Study protocol to investigate biomolecular muscle profile as predictors of long-term urinary incontinence in women with gestational diabetes mellitus.

BACKGROUND: Pelvic floor muscles (PFM) and rectus abdominis muscles (RAM) of pregnant diabetic rats exhibit atrophy, co-localization of fast and slow fibers and an increased collagen type I/III ratio. However, the role of similar PFM or RAM hyperglycemic-related myopathy in women with gestational diabetes mellitus (GDM) remains poorly investigated. This study aims to assess the frequency of pelvic floor muscle disorders and pregnancy-specific urinary incontinence (PS-UI) 12 months after the Cesarean (C) section in women with GDM. Specifically, differences in PFM/RAM hyperglycemic myopathy will be evaluated. METHODS: The Diamater is an ongoing cohort study of four groups of 59 pregnant women each from the Perinatal Diabetes Research Centre (PDRC), Botucatu Medical School (FMB)-UNESP (São Paulo State University), Brazil. Diagnosis of GDM and PS-UI will be made at 24-26 weeks, with a follow-up at 34-38 weeks of gestation. Inclusion in the study will occur at the time of C-section, and patients will be followed at 24-48 h, 6 weeks and 6 and 12 months postpartum. Study groups will be classified as (1) GDM plus PS-UI; (2) GDM without PS-UI; (3) Non-GDM plus PS-UI; and (4) Non-GDM without PS-UI. We will analyze relationships between GDM, PS-UI and hyperglycemic myopathy at 12 months after C-section. The mediator variables to be evaluated include digital palpation, vaginal squeeze pressure, 3D pelvic floor ultrasound, and 3D RAM ultrasound. RAM samples obtained during C-section will be analyzed for ex-vivo contractility, morphological, molecular and OMICS profiles to further characterize the hyperglycemic myopathy. Additional variables to be evaluated include maternal age, socioeconomic status, educational level, ethnicity, body mass index, weight gain during pregnancy, quality of glycemic control and insulin therapy. DISCUSSION: To our knowledge, this will be the first study to provide data on the prevalence of PS-UI and RAM and PFM physical and biomolecular muscle profiles after C-section in mothers with GDM. The longitudinal design allows for the assessment of cause-effect relationships between GDM, PS-UI, and PFMs and RAMs myopathy. The findings may reveal previously undetermined consequences of GDM.

Effects of a phytogenic, alone and associated with potassium diformate, on tilapia growth, immunity, gut microbiome and resistance against francisellosis.

This work evaluated the effects of dietary supplementation of A-Live (phytogenic) either individually or in combination with Aquaform (potassium diformate, acidifier) on juvenile Nile tilapia (Oreochromis niloticus) growth performance, innate immune parameters, gut microbiome, and resistance against Francisella noatunensis subsp. orientalis challenge. Each experimental group contained 140 fishes (34.3 ± 0.33) in two 150L tanks. The experimental design consisted of five groups: a negative control; treated groups (G1, G2, G3) supplemented with different concentrations of A-Live and Aquaform in the feed; and a positive control (PC) for pathogen infection. Groups G1, G2, G3, and PC were challenged with Francisella spp. after 15 days. After infection, the mortality was significantly lower in groups G1, G2, and G3 (p < 0.01). Furthermore, these groups showed significant increase (p < 0.05) in daily weight gain, feed conversion rate, and specific growth rate. The PC group presented increase (p < 0.05) in the leukocytes and neutrophils number. Innate immunity parameters showed no difference between treatments after infection. Microbiome analysis revealed an increased number of bacteria belonging to the Vibrionaceae family after pathogen infection suggesting a secondary pathogen function of these bacteria. These results validate the beneficial effects of these products in tilapia farming.

