Immunothrombotic dysregulation in chagas disease and COVID-19: a comparative study of anticoagulation.

Chagas and COVID-19 are diseases caused by Trypanosoma cruzi and SARS-CoV-2, respectively. These diseases present very different etiological agents despite showing similarities such as susceptibility/risk factors, pathogen-associated molecular patterns (PAMPs), recognition of glycosaminoglycans, inflammation, vascular leakage hypercoagulability, microthrombosis, and endotheliopathy; all of which suggest, in part, treatments with similar principles. Here, both diseases are compared, focusing mainly on the characteristics related to dysregulated immunothrombosis. Given the in-depth investigation of molecules and mechanisms related to microthrombosis in COVID-19, it is necessary to reconsider a prompt treatment of Chagas disease with oral anticoagulants.

The influence of hydrogen ions on coagulation in traumatic brain injury, explored by molecular dynamics.

Background: Patients in intensive care units with traumatic brain injuries (TBI) frequently present acid-base abnormalities and coagulability disorders, which complicate their condition.Objective: To identify protonation through in silico simulations of molecules involved in the process of coagulation in standard laboratory tests.Materials and methods: Ten patients with TBI were selected from the intensive care unit in addition to ten "healthy control subjects", and another nine patients as "disease control subjects"; the latter being a comparative group, corresponding to subjects with diabetes mellitus 2 (DM2). Fibrinogen, FVII, FVIII, FIX, FX, and D-dimer in the presence of acidification were evaluated in 20 healthy subjects in order to compare clinical results with molecular dynamics (MD), and to explain proton interactions and coagulation molecules.Results: The TBI group presented a slight, non-significant increase in D-dimer; but this was not present in "disease control subjects". Levels of fibrinogen, FVII, FIX, FX, and D-dimer were affected in the presence of acidification. We observed that various specific residues of coagulation factors "trap" ions.Conclusion: Protonation of tissue factor and factor VIIa may favor anticoagulant mechanisms, and protonation does not affect ligand binding sites of GPIIb/IIIa (PAC1) suggesting other causes for the low affinity to PAC1.

Analysis of SARS-CoV-2 mutations in Mexico, Belize, and isolated regions of Guatemala and its implication in the diagnosis.

The genomic sequences of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) worldwide are publicly available and are derived from studies due to the increase in the number of cases. The importance of study of mutations is related to the possible virulence and diagnosis of SARS-CoV-2. To identify circulating mutations present in SARS-CoV-2 genomic sequences in Mexico, Belize, and Guatemala to find out if the same strain spread to the south, and analyze the specificity of the primers used for diagnosis in these samples. Twenty three complete SARS-CoV-2 genomic sequences, available in the GISAID database from May 8 to September 11, 2020 were analyzed and aligned versus the genomic sequence reported in Wuhan, China (NC_045512.2), using Clustal Omega. Open reading frames were translated using the ExPASy Translate Tool and UCSF Chimera (v.1.12) for amino acid substitutions analysis. Finally, the sequences were aligned versus primers used in the diagnosis of COVID-19. One hundred and eighty seven distinct variants were identified, of which 102 are missense, 66 synonymous and 19 noncoding. P4715L and P5828L substitutions in replicase polyprotein were found, as well as D614G in spike protein and L84S in ORF8 in Mexico, Belize, and Guatemala. The primers design by CDC of United States showed a positive E value. The genomic sequences of SARS-CoV-2 in Mexico, Belize, and Guatemala present similar mutations related to a virulent strain of greater infectivity, which could mean a greater capacity for inclusion in the host genome and be related to an increased spread of the virus in these countries, furthermore, its diagnosis would be affected.

Molecules and Prostaglandins Related to Embryo Tolerance.

