Characterization of expression patterns and dynamic relocation of Notch proteins during acrosome reaction of bull spermatozoa.

Notch is a conserved cell-signaling pathway involved in spermatogenesis regulation. This study firstly evaluated the presence, localization patterns, acquisition origin and relation to acrosome reaction of Notch proteins in bull sperm. Western Blot analysis detected all Notch proteins in ejaculated bull sperm, and immunostaining described their specific sperm localization. Recovery of sperm from different segments showed that Notch proteins have testicular origin (NOTCH1, NOTCH2, DLL4), are sequentially acquired during sperm maturation along epididymal transit (NOTCH3, DLL3, JAGGED1-2), or post-ejaculation (DLL1, NOTCH4). Testis NOTCH2 is ubiquitously expressed in all germ-cell lines, whereas DLL4 is expressed in round and elongated spermatids during the Golgi, Cap, Acrosome and Maturation phases. In vitro spontaneous and induced sperm acrosome reaction induce consistent sperm regional relocation of NOTCH2, DLL4 and JAGGED1, and these relocation patterns are significantly associated to sperm acrosome status. NOTCH2 and JAGGED1 are relocated from the head apical to the post-equatorial regions, whereas DLL4 is lost along with the acrosome, evidencing that sperm spatial redistribution of NOTCH2 and JAGGED1 is linked to acrosome reaction onset, whereas DLL4 loss is linked to AR completion. Overall, results prompt for a relevant Notch role in bull sperm acrosome testicular development, epididymal maturation and acrosome reaction.

Frequency of low-dose dexamethasone suppression test (LDDST) response patterns and their correlation with clinicopathologic signs in dogs suspected of having Cushing's syndrome: A retrospective study.

A retrospective cross-sectional study was conducted to assess the frequency of low-dose dexamethasone suppression test (LDDST) patterns in canine patients that had clinicopathologic signs consistent with Cushing's syndrome (CS). Medical records for patients of interest (N = 128) were reviewed between January 2014 and December 2020 to analyse and classify LDDST results based upon the following patterns: lack of suppression, partial suppression, complete suppression, escape, or inverse. Complete suppression, lack of suppression, partial suppression, escape, and inverse patterns were identified in 39.1%, 31.2%, 14.1%, 10.1% and 5.5% of cases respectively. LDDST results were also evaluated with respect to clinical signs, serum alkaline phosphatase (ALP) activity, urine specific gravity (USG) and adrenal ultrasonographic findings. There was no association between LDDST patterns and clinical signs (p = 0.11), increased ALP (p = 0.32), USG (p = 0.33) or adrenal ultrasonographic findings (p = 0.19). In all dogs that demonstrated complete suppression or an inverse pattern, CS was excluded by the attending clinician. The diagnosis of CS was also excluded without further exploration in 23.1%, 7.5% and 5.6% of dogs that demonstrated an escape pattern, lack of suppression and partial suppression pattern, respectively. These results suggest that the clinical significance of LDDST patterns, particularly escape and inverse patterns, are misunderstood by some clinicians, leading them to prematurely exclude the diagnosis of CS.

Design of a multi-epitope-based vaccine candidate against Bovine Genital Campylobacteriosis using a reverse vaccinology approach.

BACKGROUND: Bovine Genital Campylobacteriosis (BGC), a worldwide distributed venereal disease caused by Campylobacter fetus subsp. venerealis (Cfv), has a relevant negative economic impact in cattle herds. The control of BGC is hampered by the inexistence of globally available effective vaccines. The present in silico study aimed to develop a multi-epitope vaccine candidate against Cfv through reverse vaccinology. RESULTS: The analysis of Cfv strain NCTC 10354 proteome allowed the identification of 9 proteins suitable for vaccine development. From these, an outer membrane protein, OmpA, and a flagellar protein, FliK, were selected for prediction of B-cell and T-cell epitopes. The top-ranked epitopes conservancy was assessed in 31 Cfv strains. The selected epitopes were integrated to form a multi-epitope fragment of 241 amino acids, which included 2 epitopes from OmpA and 13 epitopes from FliK linked by GPGPG linkers and connected to the cholera toxin subunit B by an EAAAK linker. The vaccine candidate was predicted to be antigenic, non-toxic, non-allergenic, and soluble upon overexpression. The protein structure was predicted and optimized, and the sequence was successfully cloned in silico into a plasmid vector. Additionally, immunological simulations demonstrated the vaccine candidate's ability to stimulate an immune response. CONCLUSIONS: This study developed a novel vaccine candidate suitable for further in vitro and in vivo experimental validation, which may become a useful tool for the control of BGC.

