Occult cancer in patients with unprovoked venous thromboembolism: A nested case-control study.

OBJECTIVES: Detecting occult cancer in patients with unprovoked venous thromboembolism (VTE) remains a significant challenge. Our objective was to investigate the potential predictive role of coagulation-related biomarkers in the diagnosis of occult malignancies. METHODS: We conducted a nested case-control study with a 1-year prospective cohort of 214 patients with unprovoked VTE, with a focus on identifying occult cancer. At the time of VTE diagnosis, we measured various biomarkers, including soluble P-selectin (sP-selectin), dimerized plasmin fragment D (D-dimer), platelets, leukocytes, hemoglobin, total extracellular vesicles (EVs), EVs expressing tissue factor on their surface (TF+EVs), and EVs expressing P-selectin on their surface (Psel+EVs) in all participants. RESULTS: We observed statistically significant increased levels of sP-selectin (P = .015) in patients with occult cancer. Despite an increase in Psel+EVs, TF+EVs, D-dimer, and platelets within this group, however, no significant differences were found. When sP-selectin exceeded 62 ng/mL and D-dimer surpassed 10,000 µg/L, the diagnosis of occult cancer demonstrated a specificity of up to 91% (95% CI, 79.9%-96.7%). CONCLUSIONS: The combination of sP-selectin and D-dimer can be a valuable biomarker in detecting occult cancer in patients with unprovoked VTE. Further research is necessary to ascertain whether easily measurable biomarkers such as sP-selectin and D-dimer can effectively distinguish between patients who have VTE with and without hidden malignancies.

Presurgical circulating platelet-derived microparticles level as a risk factor of blood transfusion in patients with valve heart disease undergoing cardiac surgery.

BACKGROUND: Cell-derived microparticles (MPs) are membrane vesicles that have emerged as a potential biomarker for various diseases and their clinical complications. This study investigates the role of MPs as a risk factor for blood transfusion in patients with valve heart disease undergoing cardiac surgery. METHODS: Forty adult patients undergoing heart valve surgery with cardiopulmonary bypass (CPB) were enrolled, and venous blood samples were collected prior to surgical incision. Plasma rich in MPs was prepared by double centrifugation, and the concentration of MPs was determined using the Bradford method. Flow cytometry analysis was performed to determine MPs count and phenotype. Patients were divided into "with transfusion" (n = 18) and "without transfusion" (n = 22) groups based on red blood cell (RBC) transfusion. RESULTS: There was no significant difference in MPs concentration between the "with transfusion" and "without transfusion" groups. Although the count of preoperative platelet-derived MPs (PMPs), monocyte-derived MPs (MMPs), and red cell-derived MPs (RMPs) was higher in "without transfusion" group, these differences were not statistically significant. The preoperative PMPs count was negatively correlated with RBC transfusion (P = 0.005, r = -0.65). Multivariate logistic regression analysis revealed that the count of CD41+ PMPs, Hemoglobin (Hb), and RBC count were risk factors for RBC transfusion. CONCLUSION: This study suggests that the presurgical levels of PMPs, Hb, and RBC count can serve as risk factors of RBC transfusion in patients with valve heart disease undergoing cardiac surgery. The findings provide insights into the potential use of MPs as biomarkers for blood transfusion prediction in cardiac surgery.

Association of Acute Hyperglycemia and Diabetes Mellitus with Platelet-derived Microparticle (PDMP) Levels During Acute Myocardial Infarction.