Variabilidade genética de larvas e alevinos de piracanjuba (Brycon orbignyanus) / Genetic variability of larvae and fingerlings of Piracanjuba (Brycon orbignyanus)

O objetivo do presente trabalho foi avaliar a variabilidade genética de larvas e alevinos de piracanjuba em programa de repovoamento. Foram coletadas 180 larvas de piracanjuba de três dias e 90 alevinos de três meses de idade. Foram avaliados cinco loci microssatélites, os quais produziram 19 alelos. Não houve presença de alelos raros nem perdas de alelos ao longo do período. A heterozigosidade observada foi superior nas larvas em relação aos alevinos. Houve desvio no equilíbrio de Hardy-Weinberg na maioria dos loci em ambos os grupos. O coeficiente de endogamia foi positivo em ambos os grupos, sendo a média dos alevinos superior em relação às larvas. O excesso de heterozigotos foi significativo no modelo Stepwise Mutation Model para os alevinos, indicando a possibilidade de efeito gargalo recente. Conclui-se que, apesar da adequada variabilidade genética encontrada, os valores do coeficiente de endogamia e a possibilidade de efeito gargalo nos alevinos atentam para a necessidade de constante monitoramento genético desses estoques antes da liberação no ambiente.(AU)

The objective of this study was to evaluate the genetic variability of Piracanjuba larvae and fingerlings in restocking program. A total of 180 three-day Piracanjuba larvae and 90 three-month-old fish were sampled. Five microsatellite loci were evaluated, which produced 19 alleles. There were no rare alleles or loss of alleles over the period. The observed heterozygosity was higher in larvae compared to fingerlings. There was a deviation in Hardy-Weinberg equilibrium in most loci in both groups. The inbreeding coefficient was positive in both groups, with the average of the fingerlings superior to the larvae. The excess heterozygotes were significant in the Stepwise Mutation Model for the fingerlings, indicating the possibility of a recent bottleneck effect. Despite the adequate genetic variability found, the values of the inbreeding coefficient and the possibility of bottleneck effect in the fingerlings show the need for constant genetic monitoring of these stocks prior to release into the environment.(AU)

Effect of propolis ethanol extract on myostatin gene expression and muscle morphometry of Nile tilapia in net cages.

Propolis can be used as growth enhancer due to its antimicrobial, antioxidant, and immune-stimulant properties, but its effects on morphometry and muscle gene expression are largely unknown. The present study evaluates the influence of propolis on muscle morphometry and myostatin gene expression in Nile tilapia (Oreochromis niloticus) bred in net cages. Reversed males (GIFT strain) with an initial weight of 170 ± 25 g were distributed in a (2 x 4) factorial scheme, with two diets (DPRO, commercial diet with 4% propolis ethanol extract and DCON, commercial diet without propolis, control) and four assessment periods (0, 35, 70, and 105 experimental days). Muscles were evaluated at each assessment period. Histomorphometric analysis classified the fiber diameters into four groups: <20 µm; 20-30 µm; 30-50 µm; and > 50 µm. RT-qPCR was performed to assess myostatin gene expression. Fibers < 20 µm diameter were more frequent in DPRO than in DCON at all times. Fiber percentages >30 µm (30-50 and > 50 µm) at 70 days were 25.39% and 40.07% for DPRO and DCON, respectively. There was greater myostatin gene expression at 105 days, averaging 1.93 and 1.89 for DCON and DPRO, respectively, with no significant difference in any of the analyzed periods. Propolis ethanol extract did not affect the diameter of muscle fibers or the gene expression of myostatin. Future studies should describe the mechanisms of natural products' effects on muscle growth and development since these factors are highly relevant for fish production performance.

Morphology and muscle gene expression in GIFT and Supreme Nile tilapia varieties reared in two cultivation systems.

Tissue growth in most fishes occurs by muscular hyperplasia and hypertrophy, which are influenced by different regulatory factors, such as myostatin. The current study evaluated the influence of cultivation in hapas and earthen ponds on the diameter of white muscle fibers and on the myostatin (MSTN-1) gene in GIFT and Supreme varieties of tilapia. Fish of both varieties were reared for 204 days and then divided into four developmental stages. White muscle samples, corresponding to 100 fibers per slide, were collected from the middle region of fish of each variety and cultivation system, and were measured and divided into two classes representing hyperplasia and hypertrophy. Samples were subjected to real-time PCR to analyze gene expression. Hyperplasia decreased during the developing stages, coupled with increased hypertrophy. There was a higher rate of hypertrophy in fish raised in earthen ponds when compared to those raised in hapas, during juvenile and developing phases, and greater hypertrophic growth was observed in GIFT specimens when compared to Supreme specimens in earthen ponds. Since increased MSTN-1 gene expression was observed in GIFT specimens during the developing phase in pond cultivations, and in Supreme tilapia in hapas, MSTN-1 expression is related to greater hypertrophy. These results demonstrate the capacity for increased muscle growth in earthen pond cultivation in which the GIFT variety developed best. How the environment affects the growth of different tilapia varieties may be employed to optimize culture management and genetic improvement programs. Further investigations should aim to describe mechanisms affecting muscle growth and development.