It is generally understood that the entry of semen into the female reproductive tract provokes molecular and cellular changes facilitating conception and pregnancy. We show a broader picture of the participation of prostaglandins in the fertilization, implantation and maintenance of the embryo. A large number of cells and molecules are related to signaling networks, which regulate tolerance to implantation and maintenance of the embryo and fetus. In this work, many of those cells and molecules are analyzed. We focus on platelets, polymorphonuclear leukocytes, and group 2 innate lymphoid cells involved in embryo tolerance in order to have a wider view of how prostaglandins participate. The combination of platelets and neutrophil extracellular traps (Nets), uterine innate lymphoid cells (uILC), Treg cells, NK cells, and sex hormones have an important function in immunological tolerance. In both animals and humans, the functions of these cells can be regulated by prostaglandins and soluble factors in seminal plasma to achieve an immunological balance, which maintains fetal-maternal tolerance. Prostaglandins, such as PGI2 and PGE2, play an important role in the suppression of the previously mentioned cells. PGI2 inhibits platelet aggregation, in addition to IL-5 and IL-13 expression in ILC2, and PGE2 inhibits some neutrophil functions, such as chemotaxis and migration processes, leukotriene B4 (LTB4) biosynthesis, ROS production, and the formation of extracellular traps, which could help prevent trophoblast injury and fetal loss. The implications are related to fertility in female when seminal fluid is deposited in the vagina or uterus.

TSH Levels in Subclinical Hypothyroidism in the 97.5th Percentile of the Population.

The debate regarding the cutoff point in the treatment of patients with subclinical hypothyroidism (Shypo) is ongoing. Generally, two different groups are identified for treatment by levels of 10 and 20 mIU/L. Nevertheless, the question remains, "what cutoff point should be chosen?" We have written a selective nonsystematic review focused on the 97.5 percentile reference value reported in healthy subjects in a number of countries and observed important disparities, which partly show the challenge of identifying a single cutoff point for those patients needing medication. We identified studies of TSH on the natural history of subclinical hypothyroidism from population-based prospective cohort studies, which follow up patients for several years. The evolution of TSH levels in these patients is variable. Some cases of TSH may return to lower levels at different stages over the years, but others may not, possibly even developing into overt thyroid failure, also variable. We analyzed factors that may explain the normalization of serum TSH levels. In addition, we found that thorough population-based prospective cohort studies following up on TSH levels, thyroid antibodies, and ultrasonography are important in decisions made in the treatment of patients. However, the 97.5 percentile reference value varies in different countries; therefore, an international cutoff point for subclinical hypothyroidism cannot be recommended.

Aggregation and Molecular Properties of ß-Glucosidase Isoform II in Chayote (Sechium edule).

The presence of isoforms of ß-glucosidase has been reported in some grasses such as sorghum, rice and maize. This work aims to extract and characterize isoform II in ß-glucosidase from S. edule. A crude extract was prepared without buffer solution and adjusted to pH 4.6. Contaminating proteins were precipitated at 4 °C for 24 h. The supernatant was purified by chromatography on carboxymethyl cellulose (CMC) column, molecular exclusion on Sephacryl S-200HR, and exchange anionic on QFF column. Electrophoretic analyzes revealed a purified enzyme with aggregating molecular complex on SDS-PAGE, Native-PAGE, and AU-PAGE. Twelve peptides fragments were identified by nano liquid chromatography-tandem mass spectrometry (nano LC-ESI-MS/MS), which presented as 61% identical to Cucurbita moschata ß-glucosidase and 55.74% identical to ß-glucosidase from Cucumis sativus, another Cucurbitaceous member. The relative masses which contained 39% hydrophobic amino acids ranged from 982.49 to 2,781.26. The enzyme showed a specificity to ß-d-glucose with a Km of 4.59 mM, a Vmax value of 104.3 µM∙min-1 and a kcat of 10,087 µM∙min-1 using p-nitrophenyl-ß-D-glucopyranoside. The presence of molecular aggregates can be attributed to non-polar amino acids. This property is not mediated by a ß-glucosidase aggregating factor (BGAF) as in grasses (maize and sorghum). The role of these aggregates is discussed.