Objectives: This research investigates whether there is an association between acute hyperglycemia and diabetes mellitus and the level of circulating platelet-derived microparticles (PDMPs) during an initial episode of acute myocardial infarction (AMI). Methodology: This was a cross-sectional study involving hospitalized AMI patients. Demographic and clinical data were obtained from hospital records. Diabetes mellitus was defined by the history of the disease, anti-diabetes medication use and/ or level of HbA1C ≥6.5%. Levels of HbA1c, admission random and fasting blood glucose levels were measured. Flow-cytometry method was used to determine the levels of PDMPs from collected venous blood through tagging with CD-41 FITC and CD-62 PE markers and a threshold size of <1 µm. The number of circulating PDMPs was compared according to glucometabolic state, namely acute hyperglycemia (admission random glucose ≥200 mg/dL and fasting glucose ≥140 mg/dL) and diabetes mellitus. The comparative analysis between groups was conducted with Student T-test or Mann-Whitney test, where applicable. Results: A total of 108 subjects were included and their data analyzed. The level of circulating PDMPs was significantly lower in subjects with admission random glucose ≥200 mg/dL as compared to those with below level [median (interquartile range (IQR)]: 2,710.0 (718.0-8,167.0) count/mL vs. 4,452.0 (2,128.5-14,499.8) count/mL, p = 0.05) and in subjects with fasting glucose ≥140 mg/dL as compared to those with below level (median (IQR): 2,382.0 (779.0-6,619.0) count/mL vs. 5,972.0 (2,345.7-14,781.3) count/mL, p = 0.006). The level of circulating PDMPs was also significantly lower in patients with diabetes mellitus as compared to those without (median (IQR): 2,655.0 (840.0-5,821.0) count/mL vs. 4,562.0 (2,128.5-15,055.8) count/mL; p = 0.007). Conclusion: Acute hyperglycemia and diabetes mellitus are significantly associated with a lower circulating PDMP level during an initial AMI episode.

Hydrophobicity-Adaptive Polymers Trigger Fission of Tumor-Cell-Derived Microparticles for Enhanced Anticancer Drug Delivery.

Tumor-cell-derived microparticles (MPs) can function as anticancer drug-delivery carriers. However, short blood circulation time, large-size-induced insufficient tumor accumulation and penetration into tumor parenchyma, as well as limited cellular internalization by tumor cells and cancer stem cells (CSCs), and difficult intracellular drug release restrict the anticancer activity of tumor-cell-derived MP-based drug-delivery systems. In this work, hydrophobicity-adaptive polymers based on poly(N-isopropylacrylamide) are anchored to tumor-cell-derived MPs for enhanced delivery of the anticancer drug doxorubicin (DOX). The polymers are hydrophilic in blood to prolong the circulation time of DOX-loaded MPs (DOX@MPs), while rapidly switching to hydrophobic at the tumor acidic microenvironment. The hydrophobicity of polymers drives the fission of tumor-cell-derived MPs to form small vesicles, facilitating tumor accumulation, deep tumor penetration, and efficient internalization of DOX@MPs into tumor cells and CSCs. Subsequently, the hydrophobicity of polymers in acidic lysosomes further promotes DOX release to nuclei for strong cytotoxicity against tumor cells and CSCs. The work provides a facile and simple strategy for improved anticancer drug delivery of tumor-cell-derived MPs.

Mechanism of trophoblast cell-derived microparticles mediated immunocontraceptive response.

PROBLEM: Immunomodulation profoundly affects the process of human implantation. Trophoblast cell-derived microparticles (Tr-MPs) may activate specific T cells to attack trophoblast cells, thus potentially acting as an immunocontraceptive vaccine. The safety and persistence of Tr-MP vaccine are needed to address. METHOD OF STUDY: Flow cytometry and confocal fluorescent microscopy were conducted to detect cellular absorptivity and localization of Tr-MPs in bone marrow-derived dendritic cells (BMDCs). The phenotype and cytokine secretion of BMDC and T cells were performed by flow cytometry and enzyme-linked immuno sorbent assay (ELISA). The constructed vaccine female moused model were used to observe the infertile effect and safety of Tr-MPs. RESULTS: As compared with non-irradiation exposure groups, the number of MPs released by trophoblast cells in ultraviolet immunized groups significantly increased. The phagocytosis of Tr-MPs led to the maturation of dendritic cells (DCs), which, in turn, activate T cells. Then cytotoxic T cells attacking trophoblast cells. In mouse model, female mice were infertile after receiving Tr-MPs, and the effect of contraception is transient and safety. CONCLUSION: Using Tr-MPs to initiate an adaptive immune response against alloantigens in trophoblast cells. Tr-MPs may be a new candidate for the development of contraceptive vaccines due to its effectiveness, safety, and reversibility.

Liquid Biopsy in Endometriosis: A Systematic Review.

