Understanding the complexity of Tityus serrulatus venom: A focus on high molecular weight components

Tityus serrulatus scorpion is responsible for a significant number of envenomings in Brazil, ranging from mild to severe, and in some cases, leading to fatalities. While supportive care is the primary treatment modality, moderate and severe cases require antivenom administration despite potential limitations and adverse effects. The remarkable proliferation of T. serrulatus scorpions, attributed to their biology and asexual reproduction, contributes to a high incidence of envenomation. T. serrulatus scorpion venom predominantly consists of short proteins acting as neurotoxins (α and ß), that primarily target ion channels. Nevertheless, high molecular weight compounds, including metalloproteases, serine proteases, phospholipases, and hyaluronidases, are also present in the venom. These compounds play a crucial role in envenomation, influencing the severity of symptoms and the spread of venom. This review endeavors to comprehensively understand the T. serrulatus scorpion venom by elucidating the primary high molecular weight compounds and exploring their potential contributions to envenomation. Understanding these compounds' mechanisms of action can aid in developing more effective treatments and prevention strategies, ultimately mitigating the impact of scorpion envenomation on public health in Brazil.

Intradermal hyaluronidase: The answer to treatment in softening a fibrous thick supratip skin in rhinoplasty?

@# It is a common goal for rhinoplastic surgeons to make the best-looking tip with proper projection, maintaining the tip lobule appearance with a supratip break. (Figure 1) However, a fibrous thick skin with fullness may not achieve the ideal tip. It is one of the nuisances in rhinoplasty that makes tip definition surgery difficult. The supratip area remains firm and convex causing a wide bulbous feature of the tip. Several techniques have been introduced with good results however some may still result in supratip fullness because of the firm fibrous nature of thick skin.1-3 Hyaluronidase is an enzyme that depolymerizes hyaluronic acid which is present in the epithelium.4 The use of intradermal hyaluronidase for thick skin was discovered by the junior author (JMP) in one of his rhinoplasties when he injected hyaluronidase in a nose with fillers containing hyaluronic acid. The fillers not only instantly dissolved but the skin also softened, so he tried injecting intradermally in his subsequent rhinoplasties on non-filler noses with fibrous thick skin and indeed found the same effect of softening of the fibrous supratip skin. We here describe the technique used in this preliminary clinical series.

Comparative venomic profiles of three spiders of the genus Phoneutria

Spider venoms induce different physio-pharmacological effects by binding with high affinity on molecular targets, therefore being of biotechnological interest. Some of these toxins, acting on different types of ion channels, have been identified in the venom of spiders of the genus Phoneutria, mainly from P. nigriventer. In spite of the pharmaceutical potential demonstrated by P. nigriventer toxins, there is limited information on molecules from venoms of the same genus, as their toxins remain poorly characterized. Understanding this diversity and clarifying the differences in the mechanisms of action of spider toxins is of great importance for establishing their true biotechnological potential. This prompted us to compare three different venoms of the Phoneutria genus: P. nigriventer (Pn-V), P. eickstedtae (Pe-V) and P. pertyi (Pp-V). Methods: Biochemical and functional comparison of the venoms were carried out by SDS-PAGE, HPLC, mass spectrometry, enzymatic activities and electrophysiological assays (whole-cell patch clamp). Results: The employed approach revealed that all three venoms had an overall similarity in their components, with only minor differences. The presence of a high number of similar proteins was evident, particularly toxins in the mass range of ~6.0 kDa. Hyaluronidase and proteolytic activities were detected in all venoms, in addition to isoforms of the toxins Tx1 and Tx2-6. All Tx1 isoforms blocked Nav1.6 ion currents, with slight differences. Conclusion: Our findings showed that Pn-V, Pe-V and Pp-V are highly similar concerning protein composition and enzymatic activities, containing isoforms of the same toxins sharing high sequence homology, with minor modifications. However, these structural and functional variations are very important for venom diversity. In addition, our findings will contribute to the comprehension of the molecular diversity of the venoms of the other species from Phoneutria genus, exposing their biotechnological potential as a source for searching for new active molecules.(AU)

Effects of photobiomodulation (660 nm laser) on anthracycline extravasation: An experimental study / Efeitos da fotobiomodulação (laser 660 nm) no extravasamento de antraciclina: estudo experimental / Efectos de la fotobiomodulación (láser de 660 nm) sobre la extravasación de antraciclinas: estudio experimental

