maxilar sin injerto e implante dental simultáneo, con técnica transalveolar, mediante el uso de piezoeléctrico debido a una altura ósea reducida de 8,33 mm. Luego del período de seguimiento radiográfico de 4 meses, antes de la carga protésica, se logró una altura ósea vertical de 11,55 mm, lo que dio como resultado una ganancia ósea vertical de 3,19 mm. Conclusiones: Con base en estos 2 informes de casos, la elevación de seno maxilar e implante dental simultáneo sin injerto se asocian con la reducción de la morbilidad quirúrgica, menor probabilidad de procesos infecciosos y menor costo de la cirugía. Por lo tanto, puede considerarse una alternativa quirúrgica para la colocación de implantes en el maxilar posterior superior debido a deficiencias óseas verticales, independientemente de la técnica (lateral o transalveolar(AU)

Introduction: Maxillary sinus lift with simultaneous dental implantation without using bone substitutes, reapproaches the theory about the bone neoformation potential of the blood clot housed under the Schneider membrane, all by itself, thus excluding the need to use exogenous materials. Objective: Discuss the effectiveness of graftless maxillary sinus lift with simultaneous dental implant placement through an evaluation of the amount of vertical bone gain. Case presentation: Case 1: Female 62-year-old patient undergoing graftless maxillary sinus lift with simultaneous dental implantation by lateral window technique due to a reduced bone height of 5.24 mm. After a 12-month tomographic follow-up period subsequent to prosthetic loading, a 10.2 mm vertical bone height was achieved, resulting in 4.96 mm vertical bone gain. Case 2: Female 48-year-old patient undergoing graftless maxillary sinus lift with simultaneous dental implantation by transalveolar technique using a piezoelectric generator due to a reduced bone height of 8.33 mm. After a 4-month radiographic follow-up period, before prosthetic loading, an 11.55 mm vertical bone height was achieved, resulting in 3.19 mm vertical bone gain. Conclusions: According to these two case reports, graftless maxillary sinus lifting with simultaneous dental implantation is associated to reduced surgical morbidity, a lesser probability of infectious processes and lower surgical costs. It may therefore be considered to be a surgical alternative for implant placement in the posterior maxilla due to vertical bone deficiencies, regardless of the technique used (lateral or transalveolar(AU)

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Body bones play diverse pivotal roles, including the protection of vital organs. For instance, the integrative functions of the brain controlling diverse peripheral actions can be affected by a traumatic injury on the calvaria and the reparative process of a large defect is a challenge in the integrative physiology. Therefore, the development of biomaterials and approaches to improve such defects still requires substantial advances. In this regard, the most attractive approaches have been covering the cavity with inorganic bovine bone (IBB) and, more recently, also using low-level laser therapy (LT), but this issue has opened many questions. Here, it was determined the number of LT sessions required to speed up and to intensify the recovery process of two 5-mm-diameter defects promoted in the calvaria of each subgroup of six adult Wistar rats. The quantitative data showed that 30 days post-surgery, the recovery process by using blood clot-filling was not significantly influenced by the number of LT sessions. However, in the IBB-filled defects, the number of LT sessions markedly contributed to the improvement of the reparative process. Compared to the Control group (non-irradiated), the percentage of mineralization (formation of new bone into the cavities) gradually increased 25, 49, and 52% with, respectively, 4, 7, and 11 sessions of LT. In summary, combining the use of IBB with seven sessions of LT seems to be an optimal approach to greatly improve the recovery of calvarial defects. This translational research opens new avenues targeting better conditions of life for those suffering from large bone traumas and in the present field could contribute to preserve the integrative functions of the brain.

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Recent advances in ultrasound technology have made modern handheld ultrasound devices and are contributing to make bedside ultrasound evaluation a practice available to all physicians. A 46-year-old woman with history of systemic erythematosus presented to our hospital with 14 days of COVID-19. The patient suddenly presented greater respiratory distress, tachycardia, hypotension, and increased supplemental oxygen requirements; so she required mechanical ventilation. Point-of-care ultrasound assessment with handheld ultrasound device was observed on the apical view an apical thrombus in the right ventricle, McConnell's sign. The patient underwent systemic thrombolysis with alteplase showing improvement in mechanical ventilation parameters and is currently continuing treatment for COVID-19 in the intensive care unit of our hospital. Emerging technologies such as handheld ultrasound devices can provide high-quality care to the patients. Routine screening of patients with COVID-19 using handheld ultrasound is feasible, may be able to define prognosis and treatment of cardiovascular complications.