Despite laparoscopy being a standardized option to diagnose pelvic endometriotic implants, non-invasive biomarkers are necessary to avoid the discomfort of invasive procedures. Recent evidence suggests a potential role of microRNAs (miRNAs) as feasible biomarkers for the early diagnosis of endometriosis. Following the recommendations in the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement, we systematically searched PubMed, EMBASE, Scopus, Cochrane Library, and Science Direct in January 2023. We provided no restriction on the country and year of publication, and considered English published articles. We selected studies including patients with endometriosis and describing miRNA regulation in the context of endometriosis. Overall, 45 studies fulfilled the inclusion criteria, and 2045 patients with endometriosis and 1587 controls were screened. Patients were analyzed concerning miRNAs expression and sources, stage of disease, and symptoms, and compared to controls. Among DEMs, the ones with the widest delta between endometriosis patients and controls-Relative Expression ≥ 4 Log2(ratio)-were miR-145, miR-191, miR-195, miR-21-5p, miR-106b-5p, miR-195-5p, miR-451a, miR-200c, miR-20a-5p, and miR-15a-5p. Although the epigenetic regulation is partially unclear, miRNAs are valid biomarkers to diagnose endometriotic lesions in symptomatic and non-symptomatic women. MiRNAs modulation should be clarified, especially during therapies or relapse, to plan targeted management protocols.

Paclitaxel-loaded tumor cell-derived microparticles improve radiotherapy efficacy in triple-negative breast cancer by enhancing cell killing and stimulating immunity.

Triple-negative breast cancer (TNBC) is a highly heterogeneous tumor characterized by high recurrence and metastasis, with a very poor prognosis, and there are still great challenges in its clinical treatment. Here, we describe the development of a novel modality for the treatment of TNBC with tumor cell-derived microparticles loaded with paclitaxel (MP-PTX) in combination with radiotherapy. We show that MP can deliver agents to tumor cells by homologous targeting, thereby increasing the absorption rate of the chemotherapeutic agent and enhancing its killing effects on tumor cells. We further demonstrate that MP-PTX combined with radiotherapy shows a synergistic antitumor effect by enhancing the inhibition of tumor cell proliferation, promoting tumor cell apoptosis, reducing the immunosuppressive microenvironment at the tumor site, and activating the antitumor immune response. Altogether, this study provides a referable and optional method for the clinical treatment of refractory tumors such as TNBC based on the combination of T-MP-delivered chemotherapeutic drugs and radiotherapy. Chemical compounds: paclitaxel (PTX), paclitaxel-loaded tumor cell-derived microparticles (MP-PTX).

Effect of lower-leg trauma and knee arthroscopy on procoagulant phospholipid-dependent activity.

Background: Lower-leg injury and knee arthroscopy are both associated with venous thromboembolism (VTE). The mechanism of VTE in both situations is unknown, including the role of procoagulant microparticles. This may provide useful information for individualizing thromboprophylactic treatment in both patient groups. Objective: We aimed to study the effect of (1) lower-leg trauma and (2) knee arthroscopy on procoagulant phospholipid-dependent (PPL) activity plasma levels. Methods: POT-(K)CAST trial participants who did not develop VTE were randomly selected for the current study. Plasma was collected shortly after lower-leg trauma or before and after knee arthroscopy. For aim 1, samples of 67 patients with lower-leg injury were compared with control samples (preoperative samples of 74 patients undergoing arthroscopy). Linear regression was used to obtain mean ratios (natural logarithm retransformed data), adjusted for age, sex, body mass index, infections, and comorbidities. For aim 2, pre- and postoperative samples of 49 patients undergoing arthroscopy were compared using paired t tests. PPL activity was measured using modified activated factor X-dependent PPL clotting assay. Results: For aim 1, PPL activity levels were almost threefold higher in patients with lower-leg injury compared with controls, that is, mean ratio, 2.82 (95% confidence interval [CI], 1.98-4.03). For aim 2, postoperative PPL activity levels did not change significantly, that is, mean change, -0.72 mU/mL (95% CI, -2.03 to 0.59). Conclusion: Lower-leg trauma was associated with increased plasma levels of PPL activity, in contrast to knee arthroscopy. Lower-leg trauma triggers the release of procoagulant microparticles.

Biodegradable electrospun nanofibrous platform integrating antiplatelet therapy-chemotherapy for preventing postoperative tumor recurrence and metastasis.

