Structural characterization of DNA amplicons by ATR-FTIR spectroscopy as a guide for screening metainflammatory disorders in blood plasma.

Attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopy is a promising rapid, reagent-free, and low-cost technique considered for clinical translation. It allows to characterize biofluids proteome, lipidome, and metabolome at once. Metainflammatory disorders share a constellation of chronic systemic inflammation, oxidative stress, aberrant adipogenesis, and hypoxia, that significantly increased cardiovascular and cancer risk. As a result, these patients have elevated concentration of cfDNA in the bloodstream. Considering this, DNA amplicons were analyzed by ATR-FTIR at 3 concentrations with 1:100 dilution: (IU/mL): 718, 7.18, and 0.0718. The generated IR spectrum was used as a guide for variable selection. The main peaks in the biofingerprint (1800-900 cm-1) give important information about the base, base-sugar, phosphate, and sugar-phosphate transitions of DNA. To validate our method of selecting variables in blood plasma, 38 control subjects and 12 with metabolic syndrome were used. Using the wavenumbers of the peaks in the biofingerprint of the DNA amplicons, was generated a discriminant analysis model with Mahalanobis distance in blood plasma, and 100 % discrimination accuracy was obtained. In addition, the interval 1475-1188 cm-1 showed the greatest sensitivity to variation in the concentration of DNA amplicons, so curve fitting with Gaussian funcion was performed, obtaining adjusted-R2 of 0.993. PCA with Mahalanobis distance in the interval 1475-1188 cm-1 obtained an accuracy of 96 % and PLS-DA modeling in the interval 1475-1088 cm-1 obtained AUC = 0.991 with sensitivity of 95 % and specificity of 100 %. Therefore, ATR-FTIR spectroscopy with variable selection guided by DNA IR peaks is a promising and efficient method to be applied in metainflammatory disorders.

Are pediatricians familiar with hereditary angioedema?

Background: Hereditary angioedema (HAE) is an autosomal dominant disease characterized by recurrent episodes of subcutaneous or mucosal edema caused by excess bradykinin. The aim of the present study was to assess the knowledge of pediatricians about hereditary angioedema. Methods: An online survey with 12 HAE-related and 14 demographics-related questions was e-mailed to all pediatricians who were members of the Brazilian Society of Pediatrics (n = 17 145) once a week during the months of June and July 2021. The electronic questionnaire assessed clinical manifestations, diagnosis, and treatment of hereditary angioedema in children and adolescents. Results: Four hundred and fifty-five pediatricians responded to the questionnaire (2.6%), of whom 55 (12.1%) were board certified in Allergy and Immunology (A/I), while 400 (87.9%) were not (N-A/I). Three hundred and sixty-eight (80.9%) were female, 289 (55.7%) were under 50 years of age, 286 (62.9%) graduated from Medical School more than 10 years previously, 83 (18.2%) held an MSc/PhD degree, and 253 (55.6%) were living in the Southeast Region of Brazil. The median number of correct answers to the questions related to HAE among A/I was 7 out of 12 (58.3%), with median ranging from 4.5 to 8 correct answers, while for N-A/I it was 3 (25%), with median ranging from 2.5 to 4 correct answers (p < 0.001). Conclusion: Knowledge about HAE among Brazilian pediatricians, whether board certified in Allergy and Immunology or not, was unsatisfactory. HAE is a rare disease, largely unknown among physicians; therefore, increasing awareness may lead to improvement in diagnosis and treatment.

Amoxicillin degradation by iron photonanocatalyst synthetized by green route using pumpkin (Tetsukabuto) peel extract.

Amoxicillin is a pharmaceutical compound that is not degraded in wastewater treatment plants, causing harm to the environment. In this work, an iron nanoparticle (IPP) was synthesized using pumpkin (Tetsukabuto) peel extract for the degradation of amoxicillin under UV light. The IPP was characterized using scanning electron microscopy/energy dispersive x-ray spectroscopy, transmission electron microscopy, X-ray diffraction, Fourier-transform infrared spectroscopy, thermogravimetric analysis, and Raman spectroscopy techniques. The photocatalytic efficiency of IPP was analyzed by investigating the effect of IPP dosage (1-3 g L-1), initial amoxicillin concentration (10-40 mg L-1), pH (3-9), reaction time (10-60 min), and the effect of inorganic ions (1 g L-1). The optimum conditions for the maximum photodegradation removal (≈60%) were IPP = 2.5 g L-1, initial amoxicillin concentration = 10 mg L-1, pH = 5.6, and irradiation time = 60 min. The results of this study showed that inorganic ions (Mg2+, Zn2+, and Ca2+) negatively affect the photodegradation of amoxicillin by IPP; the quenching test showed that hydroxyl radical (OH•) is the primary reactive species of the reaction; NMR analysis revealed changes in amoxicillin molecules after photoreaction; the subproducts of photodegradation were identified by LC-MS; the proposed kinetic model demonstrated good applicability, predicting the behavior of OH• and determining the kinetic constant, and the cost analysis based on required energy (238.5 kWh m-3 order-1) indicated that the amoxicillin degradation method by IPP is economically viable. This study developed a new efficient iron nanocatalyst for the removal of antibiotics from aqueous environments and provided optimal conditions and relevant information in the area of advanced oxidative processes.

Effect of rACLF, a recombinant snake venom metallopeptidase on cell viability, chemokine expression and degradation of extracellular matrix proteins.

Snake venom metallopeptidases (SVMPs) comprise a family of zinc-dependent enzymes, which display many different biological activities. ACLF is a 23kDa fibrinolytic non-hemorrhagic metallopeptidase from the venom of the snake Agkistrodon contortrix laticinctus. We have previously developed an expression system for production of recombinant ACLF (rACLF) in bacteria. To achieve a better understanding of the role of such enzyme in envenoming cases, we have studied the biological properties of rACLF, including the ability of enzyme to degrade extracellular proteins, as well its cytotoxic effect in human fibroblasts and HeLa cells. Our results showed that rACLF hydrolyzed laminin, fibronectin, type IV collagen and thrombospondin. rACLF decreased HeLa cell viability, changed cell morphology and induced detachment, while for human fibroblasts no cytotoxic effects were observed after treatment with rACLF. In addition, growth-related oncogene (GRO) and monocyte chemoattractant protein 1 (MCP-1/CCL2) were chemokines detected in the culture supernatant of human fibroblasts incubated with rACLF for 48h. These chemokines could contribute to the severe local lesion induced by Agkistrodon contortrix lacticinctus venom. These findings suggest a relevant role for ACLF in envenomation.

Regulation of extracellular chitinases and proteases in the entomopathogen and acaricide Metarhizium anisopliae.

Metarhizium anisopliae infects insects and ticks via a combination of specialized structures and cuticle degradation. Hydrolytic enzymes are accepted as key factors for the penetration step. The search for pathogenicity determinants has demonstrated that the process is multifactorial. Host specificity is an important factor to be addressed. The study of the enzymes produced during infection is important to discover those with a role in the process. To address some of the enzymes that take part during the infection of the tick, Boophilus microplus, we have analyzed the secretion of proteases and chitinases in single and combined carbon/nitrogen sources as compared with such complex substrates as chitin and B. microplus cuticles. Two chitinases, endo- and N-acetylglucosaminidases, and two proteases, subtilisin and trypsin-like proteases, were analyzed. Enzyme activities were detected in all carbon sources tested, but higher levels were found when combinations of carbon sources were used. A major 30-kDa protein apparently secreted during M. anisopliae growth on all carbon/nitrogen sources tested was demonstrated by SDS-PAGE.