Polimorfismo rs4998 del gen ADRB3 y su relación con el índicede masa corpora / Polymorphism rs4998 of the ADRB3 gene and its relationship with the bodymass index

Introducción: Para el 2007 en Colombia se estimó que el 56% de la población adulta era obesa, condición que se considera un problema de salud pública con implicaciones en el desarrollo de enfermedad coronaria. Objetivo: Determinar la distribución de las frecuencias alélicas y genotípicas y la relación del polimorfismo rs4998 del gen ADRB3 con el índice de masa corporal en una población de docentes y administrativos. Materiales y Métodos: La extracción de ADN se llevó a cabo mediante un método comercial, para la amplificación de ADN se emplearon cebadores específicos no marcado con fluorocromos, los fragmentos obtenidos fueron purificados mediante las enzimas SAP y EXOSAP, finalmente se realizó minisecuenciación en el ABI PRISM 310 y posterior lectura de electroferogramas. El análisis de las variables sociode mográficas se realizó mediante métodos univariados y métodos bivariados empleando pruebas de significancia como: test Chi2 de Pearson y test exacto de Fisher. Modelos logísticos fueron empleados para evaluar asociación. Resultados: Las frecuencias genotípicas encontradas fueron 0.96 GG, 0.014 CG y 0,020 CC. Las frecuencias alélicas fueron 0,027 C y 0,973 G. La muestra se encontró en equilibrio de Hardy-Weinberg con Chi2 = 81.153(1) (p = 0.000) y se calculó la estructura poblacional mediante la prueba Fst = 0,743. Discusión: Encontrar la población en equilibrio de Hardy-Weinberg indica que los individuos se están mezclando al azar. El coeficiente de endogamia o estadístico Fst, indica que la población presenta una alta estructura o diferenciación genética y existe fijación del alelo mutado G; esto podría obtener falsas asociaciones con la patología de interés. Conclusión: Las frecuencias obtenidas en este estudio no se ajustan a los modelos génicos propuestos para estudios de asociación; por lo tanto, se recomienda aumentar el tamaño de la muestra para futuras investigaciones o plantear estudios de casos y controles

Introduction: At 2007 in Colombia it was estimated that a portion 56 % from the adult population was obese, a condition that is considered a public health problem with implications in the development of coronary heart disease.Objective: Determine the distribution of allelic and genotypic frequencies and the relation of the polymorphism rs4998 of the ADRB3 gene with the Body Mass Index in a population of teachers and administrators of an university institution in Colombia. Methods: DNA extraction was carried out by a commercial method, for specific DNA amplification, specific primers not labeled with fluorochromes were used, fragments obtained were purified by SAP and EXOSAP enzymes, and finally minisequencing was performed in the ABI PRISM 310 and later electropherogram reading. The analysis of the sociodemographic variables was performed using univariate methods and bivariate methods using significance tests such as: Pearson's Chi2 test and Fisher's exact test. Logistic models were used to evaluate association. Results: The genotype frequencies found were 0.96 GG, 0.014 CG and 0.020 CC. The allele frequencies were 0.027 C and 0.973 G. The sample was found in Hardy-Weinberg equilibrium with Chi2 = 81.153 (1) (p = 0.000) and the population structure was calculated by means of the Fst test = 0.743. Discussion: Finding the population in Hardy-Weinberg equilibrium, indicates that individuals were mixed randomly. The coefficient of inbreeding or statistical Fst, is very close to one, which indicates that the population has a high structure or genetic differentiation and there is fixation of the mutated allele G. This could obtain false associations with the pathology of interest. Conclusion: The frequencies obtained in this study do not fit the proposed gene models for association studies, therefore, it is recommended to increase the sample size for future researches or to propose case-control studies