The perioperative trauma-related platelet recruitment and activation severely affect tumor recurrence and metastasis. Therefore, efficiently killing residual tumor cells and simultaneously inhibiting platelet activation to block platelet-cancer cell interaction might be a promising strategy to prevent postoperative tumor recurrence and metastasis. Methods: Biodegradable PLGA electrospun nanofibrous films co-delivering doxorubicin-loaded tumor repopulating cell-derived microparticles (DOX-MPs) and aspirin (ASA) were developed as the implant materials (DOX-MPs/ASA@NF) for postoperative in-situ treatment. The characterization, cytotoxicity against tumor cells, inhibition in platelet activation-triggered proliferation, migration and metastasis of tumor cells and in vivo anti-recurrence and anti-metastasis activity induced by DOX-MPs/ASA@NF were systematically evaluated. Results: PLGA nanofibrous films facilitate the enhanced distribution of DOX-MPs as well as DOX-MPs and ASA release in a time-programmed manner within the tumor resection cavity. The released DOX-MPs efficiently kill the residual tumor cells, while ASA decreases platelet activation and inhibits platelet-promoted proliferation, migration and metastasis of tumor cells, resulting in the remarkable inhibition of postoperative tumor recurrence and metastasis. Conclusions: DOX-MPs/ASA@NF may be a promising candidate to prevent the recurrence and metastasis of resectable tumors.

Increased suicidal erythrocyte death in patients with hepatitis B-related acute-on-chronic liver failure.

Anemia is a common complication of hepatitis B-related acute-on-chronic liver failure (HB-ACLF). Eryptosis, a suicidal erythrocyte death characterized by phosphatidylserine (PS) externalization and red blood cell-derived microparticle (RMP) generation, decreases erythrocyte lifespan. Herein, we investigated whether enhanced eryptosis is involved in the anemia pathophysiology associated with HB-ACLF. PS exposure, cell volume, cytosolic Ca2+, and reactive oxygen species (ROS) production were determined using flow cytometry. RMPs were extracted using a polyethylene glycol (PEG)-based method. We found that hemoglobin (Hb) and hematocrit (Hct) were significantly lower in patients with HB-ACLF than in healthy controls (HC), patients with chronic hepatitis B (CHB), and patients with cirrhosis. The direct antiglobulin test positive rate was 75.9% in patients with HB-ACLF while its intensity was associated with anemia. The ratio of abnormal erythrocytes was higher in patients with HB-ACLF than in HC, CHB, and cirrhosis. The percentage of PS-exposed erythrocytes was higher in patients with HB-ACLF (2.07 ± 0.11%) compared with HC (0.37 ± 0.05%), CHB (0.38 ± 0.03%), and cirrhosis (0.38 ± 0.04%). The cytosolic Ca2+ and ROS abundance were also higher in patients with HB-ACLF compared with HC, patients with CHB, and patients with cirrhosis, and were inversely correlated with the anemia in patients with HB-ACLF. PS exposure of erythrocytes collected from HC was significantly pronounced following incubation in plasma from patients with HB-ACLF compared with incubation in plasma from HC. The protein concentration and RMPs size significantly increased in patients with HB-ACLF compared with HC. Thus, the anemia in patients with HB-ACLF is associated with increased eryptosis, which is partially triggered by increased cytosolic Ca2+ and oxidative stress.NEW & NOTEWORTHY Acute chronic liver failure (ACLF) is a critical syndrome characterized by multiple organ failures and high short-term mortality. A common complication of HB-ACLF is anemia, however, the mechanism of anemia in HB-ACLF remains to be elucidated. We confirm that the accelerated eryptosis is involved in the pathophysiology of anemia associated with HB-ACLF, which progressively aggravates the clinical outcome. Our study illustrates the mechanism regarding the anemia pathogenesis of HB-ACLF, which may be utilized further toward therapeutic ends.

Assessment of extracellular vesicle isolation methods from human stool supernatant.