Platelets and Platelet-Derived Microvesicles as Immune Effectors in Type 2 Diabetes.

The association between type 2 diabetes mellitus (T2DM) and systemic inflammation may increase platelet reactivity and the accelerated development of vascular disease. Platelets are able to modulate the function of immune cells via the direct release of growth factors and pro-inflammatory chemokines through the production of microvesicles. The microvesicles trigger a transcellular delivery system of bioactive molecules to other cells acting as vectors in the exchange of biological information. Here, we consider the influence of platelets and platelet-derived microvesicles on cells of the immune system and the implications in the pathogenesis of T2DM.

Molecules in seminal plasma related to platelets in preeclampsia.

OBJECTIVE: Prostaglandins present in seminal fluid are actively involved in vascular and non-vascular smooth muscle maintenance, reproduction, and inflammatory processes. Seminal plasma contains molecules, such as oxylipins, which possess cell signaling functions. Several studies have shown that specific molecules in seminal fluid can increase passive diffusion, and cause interactions in the female reproductive tract. This may provoke a cascade of cellular and molecular changes in general health and certain diseases. This study examines the hypothesis that the molecules in seminal fluid are involved in platelet activity. The molecules diffuse through cells and membranes, affecting Hoxa 10, binding ganglioside pathways, and acting over platelet function. When these molecules are at low levels, they may trigger prothrombotic states, explaining the pathophysiology of haemostatic response, such as preeclampsia, and increased risk of cardiovascular disease.

Purification and Partial Characterization of ß-Glucosidase in Chayote (Sechium edule).

ß-Glucosidase (EC 3.2.1.21) is a prominent member of the GH1 family of glycoside hydrolases. The properties of this ß-glucosidase appear to include resistance to temperature, urea, and iodoacetamide, and it is activated by 2-ME, similar to other members. ß-Glucosidase from chayote (Sechium edule) was purified by ionic-interchange chromatography and molecular exclusion chromatography. Peptides detected by LC-ESI-MS/MS were compared with other ß-glucosidases using the BLAST program. This enzyme is a 116 kDa protein composed of two sub-units of 58 kDa and shows homology with Cucumis sativus ß-glucosidase (NCBI reference sequence XP_004154617.1), in which seven peptides were found with relative masses ranging from 874.3643 to 1587.8297. The stability of ß-glucosidase depends on an initial concentration of 0.2 mg/mL of protein at pH 5.0 which decreases by 33% in a period of 30 h, and then stabilizes and is active for the next 5 days (pH 4.0 gives similar results). One hundred µg/mL ß-D-glucose inhibited ß-glucosidase activity by more than 50%. The enzyme had a Km of 4.88 mM with p-NPG and a Kcat of 10,000 min(-1). The optimal conditions for the enzyme require a pH of 4.0 and a temperature of 50 °C.

Better detection of platelet aggregation in patients with metabolic syndrome using epinephrine and ADP.

BACKGROUND: Patients with metabolic syndrome (MS) often have increased platelet aggregation. In order to determine which concentration detects a higher level of platelet aggregation in patients with MS, the agonists ADP and epinephrine were compared. METHODS: The study included 56 subjects with MS and 53 healthy subjects. Blood pressure, weight, body-mass index, and hip-to-waist ratio were collected from all subjects. Insulin, glucose, total serum cholesterol, HDL-C, LDL-C, total triglycerides, markers of plasma atherogenicity, and indices of insulin resistance were measured in all participants. For aggregometry assays, the Born method was used. Platelets were treated with ADP and epinephrine in decreasing concentrations of 2.34, 1.17, and 0.58 µM, as well as, 11.0, 1.1, and 0.55 µM, respectively. ROC curves were plotted to define the diagnostic efficiency of epinephrine levels for MS. RESULTS: Among healthy individuals and MS patients significant differences were observed in body weight, body-mass index, waist-circumference, levels of insulin, indices of insulin resistance, and levels of HDL-cholesterol, LDL-cholesterol and total triglycerides. There was a significant difference in the detection of increased platelet aggregation using 11.0 µM and 0.55 µM epinephrine and 0.58 µM ADP. With both agonists, ROC analysis showed an area under the curve of >0.8 for 11.0 µM epinephrine and 2.34 µM ADP. However, for MS patients, 11.0 µM epinephrine had a slightly better diagnostic efficiency than 2.34 µM ADP. CONCLUSIONS: It was found that 11.0 µM epinephrine and 2.34 µM ADP detected better platelet aggregation in patients with MS than in healthy subject. Both concentrations detected increased platelet aggregation in patients with MS.