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Bats are infected with several trypanosomatid species; however, assessing the diversity of this interaction remains challenging since there are species apparently unable to grow in conventional culture media. Accordingly, the ecology and biology of the Molecular Operational Taxonomic Units (MOTUs) Trypanosoma spp. Neobats are unknown. Therefore, we performed the molecular characterization targeting the 18S small subunit rDNA from the blood clot of 280 bats of three Brazilian regions (Paraíba, Rio de Janeiro and Acre states), bypassing the selective pressure of hemoculture. From 68 (24%) positive blood clot samples, we obtained 49 satisfactory sequences. Of these successfully sequenced results, T. spp. Neobats (1, 3 and 4) represented 67%, with the most abundant T. sp. Neobat 4 (53%). Our results show: (1) high abundance and wide geographic range of T. sp. Neobat 4, restricted to Carollia bats; (2) high infection rate of T. sp. Neobat 4 in Carollia perspicillata populations (mean 26%); (3) infection with the monoxenous Crithidia mellificae; and (4) a new MOTU (T. sp. Neobat 5) in Artibeus cinereus, positioning in the Trypanosoma wauwau clade. These data corroborate the importance of bats as hosts of many Trypanosoma species and C. mellificae. They also show that the diversity of the T. wauwau clade is underestimated and warn about the high magnitude of trypanosomes we overpass with the hemoculture. Our findings combined with previous data show that T. spp. Neobats include host-specific and host-generalist species, probably playing different ecological roles: T. sp. Neobat 1 shows broad host range; T. spp. Neobat 3 and 4 are restricted to Artibeus and Carollia, respectively. Finally, T. Neobat 4 seems to be a well-succeeded parasite, especially within C. perspicillata metapopulations across a wide geographical distribution. This work is a step forward to understand the biology and life history of T. spp. Neobats.

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Leptospirosis is a neglected zoonosis, caused by pathogenic spirochetes bacteria of the genus Leptospira. The molecular mechanisms of leptospirosis infection are complex, and it is becoming clear that leptospires express several functionally redundant proteins to invade, disseminate, and escape the host's immune response. Here, we describe a novel leptospiral protein encoded by the gene LIC13086 as an outer membrane protein. The recombinant protein LIC13086 can interact with the extracellular matrix component laminin and bind plasminogen, thus possibly participating during the adhesion process and dissemination. Also, by interacting with fibrinogen and plasma fibronectin, the protein LIC13086 probably has an inhibitory effect in the fibrin clot formation during the infection process. The newly characterized protein can also bind molecules of the complement system and the regulator C4BP and, thus, might have a role in the evasion mechanism of Leptospira. Taken together, our results suggest that the protein LIC13086 may have a multifunctional role in leptospiral pathogenesis, participating in host invasion, dissemination, and immune evasion processes.

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The rise in musculoskeletal disorders has prompted medical experts to devise novel effective alternatives to treat complicated orthopedic conditions. The ever-expanding field of regenerative medicine has allowed researchers to appreciate the therapeutic value of bone marrow-derived biological products, such as the bone marrow aspirate (BMA) clot, a potent orthobiologic which has often been dismissed and regarded as a technical complication. Numerous in vitro and in vivo studies have contributed to the expansion of medical knowledge, revealing optimistic results concerning the application of autologous bone marrow towards various impactful disorders. The bone marrow accommodates a diverse family of cell populations and a rich secretome; therefore, autologous BMA-derived products such as the "BMA Matrix", may represent a safe and viable approach, able to reduce the costs and some drawbacks linked to the expansion of bone marrow. BMA provides -it eliminates many hurdles associated with its preparation, especially in regards to regulatory compliance. The BMA Matrix represents a suitable alternative, indicated for the enhancement of tissue repair mechanisms by modulating inflammation and acting as a natural biological scaffold as well as a reservoir of cytokines and growth factors that support cell activity. Although promising, more clinical studies are warranted in order to further clarify the efficacy of this strategy.