Abstract Objective: to investigate the effect of using different agents (topical hyaluronidase, photobiomodulation, and the association of photobiomodulation with topical hyaluronidase) in preventing the formation of lesions caused by doxorubicin extravasation, as well as in the reduction of lesions formed by extravasation of this drug. Method: a quasi-experimental study conducted with 60 Wistar rats, randomized into four groups with 15 animals each. Group 1 (Control); Group 2 (Hyaluronidase); Group 3 (Photobiomodulation); and Group 4 (Hyaluronidase + Photobiomodulation). A wound was induced by applying 1 mg of doxorubicin to the subcutaneous tissue of the back of the animals. The concentration of topical hyaluronidase was 65 turbidity units/g and the energy employed was 1 joule of 100 mW red laser per square centimeter. With macroscopic evaluation every two days for 28 days, the following variables were observed: skin integrity, presence of blisters, hyperemia, exudate, bleeding, edema, crust, peeling and granulation tissue. Results: the animals from the groups subjected to photobiomodulation obtained better results in the assessment of the following variables: bleeding, hyperemia, exudate, intact skin and edema. Conclusion: it was evidenced that the association of photobiomodulation with topical hyaluronidase was effective in reducing the local effects and assisted in the wound healing process, and that PBM alone was able to prevent appearance of lesions.

Resumo Objetivo: investigar o efeito do uso de diferentes agentes (hialuronidase tópica, fotobiomodulação e da associação da fotobiomodulação com a hialuronidase tópica) na prevenção de formação de lesões causadas por extravasamento de doxorrubicina bem como na diminuição de lesões formadas pelo extravasamento desta droga. Método: estudo experimental com 60 ratos Wistar, randomizados em quatro grupos de 15 animais. Grupo 1 (Controle); Grupo 2 (Hialuronidase); Grupo 3 (Fotobiomodulação) e Grupo 4 (Hialuronidase + Fotobiomodulação). Induziu-se ferida aplicando 1 mg de doxorrubicina no subcutâneo do dorso dos animais. A concentração da hialuronidase tópica foi de 65 unidades de turbidez/g, a energia empregada foi de 1 joule de laser vermelho 100 mW por centímetro quadrado. Com avaliação macroscópica a cada dois dias por 28 dias, observou-se as variáveis: integridade da pele, presença de flictema, hiperemia, exsudato, sangramento, edema, crosta, descamação e tecido de granulação. Resultados: os animais dos grupos com fotobiomodulação obtiveram melhores resultados na avaliação das variáveis: sangramento, hiperemia, exsudato, pele íntegra e edema. Conclusão: evidenciou-se que a associação da fotobiomodulação com a hialuronidase tópica foi eficaz na diminuição dos efeitos locais e auxiliou no processo de cicatrização da ferida e que a FBM isolada foi capaz de prevenir o aparecimento de lesões.

Resumen Objetivo: investigar el efecto del uso de diferentes agentes (hialuronidasa tópica, fotobiomodulación y la combinación de fotobiomodulación y hialuronidasa tópica) en la prevención de la formación de lesiones causadas por la extravasación de doxorrubicina y en la reducción de las lesiones formadas por la extravasación de ese fármaco. Método: estudio experimental con 60 ratas Wistar, distribuidos aleatoriamente en cuatro grupos de 15 animales. Grupo 1 (Control); Grupo 2 (Hialuronidasa); Grupo 3 (Fotobiomodulación) y Grupo 4 (Hialuronidasa + Fotobiomodulación). La herida se indujo aplicando 1 mg de doxorrubicina por vía subcutánea en el lomo de los animales. La concentración de hialuronidasa tópica fue de 65 unidades de turbidez/g, la energía utilizada fue de 1 joule de láser rojo de 100 mW por centímetro cuadrado. En la evaluación macroscópica cada dos días durante 28 días se observaron las siguientes variables: piel intacta, presencia de flictena, hiperemia, exudado, sangrado, edema, costra, descamación y tejido de granulación. Resultados: los animales de los grupos con fotobiomodulación obtuvieron mejores resultados en la evaluación de las variables: sangrado, hiperemia, exudado, piel intacta y edema. Conclusión: se demostró que la combinación de fotobiomodulación y hialuronidasa tópica fue eficaz para reducir los efectos locales y ayudó en el proceso de cicatrización de heridas y que la FBM por sí sola previno la aparición de lesiones.