Monitoramento e conservação genética de populações naturais de Prochilodus lineatus dos rios Pardo, Mogi-Guaçu e Tietê, São Paulo / Monitoring and conservation genetics of Prochilodus lineatus wild populations of Pardo, Mogi Guaçu and Tiete rivers, São Paulo

O objetivo do presente estudo foi determinar a diversidade e a estrutura genética de seis populações naturais de Prochilodus lineatus em usinas hidrelétricas (UHE) dos rios Pardo (UHE Limoeiro - LMO), Mogi-Guaçu (UHE Mogi-Guaçu - MOG) e Tietê (UHE Promissão - PRO, UHE Barra Bonita - BAB, UHE Nova Avanhandava - NAV e UHE Bariri - BAR). Foi encontrado um total de 47 alelos, com tamanhos entre 118pb e 330pb. Os resultados de heterozigosidade média observada (0,490 a 0,625) refletiram uma alta variabilidade genética intrapopulacional. Os valores de distância genética (0,149 a 0,773), Fst (0,006 a 0,218) e Nm (1,2 a 4,2) mostraram a presença de similaridade genética entre as populações. De acordo com a AMOVA, houve maior variação dentro das populações do que entre elas. O dendograma mostrou a formação de dois agrupamentos (LMO-PRO-MOG e BAR-BAB-NAV). Concluiu-se que as populações naturais apresentaram alta variabilidade genética, com similaridade genética entre elas, possivelmente causada pelo programa de repovoamento realizado nesses rios.(AU)

The aim of this study was to determine the genetic diversity and structure of six wild populations of Prochilodus lineatus in Hydroelectric Power Plants (HPP) of the Pardo (HPP Limoeiro - LMO), Mogi Guacu (HPP Mogi-Guaçu - MOG), and Tiete (HPP Promissão - PRO, HPP Barra Bonita - BAB, HPP Nova Avanhandava - NAV and HPP Bariri - BAR) rivers. A total of 47 alleles, ranging in size from 118bp to 330bp were found. The results of observed heterozygosity average (0.490 to 0.625) reflected a high intra-population genetic variability. The values of genetic distance (0.149 to 0.773), Fst (0.006 to 0.218), and Nm (1.2 to 4.2) showed that between the populations there is genetic similarity. According to AMOVA there was higher variation within populations than between them. The dendrogram demonstrated the formation of two groups (LMO-PRO-MOG and BAR-BAB-NAV). It was concluded that wild populations had high genetic variability with genetic similarity between them, possibly caused by the restocking program performed in these rivers.(AU)

HFE gene mutations in Brazilian thalassemic patients.

Hereditary hemochromatosis is a disorder of iron metabolism characterized by increased iron intake and progressive storage and is related to mutations in the HFE gene. Interactions between thalassemia and hemochromatosis may further increase iron overload. The ethnic background of the Brazilian population is heterogeneous and studies analyzing the simultaneous presence of HFE and thalassemia-related mutations have not been carried out. The aim of this study was to evaluate the prevalence of the H63D, S65C and C282Y mutations in the HFE gene among 102 individuals with alpha-thalassemia and 168 beta-thalassemia heterozygotes and to compare them with 173 control individuals without hemoglobinopathies. The allelic frequencies found in these three groups were 0.98, 2.38, and 0.29% for the C282Y mutation, 13.72, 13.70, and 9.54% for the H63D mutation, and 0, 0.60, and 0.87% for the S65C mutation, respectively. The chi-square test for multiple independent individuals indicated a significant difference among groups for the C282Y mutation, which was shown to be significant between the beta-thalassemia heterozygote and the control group by the Fisher exact test (P value = 0.009). The higher frequency of inheritance of the C282Y mutation in the HFE gene among beta-thalassemic patients may contribute to worsen the clinical picture of these individuals. In view of the characteristics of the Brazilian population, the present results emphasize the need to screen for HFE mutations in beta-thalassemia carriers.