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In the clinical laboratory, knowledge of and the correct use of clot activators and anticoagulant additives are critical to preserve and maintain samples in optimal conditions prior to analysis. In 2017, the Latin America Confederation of Clinical Biochemistry (COLABIOCLI) commissioned the Latin American Working Group for Preanalytical Phase (WG-PRE-LATAM) to study preanalytical variability and establish guidelines for preanalytical procedures to be applied by clinical laboratories and health care professionals. The aim of this critical review, on behalf of COLABIOCLI WG-PRE-LATAM, is to provide information to understand the mechanisms of the interactions and reactions that occur between blood and clot activators and anticoagulant additives inside evacuated tubes used for laboratory testing. Clot activators - glass, silica, kaolin, bentonite, and diatomaceous earth - work by surface dependent mechanism whereas extrinsic biomolecules - thrombin, snake venoms, ellagic acid, and thromboplastin - start in vitro coagulation when added to blood. Few manufacturers of evacuated tubes state the type and concentration of clot activators used in their products. With respect to anticoagulant additives, sodium citrate and oxalate complex free calcium and ethylenediaminetetraacetic acid chelates calcium. Heparin potentiates antithrombin and hirudin binds to active thrombin, inactivating the thrombin irreversibly. Blood collection tubes have improved continually over the years, from the glass tubes containing clot activators or anticoagulant additives that were prepared by laboratory personnel to the current standardized evacuated systems that permit more precise blood/additive ratios. Each clot activator and anticoagulant additive demonstrates specific functionality, and both manufacturers of tubes and laboratory professional strive to provide suitable interference-free sample matrices for laboratory testing. Both manufacturers of in vitro diagnostic devices and laboratory professionals need to understand all aspects of venous blood sampling so that they do not underestimate the impact of tube additives on laboratory testing.

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Leptospirosis is a neglected zoonosis, caused by pathogenic spirochetes bacteria of the genus Leptospira. The molecular mechanisms of leptospirosis infection are complex, and it is becoming clear that leptospires express several functionally redundant proteins to invade, disseminate, and escape the host’s immune response. Here, we describe a novel leptospiral protein encoded by the gene LIC13086 as an outer membrane protein. The recombinant protein LIC13086 can interact with the extracellular matrix component laminin and bind plasminogen, thus possibly participating during the adhesion process and dissemination. Also, by interacting with fibrinogen and plasma fibronectin, the protein LIC13086 probably has an inhibitory effect in the fibrin clot formation during the infection process. The newly characterized protein can also bind molecules of the complement system and the regulator C4BP and, thus, might have a role in the evasion mechanism of Leptospira. Taken together, our results suggest that the protein LIC13086 may have a multifunctional role in leptospiral pathogenesis, participating in host invasion, dissemination, and immune evasion processes.

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Resumen Se presenta el caso de un varón de 70 años de edad, quien debutó con fibrilación auricular no valvular, en quien, dos meses después y mediante ecocardiografía, se objetivaron dos masas contiguas en la aurícula derecha. Las masas eran dependientes de la válvula de Eustaquio, alcanzaban el septo interauricular y se asemejaban a un mixoma. Ambas fueron resecadas mediante cirugía, sin complicaciones intraoperatorias ni postoperatorias. El estudio anatomo-patológico concluyó que eran compatibles con trombos sin componente tumoral. Durante el seguimiento permaneció asintomático y en ritmo de fibrilación auricular.

Abstract The case is presented on a 70 year-old male who debuted with non-valvular atrial fibrillation. Two months later, and using cardiac ultrasound, two contiguous masses were observed in the right atrium. The masses were hanging from the Eustachian (inferior vena cave) valve, reached the interatrial septum and resembled a myxoma. Both were surgically resected, with no intra- or post-operative complications. The patient remained asymptomatic and in atrial fibrillation rhythm during follow-up.