Detection of DNA methylation of HYAL2 gene for differentiating malignant from benign thyroid tumors / 南方医科大学学报

OBJECTIVE@#To assess the value of DNA methylation level of HYAL2 gene as a molecular marker for differential diagnosis of malignant and benign thyroid tumors.@*METHODS@#DNA methylation of HYAL2 gene in tissue specimens of 190 patients with papillary thyroid cancer (PTC) and 190 age- and gender-matched patients with benign thyroid tumors was examined by mass spectrometry, and the protein expression of HYAL2 was detected immunohistochemically for another 55 pairs of patients. Logistic regression analysis was performed to calculate the odds ratio (OR) and evaluate the correlation of per 10% reduction in DNA methylation with PTC. Receiver operating characteristic (ROC) curve analysis was performed and the area under curve (AUC) was calculated to assess the predictive value of alterations in HYAL2 methylation.@*RESULTS@#Hypomethylation of HYAL2_CpG_3 was significantly correlated with early-stage PTC (OR=1.51, P=0.001), even in stage I cancer (OR=1.42, P=0.007). Age-stratified analysis revealed a significantly stronger correlation between increased HYAL2_CpG_ 3 methylation and early-stage PTC in patients below 50 years than in those older than 50 years (OR: 1.89 vs 1.37, P < 0.05); ROC analysis also showed a larger AUC of 0.787 in younger patients. The results of immunohistochemistry showed that patients with PTC had significantly higher protein expressions of HYAL2 than patients with benign tumors.@*CONCLUSION@#The alterations of DNA methylation level of HYAL2 gene is significantly correlated with early-stage PTC, suggesting the value of DNA methylation level as a potential biomarker for differentiation of malignant from benign thyroid tumors.

Uso terapéutico de la hialuronidasa y la triamcinolona en el pseudotumor orbitario / Therapeutic use of hyaluronidase and triamcinolone in orbital pseudotumor

RESUMEN El origen del pseudotumor orbitario no es del todo conocido. Se admite su naturaleza inflamatoria granulomatosa e inespecífica en diferentes localizaciones. El pseudotumor orbitario se define como una respuesta inflamatoria celular pleomórfica, que está usualmente confinado a estructuras de la órbita y tiene una evolución limitada. En este trabajo se presenta una paciente femenina de 16 años, con diagnóstico de pseudotumor orbitario corroborado por biopsia y tomografía axial computarizada, refractaria al tratamiento con esteroides sistémicos, por lo que se decide iniciar con la aplicación de hialuronidasa y triamcinolona en el espacio peribulbar. Los casos agudos casi siempre responden rápidamente al tratamiento con cortocoesteroides, como prednisona, pero debemos tener en cuenta que existen pacientes que son refractarios al tratamiento, por lo que es necesario buscar procedimientos alternativos. Una opción es el uso de hialuronidasa para destruir las uniones extracelulares, y difundir un esteroide de manera local, como la triamcinolona, más efectiva dentro del tejido inflamatorio para provocar un efecto localizado de este. A los tres meses del tratamiento hubo una regresión total del cuadro en esta paciente(AU)

ABSTRACT The exact etiology of orbital pseudotumor is unknown, but its granulomatous unspecific inflammatory nature at various locations has been recognized. Orbital pseudotumor is defined as a cellular pleomorphic inflammatory response of limited evolution often confined to orbital structures. A case is presented of a female 16-year-old patient diagnosed with orbital pseudotumor confirmed by biopsy and computerized axial tomography, refractory to treatment with systemic steroids, due to which it is decided to start treatment with hyaluronidase and triamcinolone in the peribulbar space. Acute cases often respond fast to treatment with corticosteroids such as prednisone. It should be borne in mind that there are patients who are refractory to treatment for whom alternative treatments should be sought. An option is the use of hyaluronidase to destroy extracellular junctions and locally spread a steroid such as triamcinolone, most effectively within the inflammatory tissue to ensure its localized effect. Total regression of the patient's status was observed at three months of treatment(AU)

Uso de enzimas como tratamiento dermatológico regenerador de las líneas de expresión / Use of enzymes as a regenerative dermatological treatment of expression lines / Uso de enzimas como tratamento dermatológico regenerativo de linhas de expressão

INTRODUCCIÓN: el tratamiento con enzimas es una alternativa estética mínimamente invasiva para mejorar la apariencia facial y disminuir las líneas de expresión. OBJETIVO: Determinar el uso de las enzimas hialuronidasa, colagenasa y lipasa como tratamiento enzimático dermatológico para las líneas de expresión facial. MATERIALES Y MÉTODO: estudio de campo, prospectivo, población 457 pacientes que acudieron a la consulta dermatológica entre los años 2013 y 2018 para tratamiento con enzimas. El instrumento de recolección de datos fue la hoja de registro y la fuente documental las historias clínicas. El método estadístico fue descriptivo, la información se presenta en tablas y gráficos. RESULTADOS: la edad promedio de los pacientes fue de 45,2 ± 10,1 años, 40,9% recibió 2 kits de enzimas con los 3 componentes básicos de colagenasa, hialuronidasas y lipasas. Se encontró diferencia significativa en la relación de atención entre hombres y mujeres, de 1:14, es decir las mujeres acudieron más a la consulta solicitando la colocación de este tratamiento. CONCLUSIÓN: el tratamiento con enzimas aporta beneficios al incrementar la permeabilidad dérmica, aumenta el flujo sanguíneo y el drenaje linfático, disminuye los tabiques fibrosos de la celulitis, la flacidez, adiposidades, y rejuvenece el aspecto general. Por lo que se plantea como un tratamiento efectivo para disminuir las líneas de expresión.