Protein C activation peptide inhibits the expression of ICAM-1, VCAM-1, and interleukin-8 induced by TNF-α in human dermal microvascular endothelial cells.

Activated protein C (APC) is generated from the cleavage of protein C by thrombin coupled to thrombomodulin and, subsequently, is released as protein C activation peptide (papC). The aim of this study was to evaluate the effect of papC on human dermal microvascular endothelial cells (HMEC-1), activated with 5 ng//mL TNF-α. Flow cytometry showed that papC inhibited the expression of VCAM-1 and ICAM-1, after activation with TNF-a. Similarly, RT-PCR analysis revealed that 2 and 4 pM papC inhibited the expression of VCAM-1 and IL-8 mRNA in TNF-α-treated HMEC-1. In addition, the expression of endothelial nitric oxide synthase(eNOS) increased in HMEC-1 treated with papC, compared to those without treatment. Furthermore, Jurkat cell adhesion to HMEC-1 induced by TNF-a was significantly inhibited after the addition of papC, compared to HMEC-1 without papC (p = 0.03). Finally, a control peptide analog to papC showed no effect on the expression of ICAM and VCAM on the surface of HMEC-1. In conclusion, our results suggest that papC exerts anti-inflammatory effects on endothelial cells.

A study on inorganic elements in psammomas from ovarian & thyroid cancer.

BACKGROUND & OBJECTIVES: Concentric lamellar calcifications known as psammoma bodies (PB) are found in benign and malignant tumours. Whether or not the inorganic element concentrations in psammomas are similar to serous adenocarcinoma of the ovary and thyroid papillary cancer tissues has not yet been ascertained. We undertook this retrospective study to establish if there is any difference in the concentrations of inorganic ions found in psammomas in serous adenocarcinoma of the ovary, and those found in thyroid papillary cancer tissue. METHODS: PB samples from patients with adenocarcinoma of the ovary (n = 10) and with thyroid papillary cancer (n = 10) were analyzed through inductively-coupled plasma spectroscopy (ICP). RESULTS: There were no significant differences in the concentrations of inorganic elements in PB from thyroid papillary cancer than in those PB from ovarian cancer. INTERPRETATION & CONCLUSIONS: Differences in the concentrations of inorganic elements may be due to the variation in environmental pollution. Our study had limitation of small sample size. Our results suggest that some inorganic elements can participate in the origin of psammoma bodies.

Color of meconium and interleukin-6.

OBJECTIVE: To test the hypothesis that the color of meconial fluid is associated with inflammatory biomarkers, by determining C-reactive protein (CRP) and Interleukin-6 (IL-6) in serum from the umbilical cord. METHODS: In this prospective study, the authors selected 30 newborns with meconium-stained amniotic fluid (MSAF): 14 with green/brown 656 R color and 16 with brown/cinnamon 654 R color, and 20 newborns which showed clear amniotic fluid without MSAF (non-MSAF); all newborns were from mothers without risk factors for neonatal sepsis. RESULTS: IL-6 concentration from umbilical cord blood, [median of 12.9 pg/mL (interquartile range {IQR} 8.7-31.0)] of MSAF-green/brown 656 R increased significantly (p < 0.05) when compared with IL-6 concentration, [median of 9.2 pg/mL (IQR 7.2-12.2)] of newborns with clear amniotic fluid and without meconium. CRP from MSAF-green/brown 656 R was median of 0.5 mg/mL (IQR 0.0-2.7), and median of 1.0 mg/mL (IQR 0.0-5.5) from clear amniotic fluid, without meconium. CONCLUSIONS: Significant association was found between MSAF-green/brown 656 R and increase in IL-6, with normal CRP values.