Extracellular vesicles (EVs) are of growing interest due to their potential diagnostic, disease surveillance, and therapeutic applications. While several studies have evaluated EV isolation methods in various biofluids, there are few if any data on these techniques when applied to stool. The latter is an ideal biospecimen for studying EVs and colorectal cancer (CRC) because the release of tumour markers by luminal exfoliation into stool occurs earlier than vascular invasion. Since EV release is a conserved mechanism, bacteria in stool contribute to the overall EV population. In this study, we assessed five EV separation methods (ultracentrifugation [UC], precipitation [EQ-O, EQ-TC], size exclusion chromatography [SEC], and ultrafiltration [UF]) for total recovery, reproducibility, purity, RNA composition, and protein expression in stool supernatant. CD63, TSG101, and ompA proteins were present in EV fractions from all methods except UC. Human (18s) and bacterial (16s) rRNA was detected in stool EV preparations. Enzymatic treatment prior to extraction is necessary to avoid non-vesicular RNA contamination. Ultrafiltration had the highest recovery, RNA, and protein yield. After assessing purity further, SEC was the isolation method of choice. These findings serve as the groundwork for future studies that use high throughput omics technologies to investigate the potential of stool-derived EVs as a source for novel biomarkers for early CRC detection.

Activation mechanism dependent surface exposure of cellular factor XIII on activated platelets and platelet microparticles.

BACKGROUND: Platelets contain a high amount of potentially active A subunit dimer of coagulation factor XIII (cellular FXIII; cFXIII). It is of cytoplasmic localization, not secreted, but becomes translocated to the surface of platelets activated by convulxin and thrombin (CVX+Thr). OBJECTIVE: To explore the difference in cFXIII translocation between receptor mediated and non-receptor mediated platelet activation and if translocation can also be detected on platelet-derived microparticles. Our aim was also to shed some light on the mechanism of cFXIII translocation. METHODS: Gel-filtered platelets were activated by CVX+Thr or Ca2+ -ionophore (calcimycin). The translocation of cFXIII and phosphatidylserine (PS) to the surface of activated platelets and platelet-derived microparticles was investigated by flow cytometry, immunofluorescence, and immune electron microscopy. Fluo-4-AM fluorescence was used for the measurement of intracellular Ca2+ concentration. RESULTS: Receptor mediated activation by CVX+Thr exposed cFXIII to the surface of more than 60% of platelets. Electron microscopy revealed microparticles with preserved membrane structure and microparticles devoid of labeling for membrane glycoprotein CD41a. cFXIII was observed on both types of microparticles but was more abundant in the absence of CD41a. Rhosin, a RhoA inhibitor, significantly decreased cFXIII translocation. Non-receptor mediated activation of platelets by calcimycin elevated intracellular Ca2+ concentration, induced the translocation of PS to the surface of platelets and microparticles, but failed to expose cFXIII. CONCLUSIONS: The elevation of intracellular Ca2+ concentration is sufficient for the translocation of PS from the internal layer of the membrane, while the translocation of cFXIII from the platelet cytoplasm requires additional receptor mediated mechanism(s).

Vitreous microparticles contain apoptotic signals suggesting a diabetic vitreopathy.

AIM: To evaluate differences in microparticle profiles in vitreous samples between diabetic and non-diabetic eyes undergoing vitrectomy. METHODS: Un-masked cross-sectional series of 34 eyes undergoing vitrectomy. Vitreous specimens were collected and processed to evaluate for membrane integrity (DAPI), apoptosis (Annexin-V), and endothelial-cell origin (V-Cadherin). A BD LSR II flow cytometer was used for analysis and standardized sub-micron-sized beads were used for size comparison. RESULTS: Thirty-four specimens underwent analysis. Greater levels of Annexin-V were found on microparticles from specimens in which blood had entered the vitreous (n=12) compared to those without blood (n=22; 52.3%±30.7% vs 19.6%±27.2%, P=0.002). Patients with diabetes having surgery with hemorrhage (n=7) had greater expression of Annexin-V than those without hemorrhage (n=8; 62.1%±31.7% vs 18.9%±20.9%, P=0.009). However, in patients with non-diabetic vitreous hemorrhage, the level of Annexin-V expression was not significantly different compared to other disease processes (38.6%±25.7%, n=5 vs 20.0%±30.9%, n=14, P=0.087). CONCLUSION: Increased expression of the apoptotic marker, Annexin-V is detected on vitreous microparticles in diabetes-related vitreous hemorrhage. When evaluating vitreous hemorrhage in patients without diabetes, the apoptotic signal is not significantly different. Vitrectomy in patients with diabetes, and improvement in visual outcomes, may be related to the removal of a serum-derived, pro-apoptotic vitreous. Further investigation is warranted in order to identify the molecular characteristics of microparticles that regulate disease.

Correlations between the Circulating Level of Cell-Derived Microparticles and Surgical Variables in Heart Valve Surgery with Cardiopulmonary Bypass.