INTRODUCTION: enzyme treatment represents a minimally invasive aesthetic alternative to improve facial appearance and decrease expression lines. OBJECTIVE: to determine the use of the enzymes hyaluronidase, collagenase and lipase as a dermatological enzyme treatment as a regenerative treatment for expression lines. METHODS: a prospective field study was conducted of a population made up of 457 patients who attended the UNIMEL dermatological consultation between 2013 and 2018 to be treated with enzymes. The data collection instrument was the record sheet and the documentary source was the medical records. The statistical method was descriptive, the information is presented in tables and graphs. RESULTS: the average age was 45.2 ± 10.1 years of age, to whom a majority of 40.9% were applied 2 kits of enzymes with the 3 basic components of collagenase, hyaluronidases and lipases to act synergistically each other enhancing functions and revitalizing the cells of the face. A significant difference was found in the care ratio between men and women, 1:14, that is, the women attended the consultation more requesting the placement of this treatment. CONCLUSION: the use of enzymes provides great benefits to increase skin permeability, increases lymphatic drainage, reduces fibrous septa of cellulite, sagging and fat, increases blood flow and rejuvenates the general appearance. So, it is proposed as an effective treatment to reduce expression lines

INTRODUÇÃO: o tratamento enzimático é uma alternativa estética minimamente invasiva para melhorar a aparência facial e diminuir as linhas de expressão. OBJETIVO: determinar o uso das enzimas hialuronidase, colagenase e lipase como tratamento enzimático dermatológico para linhas de expressão facial. MATERIAIS E MÉTODOS: estudo de campo em perspectiva, população de 457 pacientes que compareceram à consulta dermatológica entre 2013 e 2018 para tratamento enzimático. O instrumento de coleta de dados foi a folha de registros e a fonte documental foram os registros médicos. O método estatístico foi descritivo, as informações são apresentadas em tabelas e gráficos. RESULTADOS: a idade média dos pacientes foi de 45,2 ± 10,1 anos, 40,9% receberam 2 kits de enzimas com os 3 componentes básicos de colagenase, hialuronidases e lipases. Foi encontrada diferença significativa de 1:14 na relação de atenção entre homens e mulheres, ou seja, as mulheres compareceram mais frequentemente as consultas para a colocação desse tratamento do que aos homes. CONCLUSÃO: o tratamento enzimático oferece benefícios ao aumentar a permeabilidade dérmica, aumenta o fluxo sanguíneo e a drenagem linfática, reduz os septos fibrosos da celulite, flacidez, adiposidade e rejuvenesce a aparência geral. Por isso, é proposto como um tratamento eficaz para diminuir as linhas de expressão.

Randomized, Double-Blind, Placebo-Controlled Trial on the Efficacy of Hyaluronidase in Preventing Perineal Trauma in Nulliparous Women

Rol de los estreptococos del grupo Streptococcus anginosus en infecciones de piel y partes blandas: reporte de 3 casos / Role of streptococci of the Streptococcus anginosus group in skin and soft tissue infections: report of 3 cases

Los estreptococos del grupo Streptococcus anginosus (GSA) comprenden uno de los grupos que conforman el grupo viridans de estreptococos y son parte de la microbiota humana habitual de la cavidad oral, del tracto respiratorio superior y del tracto gastrointestinal. Pueden producir colecciones purulentas de órganos sólidos, abscesos cutáneos y celulitis, entre otras infecciones. Este grupo posee numerosos factores de virulencia como hialuronidasa, adhesinas, enzimas extracelulares y hemolisinas. El objetivo de este trabajo fue presentar tres casos clínicos de infecciones asociadas a traumatismos vinculados con la cavidad oral como golpes de puño o mordeduras humanas. En todos ellos, los GSA desarrollaron como germen único, lo que pone de manifiesto su importante rol en este tipo de infecciones.

The Streptococcus anginosus group (SAG) is part of the viridans group streptococci and they belong to the normal human microbiota of the oral cavity, the upper respiratory tract and the gastrointestinal tract. It can produce purulent collections of solid organs, cutaneous abscesses and cellulitis, among other infections. This group has numerous virulence factors such as hyaluronidase, adhesins, extracellular enzymes and hemolysins. The objective of this work is to present three clinical cases of infections associated with trauma related to the oral cavity such as fist bumps or human bites. In all of them, the SAG developed as a single organism, which highlights its important role in this type of infection.