The relationship of aluminium and silver to neural tube defects; a case control.

OBJECTIVE: The purpose of this study was to identify the relationship of neurotoxic inorganic elements in the hair of patients with the diagnosis of Neural Tube Defects. Our initial hypothesis was that neurotoxic inorganic elements were associated with Neural Tube Defects. METHODS: Twenty-three samples of hair from newborns were obtained from the General Hospital, "Aurelio Valdivieso" in the city of Oaxaca, Mexico. The study group included 8 newborn infants with neural tube pathology. The control group was composed of 15 newborns without this pathology. The presence of inorganic elements in the hair samples was determined by inductively-coupled plasma spectroscopy (spectroscopic emission of the plasma). FINDINGS: THE POPULATION OF NEWBORNS WITH NEURAL TUBE DEFECTS SHOWED SIGNIFICANTLY HIGHER VALUES OF THE FOLLOWING ELEMENTS THAN THE CONTROL GROUP: Aluminium, Neural Tube Defects 152.77±51.06 µg/g, control group 76.24±27.89 µg/g; Silver, Neural Tube Defects 1.45±0.76, control group 0.25±0.53 µg/g; Potassium, Neural Tube Defects 553.87±77.91 µg/g, control group 341.13±205.90 µg/g. Association was found at 75 percentile between aluminium plus silver, aluminium plus potassium, silver plus potassium, and potassium plus sodium. CONCLUSION: IN THE HAIR OF NEWBORNS WITH NEURAL TUBE DEFECTS, THE FOLLOWING METALS WERE INCREASED: aluminium, silver. Given the neurotoxicity of the same, and association of Neural Tube Defects with aluminum and silver, one may infer that they may be participating as factors in the development of Neural Tube Defects.

Lectin activity of the coagulation factor VIII/von Willebrand complex.

The human coagulation factor VIII (FVIII) is essential in the intrinsic pathway of blood coagulation and circulates mainly as a non-covalently bound complex with the von Willebrand factor (VWF). This complex (FVIII/VWF) protects FVIII from degradation and cellular uptake, although no biological role has been identified yet for this complex. The FVIII/VWF complex was purified from a healthy donor's plasma by affinity chromatography on a Sepharose 4B-Concanavalin A column and was used to determine its capability to interact with erythrocytes and platelets. The purified FVIII/VWF complex at 6.0 and 12 microg/ml agglutinates rabbit and bovine erythrocytes, and showed negative agglutination with erythrocytes from other species including human ABO. Treatment of erythrocytes with Clostridium perfringens sialidase or trypsin increased four-fold the activity toward rabbit erythrocytes and positive agglutination for human A and B erythrocytes, suggesting the presence of FVIII/VWF-cryptic receptors in these erythrocytes. Goat, pig, or human O erythrocytes were not agglutinated even after enzymatic treatment. Fucose or N-acetyl-glucosamine (GlcNAc), at 10 mM, inhibited agglutinating activity of the complex with rabbit, human A and B erythrocytes, whereas galactose and N-acetyl-galactosamine, even at 200 mM, showed no effect on the complex activity. The FVIII/VWF complex, at 1.5 microg/200,000 platelets, significantly decreased platelet aggregation (p < 0.001) when compared with the effect of platelet-rich plasma; this effect was inhibited with 15 mM GlcNAc or fucose. ELISA assays on FVIII/VWF coated polystyrene plates confirmed specific binding to fucose- or biotinylated GlcNAc-dextran derivatives. We therefore propose that the FVIII/VWF complex possesses lectin activity.