Background: Cell-derived microparticles (MPs) as membrane vesicles are procoagulant. They play a role in surgical hemostasis. In this study, the correlations between the circulating level of cell-derived MPs and surgical variables in heart valve surgery were investigated. Methods: The present prospective case-series study was conducted in Rajaie Cardiovascular Medical and Research Center from January through March 2021. Forty patients undergoing heart valve surgery with cardiopulmonary bypass (CPB) were enrolled. Before the induction of anesthesia and 30 minutes after the administration of protamine sulfate, venous blood samples were collected. After MP isolation, the concentration of MPs was determined via the Bradford method. Flow cytometry analysis was performed to determine the MP count and phenotype. Intraoperative variables and postoperative routine coagulation tests were defined as surgical variables. Postoperative coagulopathy was defined as an activated partial thromboplastin time (aPTT) ≥48 seconds or an international normalized ratio (INR) >1.5. Results: The total concentration of MPs and the MP count increased significantly after surgery compared with before surgery. The postoperative concentration of MPs was positively correlated with the CPB time (P=0.030, ρ=0.40). The preoperative concentration of MPs was significantly lower in patients with higher postoperative aPTT and INR (P=0.003, P= -0.50 and P=0.020, P= -0.40, respectively). In multivariate logistic regression analysis, the preoperative MP concentration (OR, 1.00; 95% CI, 1.00 to 1.01; P=0.017) was considered a risk factor for postoperative coagulopathy. Conclusion: The levels of MPs, especially platelet-derived MPs, rose after surgery, in correlation with the CPB time. Given the role of MPs in the induction of coagulation and inflammation, they can be considered therapeutic goals for preventing postoperative complications. In addition, the preoperative levels of MPs are a risk factor for predicting the occurrence of postoperative coagulopathy in heart valve surgery.

Challenges and opportunities in the study of extracellular vesicles: Global institutional context and national state of the art / Retos y oportunidades en el estudio de vesículas extracelulares: contexto institucional a nivel mundial y situación actual en Colombia

In the last decade, the number of studies and publications on extracellular vesicles (EV) and exosomes has boomed. Colombia has displayed interest and progress in their study as shown in the increase of research project publications and products. However, this research field is still developing and has its own analytical challenges and technical limitations. For planning research projects and developing EV studies it is necessary to consider what is the state of the scientific field worldwide concerning EV nomenclature and classification, available techniques, resources, requirements and quality specifications, and the institutions that regulate the field. Answering this question will elicit EV studies that comply with international standards and respond to institutional demands and recommendations. However, the scientific information available is scattered and not all the aspects are considered in full. In this update, the available information is condensed and the official terms and currently defined nomenclature is presented, as well as the evolution of the field, the homogenization of the experimental parameters, the establishment of scientific authorities, institutions, and resources, and the recommendations generated worldwide for their development and research including their isolation, characterization, and functional studies. Finally, I analyzed the national context in a critical way, considering institutional strengths, common mistakes, and available analytical techniques and technologies.

En la última década se ha incrementado el número de estudios y publicaciones sobre las vesículas extracelulares y los exosomas. En Colombia, ha habido interés y avances en su estudio, lo que se evidencia en el aumento de publicaciones y proyectos de investigación. Sin embargo, este es un campo de investigación aún en desarrollo, con desafíos analíticos y limitaciones técnicas, por lo cual, en el planteamiento de los proyectos de investigación y desarrollo, es necesario considerar cuál es el estado del campo científico a nivel mundial en cuanto a la nomenclatura y la clasificación de las vesículas extracelulares, las técnicas, recursos, requisitos y especificaciones de calidad y las instituciones que regulan el campo. La respuesta a esta pregunta permitirá desarrollar estudios que cumplan con los estándares internacionales, y las exigencias y recomendaciones institucionales. Sin embargo, la información científica disponible se encuentra dispersa y no todos los aspectos son tratados a cabalidad. En este actualización se condensa la información disponible y se presentan los términos oficiales para denominar las vesículas extracelulares y la nomenclatura aceptada actualmente, así como la evolución del campo, la homogenización de los parámetros experimentales, el establecimiento de autoridades científicas, instituciones y recursos, y las recomendaciones que se han generado a nivel mundial para el desarrollo de investigaciones en vesículas extracelulares, incluidos su aislamiento, caracterización y estudio funcional. Por último, se analiza el contexto nacional de una forma crítica, teniendo en cuenta las fortalezas institucionales, los errores usualmente cometidos, y las técnicas y tecnologías analíticas disponibles.