Os estreptococos do grupo do Streptococcus anginosus (GSA) são parte de um dos grupos que constituem o grupo viridans e fazem parte da microbiota normal da cavidade oral, do trato respiratório superior e do trato gastrointestinal dos seres humanos. Podem produzir coleções purulentas de órgãos sólidos, abscessos cutâneos e celulite, entre outras infecções. Este grupo possui numerosos fatores de virulência, como hialuronidases, adesinas, enzimas extracelulares e hemolisinas. O objetivo desse trabalho foi apresentar três casos clínicos de infecções associadas a traumatismos associados à cavidade oral, decorrentes de socos ou mordidas humanas. Em todos eles os GSA se desenvolveram como germe único, o que põe de manifesto seu papel importante nesse tipo de infecções.

Transient retinal artery occlusion after phacoemulsification under local anesthetic block / Oclusão arterial retiniana transitória após facoemulsificação sob bloqueio anestésico

ABSTRACT We here in report the case of a patient subjected to cataract surgery through phacoemulsification under local anesthetic block, without intra-operative complications. The patient presented important visual impairment in the first post-operative day. Fundoscopy showed pallor resembling cherry-red spots at the macula. Fluorescein angiography did not depict signs of vascular occlusion and the spectral-domain optical coherence tomography showed increased reflectivity in the inner layers of the retina, thus suggesting local thickening and edema. The current case led to the diagnostic hypothesis of transient retinal arterial occlusion.

RESUMO Relatamos um caso de um paciente submetido a facectomia por facoemulsificação sob bloqueio anestésico peribulbar, sem intercorrências per-operatória, que apresentou no primeiro dia de pós-operatório baixa visual significativa. À fundoscopia observou-se palidez em aspecto de mácula em cereja. A angiofluoresceinografia não demonstrou sinais de oclusão vascular e a tomografia de coerência óptica mostrou aumento da refletividade das camadas internas da retina, sugerindo espessamento e edema local. No caso descrito foi aventada hipótese diagnóstica de oclusão arterial retiniana transitória.

Heterologous expression and mutagenesis of recombinant Vespa affinis hyaluronidase protein (rVesA2)

Background:Crude venom of the banded tiger waspVespa affinis contains a variety of enzymes including hyaluronidases, commonly known as spreading factors.Methods:The cDNA cloning, sequence analysis and structural modelling of V. affinis venom hyaluronidase (VesA2) were herein described. Moreover, heterologous expression and mutagenesis of rVesA2 were performed.Results:V. affinis venom hyaluronidase full sequence is composed of 331 amino acids, with four predicted N-glycosylation sites. It was classified into the glycoside hydrolase family 56. The homology modelling exhibited a central core (α/β)7 composed of Asp107 and Glu109, acting as the catalytic residues. The recombinant protein was successfully expressed in E. coli with hyaluronidase activity. A recombinant mutant type with the double point mutation, Asp107Asn and Glu109Gln, completely lost this activity. The hyaluronidase from crude venom exhibited activity from pH 2 to 7. The recombinant wild type showed its maximal activity at pH 2 but decreased rapidly to nearly zero at pH 3 and was completely lost at pH 4.Conclusion:The recombinant wild-type protein showed its maximal activity at pH 2, more acidic pH than that found in the crude venom. The glycosylation was predicted to be responsible for the pH optimum and thermal stability of the enzymes activity.

Complex Regional Pain Syndrome of the Upper Limbs Caused by Facial Pain / 계명의대학술지

Polyacrylamide hydrogel is a widely used filler material in cosmetic procedures performed on the face and breasts. Recently, however, complications including inflammation, deformity, and pain have been reported. The present article addresses unregulated materials/products injected as dermal fillers. The authors report a case involving a 29-year-old woman who developed severe facial pain after undergoing a cosmetic procedure with injectable triamcinolone and hyaluronidase. Two months later, the pain spread to her upper and lower limbs, and abdomen, which eventually led to the the development and diagnosis of complex regional pain syndrome (CRPS) in the upper limbs. The authors hypothesize that CRPS in the upper limbs was responsible for the facial pain through sensitization of third-order neurons and the trigeminal nucleus caudalis extending to the upper cervical segments.