Potential use of the Macrobrachium rosenbergii lectin for diagnosis of T-cell acute lymphoblastic leukemia.

T-cell acute lymphoblastic leukemia is the most common form of cancer in children. Lectins are proteins or glycoproteins from plants or animals that recognize oligossacharides on the cell surface and have been used to characterize the structural changes of oligosaccharides in leukemias. In this study, we used the lectin from the freshwater prawn Macrobrachium (M. rosenbergii), specific for acetyl groups in sialylated glycans, because increased sialylation of glycoproteins and glycolipids has been identified in lymphoblastic leukemias. We compared the specificity of the M. rosenbergii lectin for lymphoblastic leukemias with the specificities of the lectins from Triticum vulgaris, Solanum tuberosum, Arachis hipogaea, and Phytolacca americana. By morphologic and phenotype characterization with a panel of monoclonal antibodies, we identified four types of leukemias from 106 leukemia patients: 11 cases of T-cell acute lymphoblastic leukemia, 61 cases of B-cell acute lymphoblastic leukemia, 24 cases of acute myeloblastic leukemia, and 10 cases of acute biphenotypic leukemia. As determined by cytofluorometric assays, nine of the eleven cases with T-cell acute lymphoblastic leukemia (8 +/- 3 years old) were specifically identified with the lectin from M. rosenbergii. In contrast, only six cases of B-cell leukemia, one case of myeloblastic leukemia, and 2 cases of biphenotypic leukemia were identified with this M. rosenbergii lectin. The other lectins tested showed no capacity to differentiate, in a significant manner, any of the four types of leukemias tested. Thus, the lectin from M. rosenbergii could be considered a useful tool for the diagnosis and study of T-cell acute lymphoblastic leukemia.

Role of concanavalin A lectin in recognition of pterygium remnant after surgical excision: preliminary results of a prospective study.

BACKGROUND: Pterygium is one of the most common conjunctival diseases among ophthalmic pathologies. The frequency of recurrences is high, either after surgical treatment or after treatment combined with mitomycin C or beta-radiation therapy. AIMS: The purpose of this study was to determine whether concanavalin A (ConA) lectin bound to the pterygial surface can be used to detect recurrence or remnants of pterygium after surgical excision. MATERIALS AND METHODS: This was a prospective study on 20 patients with pterygium, divided in five stages, pre-surgery, early post-surgery (24h), late post-surgery (seven days), very late post-surgery (four weeks) and two months after the procedure. A drop of fluorescein-marked Con A (35 microg/mL) was instilled in the lower conjunctival eyelid sac and the eye was exposed to the light of a Wood's lamp for an average of five seconds. RESULTS: Out of the 20 patients, eight patients were found to have fluorescent stretch marks over the scar corresponding to residual pterygial tissue at four weeks; two months after the procedure of re-surgery we observed no fluorescent remnants. All residual pterygia were confirmed through histochemistry studies. CONCLUSION: It was possible to detect remnants of pterygium in postoperative patients and recurrences in early pre-clinical stages through the visualization of fluorescent ConA bound to the pterygial surface.

Interaction of the protein C activation peptide with platelets.

The peptide NH(2)-DTEDQEDQVDPR-COOH is released during activation of protein C zymogen. We measured the effect of a synthetic peptide with an amino acid sequence similar to that of the natural peptide on platelets from healthy individuals using platelet aggregometry. We found that this synthetic peptide inhibits platelet aggregation induced by thrombin; furthermore, it diminishes mobilization of intraplatelet calcium. Molecular docking showed weak interaction between the synthetic peptide and thrombin. Our findings suggest that this synthetic peptide may interact with a receptor located on the platelet cell membrane.