Retos y oportunidades en el estudio de vesículas extracelulares: contexto institucional a nivel mundial y situación actual en Colombia / Challenges and opportunities in the study of extracellular vesicles: Global institutional context and national state of the art

Resumen En la última década se ha incrementado el número de estudios y publicaciones sobre las vesículas extracelulares y los exosomas. En Colombia, ha habido interés y avances en su estudio, lo que se evidencia en el aumento de publicaciones y proyectos de investigación. Sin embargo, este es un campo de investigación aún en desarrollo, con desafíos analíticos y limitaciones técnicas, por lo cual, en el planteamiento de los proyectos de investigación y desarrollo, es necesario considerar cuál es el estado del campo científico a nivel mundial en cuanto a la nomenclatura y la clasificación de las vesículas extracelulares, las técnicas, recursos, requisitos y especificaciones de calidad y las instituciones que regulan el campo. La respuesta a esta pregunta permitirá desarrollar estudios que cumplan con los estándares internacionales, y las exigencias y recomendaciones institucionales. Sin embargo, la información científica disponible se encuentra dispersa y no todos los aspectos son tratados a cabalidad. En este actualización se condensa la información disponible y se presentan los términos oficiales para denominar las vesículas extracelulares y la nomenclatura aceptada actualmente, así como la evolución del campo, la homogenización de los parámetros experimentales, el establecimiento de autoridades científicas, instituciones y recursos, y las recomendaciones que se han generado a nivel mundial para el desarrollo de investigaciones en vesículas extracelulares, incluidos su aislamiento, caracterización y estudio funcional. Por último, se analiza el contexto nacional de una forma crítica, teniendo en cuenta las fortalezas institucionales, los errores usualmente cometidos, y las técnicas y tecnologías analíticas disponibles.

Abstract In the last decade, the number of studies and publications on extracellular vesicles (EV) and exosomes has boomed. Colombia has displayed interest and progress in their study as shown in the increase of research project publications and products. However, this research field is still developing and has its own analytical challenges and technical limitations. For planning research projects and developing EV studies it is necessary to consider what is the state of the scientific field worldwide concerning EV nomenclature and classification, available techniques, resources, requirements and quality specifications, and the institutions that regulate the field. Answering this question will elicit EV studies that comply with international standards and respond to institutional demands and recommendations. However, the scientific information available is scattered and not all the aspects are considered in full. In this update, the available information is condensed and the official terms and currently defined nomenclature is presented, as well as the evolution of the field, the homogenization of the experimental parameters, the establishment of scientific authorities, institutions, and resources, and the recommendations generated worldwide for their development and research including their isolation, characterization, and functional studies. Finally, I analyzed the national context in a critical way, considering institutional strengths, common mistakes, and available analytical techniques and technologies.

Centrifugation Removes a Population of Large Vesicles, or "Macroparticles," Intermediate in Size to RBCs and Microvesicles.

Microparticles or microvesicles (MPs/MVs) are sub-cellular vesicles with a growing number of known biological functions. Microvesicles from a variety of parent cells within the vascular system increase in numerous pathological states. Red blood cell-derived MVs (RMVs) are relatively less studied than other types of circulating MVs despite red blood cells (RBCs) being the most abundant intravascular cell. This may be in part due the echoes of past misconceptions that RBCs were merely floating anucleate bags of hemoglobin rather than dynamic and responsive cells. The initial aim of this study was to maximize the concentration of RMVs derived from various blood or blood products by focusing on the optimal isolation conditions without creating more MVs from artificial manipulation. We found that allowing RBCs to sediment overnight resulted in a continuum in size of RBC membrane-containing fragments or vesicles extending beyond the 1 µm size limit suggested by many as the maximal size of an MV. Additionally, dilution and centrifugation factors were studied that altered the resultant MV population concentration. The heterogeneous size of RMVs was confirmed in mice models of hemolytic anemia. This methodological finding establishes a new paradigm in that it blurs the line between RBC, fragment, and RMV as well as suggests that the concentration of circulating RMVs may be widely underestimated given that centrifugation removes the majority of such RBC-derived membrane-containing particles.