Treatment of post-traumatic hematoma and fibrosis using hyaluronidase injection

Patients complaining of swelling and hematoma caused by contusion of the face can be easily seen in the emergency room. Most of the treatments were conservative treatments such as ice bag application, mild compression dressing, and massage. During the follow-up, fibrosis progression due to hematoma was frequently observed in the contusion site. When hematoma or fibrosis is confirmed, hyaluronidase (H-lase) 1,500 IU and 2 mL of normal saline were mixed and subcutaneously injected in crisscross manner. To evaluate the improvement of hematoma before and after hyaluronidase injection, three plastic surgeons evaluated using the Vancouver scar scale and compared preoperative and postoperative images. Hematoma and fibrosis after facial trauma improved after hyaluronidase injection for early treatment.

Primary cultures of mouse small intestinal epithelial cells using the dissociating enzyme type I collagenase and hyaluronidase

The epithelium is a highly dynamic system, which plays a crucial role in the homeostasis of the intestinal tract. However, studies on the physiological and pathophysiological functions of intestinal epithelial cells (IECs) have been hampered due to lack of normal epithelial cell models. In the present study, we established a reproducible method for primary culture of mouse IECs, which were isolated from the viable small intestinal crypts of murine fetuses (on embryonic day 19), using type I collagenase and hyaluronidase in a short span of time (≤20 min). With this method, continuously growing mouse IECs, which can be subcultured over a number of passages, were obtained. The obtained cell lines formed a tight cobblestone-like arrangement, displayed long and slender microvilli, expressed characteristic markers (cytokeratin 18 and Notch-1), and generated increasing transepithelial electrical resistance and low paracellular permeability during in vitro culture. The cells also had enzymatic activities of alkaline phosphatase and sucrase-isomaltase, and secreted various cytokines (IL-1β, IL-6, IL-8, and monocyte chemoattractant protein-1), responding to the stimulation of Escherichia coli. These results show that the primary-cultured mouse IECs obtained by the method established here had the morphological and immunological characteristics of IECs. This culture system can be a beneficial in vitro model for studies on mucosal immunology and toxicology.

Antioxidant and Antiaging Assays of Hibiscus sabdariffa Extract and Its Compounds

Skin aging is a complex biological process due to intrinsic and extrinsic factors. Free radical oxidative is one of extrinsic factors that induce activation of collagenase, elastase and hyaluronidase. Natural product from plants has been used as antioxidant and antiaging. This study aimed to evaluate antioxidant and antiaging properties of Hibiscus sabdariffa extract (HSE) and its compounds including myricetin, ascorbic acid, and β carotene. The phytochemical of H. sabdariffa was determined using modified Farnsworth method and presence of phenols, flavonoids and tannins were in moderate content, whereas triterpenoids and alkaloids were in low content. Total phenolic content performed using Folin-Ciocalteu method, was 23.85 µg GAE/mg. Quantitative analysis of myricetin, β-carotene, and ascorbic acid of HSE was performed with Ultra-High Performance Liquid Chromatography (UHPLC) that shows 78.23 µg/mg myricetin, 0.034 µg/mg β-carotene, whilst ascorbic acid was not detected. HSE has lower activity on DPPH (IC₅₀ = 195.73 µg/mL) compared to β-carotene, the lowest in ABTS assay (IC50 = 74.58 µg/mL) and low activity in FRAP assay (46.24 µM Fe(II)/µg) compared to myricetin, β-carotene. Antiaging was measured through inhibitory activity of collagenase, elastase, and hyaluronidase. HSE had weakest collagenase inhibitory activity (IC₅₀= 750.33 µg/mL), elastase inhibitory activity (103.83 µg/mL), hyaluronidase inhibitory activity (IC₅₀ = 619.43 µg/mL) compared to myricetin, β-carotene, and ascorbic acid. HSE contain higher myricetin compared to β-carotene. HSE has moderate antioxidants and lowest antiaging activities. Myricetin is the most active both antioxidant and antiaging activities.

Cloning, structural modelling and characterization of VesT2s, a wasp venom hyaluronidase (HAase) from Vespa tropica