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Musculoskeletal disorders are one of the major health burdens and a leading source of disability worldwide, affecting both juvenile and elderly populations either as a consequence of ageing or extrinsic factors such as physical injuries. This condition often involves a group of locomotor structures such as the bones, joints and muscles and may therefore cause significant economic and emotional impact. Some pharmacological and non-pharmacological treatments have been considered as potential solutions, however, these alternatives have provided quite limited efficacy due to the short-term effect on pain management and inability to restore damaged tissue. The emergence of novel therapeutic alternatives such as the application of orthobiologics, particularly bone marrow aspirate (BMA) clot, have bestowed medical experts with considerable optimism as evidenced by the significant results found in numerous studies addressed in this manuscript. Although other products have been proposed for the treatment of musculoskeletal injuries, the peculiar interest in BMA, fibrin clot and associated fibrinolytic mechanisms continues to expand. BMA is a rich source of various cellular and molecular components which have demonstrated positive effects on tissue regeneration in many in vitro and in vivo models of musculoskeletal injuries. In addition to being able to undergo self-renewal and differentiation, the hematopoietic and mesenchymal stem cells present in this orthobiologic elicit key immunomodulatory and paracrine roles in inflammatory responses in tissue injury and drive the coagulation cascade towards tissue repair via different mechanisms. Although promising, these complex regenerative mechanisms have not yet been fully elucidated.

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The hematopoietic niche is a specialized microenvironment that supports the survival, proliferation and differentiation of hematopoietic stem progenitor cells (HSPCs). Three-dimensional (3D) models mimicking hematopoiesis might allow in vitro and in vivo studies of the hematopoietic (HP) process. Here, we investigate the capacity of a 3D construct based on non-adherent murine bone marrow mononuclear cells (NA-BMMNCs), mesenchymal stromal cells (MSCs) and collagen microspheres (CMs), all embedded into plasma clot (PC) to support in vitro and in vivo hematopoiesis. Confocal analysis of the 3D hematopoietic construct (3D-HPC), cultured for 24 h, showed MSC lining the CM and the NA-BMMNCs closely associated with MSC. In vivo hematopoiesis was examined in 3D-HPC subcutaneously implanted in mice and harvested at different intervals. Hematopoiesis in the 3D-HPC was evaluated by histology, cell morphology, flow cytometry, confocal microscopy and hematopoietic colony formation assay. 3D-HPC implants were integrated and vascularized in the host tissue, after 3 months of implantation. Histological studies showed the presence of hematopoietic tissue with the presence of mature blood cells. Cells from 3D-HPC showed viability greater than 90%, expressed HSPCs markers, and formed hematopoietic colonies, in vitro. Confocal microscopy studies showed that MSCs adhered to the CM and NA-BMMNCs were scattered across the 3D-HPC area and in close association with MSC. In conclusion, the 3D-HPC mimics a hematopoietic niche supporting the survival, proliferation and differentiation of HSPCs, in vivo. 3D-HPC may allow evaluation of regulatory mechanisms involved in hematopoiesis.

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BACKGROUND: In healthy subjects fibrinogen γ/γ' circulates at 8-15% of the total plasma fibrinogen concentration. Elevated levels of this variant have been associated with arterial thrombosis, and its diminution with venous thrombosis. The aims of the present work were to analyze the structure of the fibrin network formed on the top of human dermal microvascular endothelial cells (HMEC-1) at different fibrinogen γ/γ' concentrations, as well as its influence on the secretion of fibrinolytic components. The kinetics of fibrin polymerization on top of HMEC-1 cells with 3, 10, and 30% fibrinogen γ/γ' was followed at 350 nm. The secretion of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type 1 (PAI 1) by HMEC-1 were measured in the supernatant and cell lysates, after incubation with 1 nM thrombin, fibrin with 3, and 30% fibrinogen γ/γ', using commercial kits. The influence of fibrinogen γ/γ' on fibrin structure on the surface of the HMEC-1 was followed with laser scanning confocal microscopy (LSCM). RESULTS: The kinetics of fibrin formation on HMEC-1 with 3 and 10% fibrinogen γ/γ' were similar. However, with 30% fibrinogen γ/γ' both the slope and final turbity were approximately 50% less. The LSCM images showed the dramatic effects of increasing fibrinogen γ/γ' from 3 to 30%. The uPA and PAI 1 concentrations in culture supernatants HMEC-1 cells treated with thrombin or 30% γ/γ' fibrin were two-fold increased as compared to basal culture supernatants and 3% γ/γ' fibrin-treated HMEC-1. In all stimulatory conditions the intracellular concentration of uPA was higher than in supernatants. In contrast, the intracellular PAI 1 concentration was decreased as compared to that measured in the supernatant, including the basal condition. CONCLUSION: A concentration of 30% fibrin γ/γ' alter drastically fibrin structure on the cell surface and affects the secretion of uPA and PAI 1 through its capacity to bind thrombin.

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