Shedding of proangiogenic microvesicles from hypertrophic scar myofibroblasts.

Hypertrophic scars are a common complication of burn injuries and represent a major challenge in terms of prevention and treatment. These scars are characterized by a supraphysiological vascular density and by the presence of pathological myofibroblasts (Hmyos) displaying a low apoptosis propensity. However, the nature of the association between these two hallmarks of hypertrophic scarring remains largely unexplored. Here, we show that Hmyos produce signalling entities known as microvesicles that significantly increase the three cellular processes underlying blood vessel formation: endothelial cell proliferation, migration and assembly into capillary-like structures. The release of microvesicles from Hmyos was dose-dependently induced by the serum protein α-2-macroglobulin. Using flow cytometry, we revealed the presence of the α-2-macroglobulin receptor-low-density lipoprotein receptor-related protein 1-on the surface of Hmyos. The inhibition of the binding of α-2-macroglobulin to its receptor abolished the shedding of proangiogenic microvesicles from Hmyos. These findings suggest that the production of microvesicles by Hmyos contributes to the excessive vascularization of hypertrophic scars. α-2-Macroglobulin modulates the release of these microvesicles through interaction with low-density lipoprotein receptor-related protein 1.

Level and clinical significance of vitreous microparticles in proliferative diabetic retinopathy / 中华眼底病杂志

Objective:To observe the level of microparticles in the vitreous of patients with proliferative diabetic retinopathy (PDR), and preliminarily explore the role of microparticles in the pathogenesis of PDR.Methods:A case control study. From January to December 2018, 54 cases of 54 eyes of PDR patients (PDR group) and 20 cases of non-diabetic retinopathy patients (control group), who were diagnosed and treated with vitrectomy (PPV) in the Department of Ophthalmology, Tianjin Medical University General Hospital vitreous samples were included in the study. Among 54 eyes in the PDR group, there were 42, 21, and 17 eyes with vitreous hemorrhage (VH), traction retinal detachment (TRD), and previous intravitreal injection of drugs, respectively. Among the 20 eyes of the control group, idiopathic macular hole, idiopathic anterior macular membrane, vitreous macular traction syndrome, and complete lens dislocation were 6, 6, 2, and 6 eyes, respectively. The PDR group was divided into uncombined TRD group and combined TRD group according to PDR stage and whether TRD occurred, with 33 and 21 eyes, respectively. According to the presence or absence of VH, they were divided into groups with VH and without VH, with 42 eyes and 12 eyes, respectively. According to whether anti-vascular endothelial growth factor (VEGF) drugs were injected into the intravitreal cavity 3 days before PPV, they were divided into anti-VEGF drug group and no anti-VEGF drug group, with 17 eyes and 37 eyes respectively. The levels of retinal photoreceptor cells (RMP), platelets (PMP), endothelial cells (EMP) and phosphatidylserine (PS-MP) expressing on the membrane surface in the sample were detected by flow cytometry. The comparison between the two groups of samples was performed by t test, and the comparison between multiple groups of samples was performed by one-way analysis of variance or Mann-Whitney test. Results:Compared with the control group, the vitreous RMP level of the PDR group was significantly decreased, and the EMP and PMP levels were significantly increased. The differences were statistically significant ( t=-2.361, 5.064, 3.531; P=0.018, <0.001, 0.001). There was no statistically significant difference in PS-MP levels between the two groups ( t=-1.617, P=0.110). Compared with the TRD group, the levels of RMP and PMP in the vitreous of the TRD group were significantly increased, and the difference was statistically significant ( t=-2.221, -2.098; P=0.031, 0.041). The level of EMP in the vitreous body of the anti-VEGF drug group was significantly lower than that of the non-anti-VEGF drug group, however, it was still higher than the control group. The difference was statistically significant ( Z=-2.430, -2.499; P=0.015, 0.012). The level of PMP in the vitreous body of the eye without VH was significantly higher than that in the group with VH, and the difference was statistically significant ( t=-3.097, P=0.003). Conclusions:The elevated levels of EMP and PMP in the vitreous of PDR patients may be related to the damage of retinal capillaries; intravitreal injection of anti-VEGF drugs before surgery can reduce the level of EMP. VH may be related to the procoagulant effect of PMP.