Wasp venom is a complex mixture containing proteins, enzymes and small molecules, including some of the most dangerous allergens. The greater banded wasp (Vespa tropica) is well-known for its lethal venom, whose one of the major components is a hyaluronidase (HAase). It is believed that the high protein proportion and activity of this enzyme is responsible for the venom potency. Methods: In the present study, cDNA cloning, sequencing and 3D-structure of Vespa tropica venom HAase were described. Anti-native HAase antibody was used for neutralization assay. Results: Two isoforms, VesT2a and VesT2b, were classified as members of the glycosidase hydrolase 56 family with high similarity (4297 %) to the allergen venom HAase. VesT2a gene contained 1486 nucleotide residues encoding 357 amino acids whereas the VesT2b isoform consisted of 1411 residues encoding 356 amino acids. The mature VesT2a and VesT2b are similar in mass and pI after prediction. They are 39119.73 Da/pI 8.91 and 39571.5 Da/pI 9.38, respectively. Two catalytic residues in VesT2a, Asp107 and Glu109 were substituted in VesT2b by Asn, thus impeding enzymatic activity. The 3D-structure of the VesT2s isoform consisted of a central core (/)7 barrel and two disulfide bridges. The five putative glycosylation sites (Asn79, Asn99, Asn127, Asn187 and Asn325) of VesT2a and the three glycosylation sites (Asn1, Asn66 and Asn81) in VesT2b were predicted. An allergenic property significantly depends on the number of putative N-glycosylation sites. The anti-native HAase serum specifically recognized to venom HAase was able to neutralize toxicity of V. tropica venom. The ratio of venom antiserum was 1:12. Conclusions: The wasp venom allergy is known to cause life-threatening and fatal IgE-mediated anaphylactic reactions in allergic individuals. Structural analysis was a helpful tool for prediction of allergenic properties including their cross reactivity among the vespid HAase.

Cloning, structural modelling and characterization of VesT2s, a wasp venom hyaluronidase (HAase) from Vespa tropica

Background: Wasp venom is a complex mixture containing proteins, enzymes and small molecules, including some of the most dangerous allergens. The greater banded wasp (Vespa tropica) is well-known for its lethal venom, whose one of the major components is a hyaluronidase (HAase). It is believed that the high protein proportion and activity of this enzyme is responsible for the venom potency. Methods: In the present study, cDNA cloning, sequencing and 3D-structure of Vespa tropica venom HAase were described. Anti-native HAase antibody was used for neutralization assay. Results: Two isoforms, VesT2a and VesT2b, were classified as members of the glycosidase hydrolase 56 family with high similarity (42-97 %) to the allergen venom HAase. VesT2a gene contained 1486 nucleotide residues encoding 357 amino acids whereas the VesT2b isoform consisted of 1411 residues encoding 356 amino acids. The mature VesT2a and VesT2b are similar in mass and pI after prediction. They are 39119.73 Da/pI 8.91 and 39571.5 Da/pI 9.38, respectively. Two catalytic residues in VesT2a, Asp107 and Glu109 were substituted in VesT2b by Asn, thus impeding enzymatic activity. The 3D-structure of the VesT2s isoform consisted of a central core (α/β)7 barrel and two disulfide bridges. The five putative glycosylation sites (Asn79, Asn99, Asn127, Asn187 and Asn325) of VesT2a and the three glycosylation sites (Asn1, Asn66 and Asn81) in VesT2b were predicted. An allergenic property significantly depends on the number of putative N-glycosylation sites. The anti-native HAase serum specifically recognized to venom HAase was able to neutralize toxicity of V. tropica venom. The ratio of venom antiserum was 1:12. Conclusions: The wasp venom allergy is known to cause life-threatening and fatal IgE-mediated anaphylactic reactions in allergic individuals. Structural analysis was a helpful tool for prediction of allergenic properties including their cross reactivity among the vespid HAase.(AU)

Hyaluronidases and hyaluronan synthases expression is inversely correlated with malignancy in lung/bronchial pre-neoplastic and neoplastic lesions, affecting prognosis

We collected a series of 136 lung/bronchial and 56 matched lung parenchyma tissue samples from patients who underwent lung/bronchial biopsies and presented invasive carcinoma after lung surgery. The lung/bronchial samples included basal cell hyperplasia, squamous metaplasia, moderate dysplasia, adenomatous hyperplasia, severe dysplasia, squamous cell carcinoma and adenocarcinoma. Matched lung parenchyma tissue samples included 25 squamous cell carcinomas and 31 adenocarcinomas. Immunohistochemistry was performed to analyze for the distribution of hyaluronidase (Hyal)-1 and −3, and hyaluronan synthases (HAS)-1, −2, and −3. Hyal-1 showed significantly higher expression in basal cell hyperplasia than in moderate dysplasia (P=0.01), atypical adenomatous hyperplasia (P=0.0001), or severe dysplasia (P=0.03). Lower expression of Hyal-3 was found in atypical adenomatous hyperplasia than in basal cell hyperplasia (P=0.01) or moderate dysplasia (P=0.02). HAS-2 was significantly higher in severe dysplasia (P=0.002) and in squamous metaplasia (P=0.04) compared with basal cell hyperplasia. HAS-3 was significantly expressed in basal cell hyperplasia compared with atypical adenomatous hyperplasia (P=0.05) and severe dysplasia (P=0.02). Lower expression of HAS-3 was found in severe dysplasia compared with squamous metaplasia (P=0.01) and moderate dysplasia (P=0.01). Epithelial Hyal-1 and −3 and HAS-1, −2, and −3 expressions were significantly higher in pre-neoplastic lesions than in neoplastic lesions. Comparative Cox multivariate analysis controlled by N stage and histologic tumor type showed that patients with high HAS-3 expression in pre-neoplastic cells obtained by lung/bronchial biopsy presented a significantly higher risk of death (HR=1.19; P=0.04). We concluded that localization of Hyal and HAS in lung/bronchial pre-neoplastic and neoplastic lesions was inversely related to malignancy, which implied that visualizing these factors could be a useful diagnostic procedure for suspected lung cancer. Finalizing this conclusion will require a wider study in a randomized and prospective trial.