HFE gene mutations in Brazilian thalassemic patients

Hereditary hemochromatosis is a disorder of iron metabolism characterized by increased iron intake and progressive storage and is related to mutations in the HFE gene. Interactions between thalassemia and hemochromatosis may further increase iron overload. The ethnic background of the Brazilian population is heterogeneous and studies analyzing the simultaneous presence of HFE and thalassemia-related mutations have not been carried out. The aim of this study was to evaluate the prevalence of the H63D, S65C and C282Y mutations in the HFE gene among 102 individuals with alpha-thalassemia and 168 beta-thalassemia heterozygotes and to compare them with 173 control individuals without hemoglobinopathies. The allelic frequencies found in these three groups were 0.98, 2.38, and 0.29 percent for the C282Y mutation, 13.72, 13.70, and 9.54 percent for the H63D mutation, and 0, 0.60, and 0.87 percent for the S65C mutation, respectively. The chi-square test for multiple independent individuals indicated a significant difference among groups for the C282Y mutation, which was shown to be significant between the beta-thalassemia heterozygote and the control group by the Fisher exact test (P value = 0.009). The higher frequency of inheritance of the C282Y mutation in the HFE gene among beta-thalassemic patients may contribute to worsen the clinical picture of these individuals. In view of the characteristics of the Brazilian population, the present results emphasize the need to screen for HFE mutations in beta-thalassemia carriers.

High incidence of single nucleotide polymorphisms in the prion protein gene of native Brazilian Caracu cattle.

Different alleles of the human and ovine prion protein gene correlate with a varying susceptibility to transmissible spongiform encephalopathies. However, the pathogenic implications of specific polymorphisms in the bovine prion protein gene (PRNP) are only poorly understood. Previous studies on the bovine PRNP gene investigated common European and North American cattle breeds. As a consequence of decades of intensive breeding for specific traits, these modern breeds represent only a small fraction of the bovine gene pool. In this study, we analysed PRNP polymorphisms in the native Brazilian Caracu breed, which developed in geographical isolation since the 16th century. A total of 10 single nucleotide polymorphisms (SNPs) were discovered in the coding region of the Caracu PRNP gene. Eight of the SNPs occurred at high frequencies in Caracu cattle (variant allele frequencies = 0.10-0.76), but were absent or only rarely observed in European and North American breeds. One of the Caracu SNPs was associated with an amino acid exchange from serine to asparagine (f = 0.17). This SNP was not detected in Holstein-Friesian, Simmental and German Gelbvieh and was only rarely detected in beef cattle (f = 0.01). We found 17 haplotypes for PRNP in the Caracu breed.

Results of a decellularized porcine heart valve implanted into the juvenile sheep model.

OBJECTIVE: This study was performed to evaluate the possibility of creating a glutaraldehyde-free porcine xenograft to improve long-term durability. METHODS: A decellularized porcine pulmonary valve was implanted into the right ventricular outflow tract of 7 juvenile sheep. Valves were explanted after 3 months (n = 4) and 6 months (n = 3). Evaluation was performed by gross examination, radiography, histology (hematoxylin-eosin and Sirius red staining), and immunohistochemistry. Quantitative determination of calcium content was investigated by atomic absorption spectrometry. RESULTS: All animals showed fast recovery without complications. At explantation, all decellularized valves showed smooth and pliable leaflets without evidence of thrombosis. The valve wall was also smooth and pliable without hardness. Light microscopy showed a monolayer of host endothelial cells covering the inner surface of the heart valves and repopulation of host fibroblasts into the deeper layers. Sirius red staining enabled visualization of the production of new collagen. Radiographic results showed an absence of calcification, confirmed by the low calcium levels (1.08 +/- 0.28 microg/g and 0.73 +/- 0.31 microg/g at 3 and 6 months, respectively) revealed by atomic absorption spectrometry. CONCLUSIONS: The results with the juvenile sheep model showed that decellularized heart valves are recellularized in vivo. Host endothelial cells form a monolayer on the inner surface of the valve matrix. Furthermore, host fibroblasts repopulate the valve matrix and produce collagen; thus, a remodeling potential can be expected.