Arthropod venom Hyaluronidases: biochemical properties and potential applications in medicine and biotechnology

Hyaluronidases are enzymes that mainly degrade hyaluronan, the major glycosaminoglycan of the interstitial matrix. They are involved in several pathological and physiological activities including fertilization, wound healing, embryogenesis, angiogenesis, diffusion of toxins and drugs, metastasis, pneumonia, sepsis, bacteremia, meningitis, inflammation and allergy, among others. Hyaluronidases are widely distributed in nature and the enzymes from mammalian spermatozoa, lysosomes and animal venoms belong to the subclass EC 3.2.1.35. To date, only five three-dimensional structures for arthropod venom hyaluronidases (Apis mellifera and Vespula vulgaris) were determined. Additionally, there are four molecular models for hyaluronidases fromMesobuthus martensii, Polybia paulista and Tityus serrulatus venoms. These enzymes are employed as adjuvants to increase the absorption and dispersion of other drugs and have been used in various off-label clinical conditions to reduce tissue edema. Moreover, a PEGylated form of a recombinant human hyaluronidase is currently under clinical trials for the treatment of metastatic pancreatic cancer. This review focuses on the arthropod venom hyaluronidases and provides an overview of their biochemical properties, role in the envenoming, structure/activity relationship, and potential medical and biotechnological applications.

Influence of anestrus and diestrus stages of oocyte on meiotic competence in bitches / Influência das fases de anestro e diestro na competência meiótica oocitária de cadelas

The bitch has reproductive peculiarities that differentiate it from other species. Several experiments have been conducted to establish efficient protocols for maturation; however, the results appear to be unsatisfactory. In this respect, the reproductive female donor should be considered, since it can be a factor of variability of findings in this species. The objective of this study was to evaluate the relationship between the estrous cycle phases phase diestrus and anestrus on canine oocyte in vitro maturation (IVM). The ovaries were transported in sodium chloride 0.9% solution, and were cut into phosphate-buffered saline (PBS) and the cumulus-oocyte complexes (COCs) selected in tissue culture medium (TCM), supplemented with 199 HEPES. A total of 469 grade 1 oocytes were collected from bitches in anestrus and diestrus. These selected oocytes were transferred to the maturation medium for a period of 72 hours, then subjected to hyaluronidase solution and stained with Hoechst 33342 to assess nuclear configuration. The comparison of anestrus and diestrus phase showed no differences (p > 0.05) between the nuclear maturation stages. Thus, the phase of the estrous cycle did not influence the in vitro maturation of canine oocytes, increasing the rates of M-II in this species.

A cadela apresenta particularidades reprodutivas que a diferencia de outras espécies. Diversos experimentos têm sido realizados visando estabelecer protocolos eficientes para a maturação, entretanto os resultados mostram-se insatisfatórios. Nesse aspecto, a fase reprodutiva da fêmea doadora deve ser considerada, já que pode ser um fator de variabilidade dos achados ate então presenciados nessa espécie. O objetivo deste estudo foi avaliar a influencia das fases do ciclo estral (anestro e diestro) na maturação in vitro (MIV) de cadelas. Os ovários foram transportados em solução de cloreto de sódio 0,9% e seccionados em solução salina fosfato tamponado (PBS) e os complexos cumulus-oócito (COCs) selecionados em meio de cultura de tecidos (TCM) 199 suplementado com HEPES. Foram obtidos 469 oócitos grau 1 de cadelas em anestro e diestro. Esses oócitos selecionados foram transferidos para o meio de maturação por um período de 72 horas, sendo posteriormente submetidos a solução de hialuronidase e corados com HOESCHT 33342 para avaliação da configuração nuclear. A comparação das fases de anestro e diestro não revelou diferença (p > 0,05) entre os estágios de maturação nuclear. Dessa maneira, a fase do ciclo estral não influenciou na maturação in vitro de oócitos caninos, incrementando os índices de M-II nesta espécie.