Comparative morphology of gonads from six species of fish belonging to the family Anostomidae (Characiformes: Anostomidae) / Morfología comparativa de las gónadas de seis especies de peces de la familia Anostomidae (Characiformes: Anostomidae)

AbstractIn fishes, gonad morphology is an important parameter to understand the reproductive biology, phylogenetic relationships and systematic studies. The main objective of this study was to make a comparative analysis of the morphology of mature ovary and testis in six fish species of the family Anostomidae. A total of 212 specimens, presenting maturing gonads, were captured from the São Francisco River, Três Marias Reservoir area, in Southeastern Brazil, between August 2008 and December 2010. The six analyzed species had mature ovaries and showed a grayish color. In vitellogenic oocytes (VO), cortical alveoli (CA) were composed by small vesicles in all Leporinus species, but in Leporellus vittatus and Schizodon knerii the CA were large vesicles. However, the CA of all species showed similar histochemical content. The pellucid zone consists of two layers to all species, but it was thicker in S. knerii (11.5 ± 1.8 µm) than in L. vittatus (9.0 ± 0.8 µm) and Leporinus species (3.5 ± 0.6 - 8.7 ± 0.9 µm). Follicular cells of the VO were respectively cubic and prismatic in animal and vegetative poles of S. knerii (22.3 ± 3.2 and 61.1 ± 9.6 µm), and squamous in the other species (1.3 ± 0.3 - 1.6 ± 0.3 µm). Females of S. knerii and males of L. vittatus showed lower values of gonadosomatic index (GSI) than the other assessed species. All evaluated species presented testicular morphology similar to most neotropical Characiformes, with unrestricted spermatogonial testis and anastomosing tubular organization of seminiferous tubules. Phenotypical differences in the vitellogenic oocyte of Anostomidae fishes, confirmed the taxonomic position of S. knerii as different genus in relation to Leporinus and L. vittatus. Despite being placed in different genera, the characteristics of the vitellogenic oocytes of L. vittatus were similar to those found in the studied Leporinus.

ResumenTeniendo en cuenta que la morfología de las gónadas es un parámetro importante para la comprensión de la biología de la reproducción, las relaciones filogenéticas y estudios sistemáticos, el objetivo principal de este estudio fue realizar un análisis comparativo de la morfología de ovarios maduros y testículos en seis especies de peces de la familia Anostomidae. Un total de 212 especímenes, con las gónadas en maduración, fueron capturados en el río São Francisco, área de embalse de Três Marias, en el sureste de Brasil, entre agosto 2008 y diciembre 2010. Las seis especies analizadas mostraron ovarios maduros de un color grisáceo. En los ovocitos vitelogénicos (VO), los alvéolos corticales (CA) están compuestos por pequeñas vesículas en todas las especies de Leporinus, pero en Leporellus vittatus y Schizodon knerii los CA son grandes vesículas. Los CA de todas las especies mostraron, no obstante, un contenido histoquímico similar. La zona pelucida se compone de dos capas en todas las especies, pero es más gruesa en S. knerii (11.5 ± 1.8 µm) que en L. vittatus (9.0 ± 0.8 µm) y las especies de Leporinus (3.5 ± 0.6 - 8.7 ± 0.9 µm). Las células foliculares de los VO son, cúbico y prismática en polos animales y vegetativos de S. knerii (22.3 ± 3.2 y 61.1 ± 9.6 µm), respectivamente, y escamosas en las otras especies (1.3 ± 0.3 - 1.6 ± 0.3 µm). Las hembras de S. knerii y los machos de L. vittatus mostraron menor GSI que las otras especies evaluadas. Todas las especies presentan una morfología testicular similar a la mayoría de los Characiformes neotropicales, los cuales poseen testículos espermatogonias sin restricciones y organización tubular anastomosis de los túbulos seminíferos. Las diferencias fenotípicas en los VO entre las especies de Anostomidae confirman la posición taxonómica de S. knerii como género diferente en relación a Leporinus y L. vittatus. A pesar de ser colocados en diferentes géneros, las características de los ovocitos vitelogénicos de L. vittatus son similares a los encontrados en los Leporinus estudiados.

Efecto del láser infrarrojo sobre células tiroideas / Infrared laser effect on thyroid cells

Diez ratas Sprague Dawley de 4 meses de vida y peso aproximado de 250 g fueron divididas en dos grupos de 5 animales cada uno, el grupo A se mantuvo como control y los animales del grupo B recibieron estimulaciones con láser infrarrojo en la tiroides con dosis de 16 J/cm2 durante 15 días consecutivos. Posteriormente las ratas fueron sacrificadas, se extrajeron las respectivas tiroides siendo procesadas para microscopía óptica y se obtuvieron placas histológicas y micrografías de tiroides con aumentos finales de hasta 1000X, las cuales fueron sometidas a estudios morfométricos para determinar en 100 células foliculares: número, áreas y perímetro tanto celular como nuclear, además de disposición coloidal y presencia de vasos sanguíneos. El análisis de los resultados entre las 100 células foliculares pertenecientes a tiroides normal y estimulada revela que existen marcadas diferencias en todos los componentes analizados los que se podría traducir en distintas funcionalidades en el metabolismo de las respectivas glándulas.

Ten 4-month-old Sprague Dawley rats weighing approximately 250 g were divided into two groups of 5 animals each. Group A was the control and the animals in group B received thyroid stimulation with infrared laser in a dose of 16 J/cm2 for 15 consecutive days. Subsequently, rats were euthanized and thyroids were removed and processed for optical microscopy. From both cell types thyroid histological slides and micrographs were obtained with final increases of 400 and 1000X. Morphometric analysis determined the number, areas and cell perimeter as well as colloidal dispersion and presence of blood vessels in 100 follicular cells. Analysis of the results among the 100 follicular cells belonging to normal and stimulated thyroids revealed marked differences in all the analyzed components, which could translate into different functionalities in the metabolism of the respective glands.

Protective effect of phloroglucinol against gamma radiation-induced oxidative stress in hair follicles

When exposed to gamma-rays, hair follicular cells immediately go through apoptosis, which hampers their rapid differentiation essential for the regeneration of hair. Phloroglucinol (PG) is a phenolic compound of Ecklonia cava, brown algae abundant in Jeju island, Korea. Containing plentiful polyphenols, PG is known for its instructive effects by inhibiting apoptosis, scavenging oxygen radicals, and protecting cells against oxidative stress. In this study, we demonstrate that PG rescues radiosensitive hair follicular cells from gamma radiation-induced apoptosis and DNA damage. To identify protective capacity of PG on hair follicles, we irradiated with 8.5 Gy (1.5 Gy/min) of gamma-rays to the whole body of C57BL/6 mice at day 6 after depilation with or without PG. In mice exposed to radiation, the expression of proapoptotic molecule p53 was downregulated in the skin of PG treated group. On immunohistochemical observation of the skin, PG inhibited the immunoreactivity of p53 and cleaved caspase-3. PG treatment protected hair follicular cells from cell death due to gamma-radiation. Our results suggest that PG presents radioprotective effects by inhibiting apoptosis of radiosensitive hair follicular cells and can protect hair follicular cells from gamma-ray induced damage.

Dexamethasone thyroid follicular cell proliferation and apoptosis in mice and its mechanisms / 中国免疫学杂志

Objective:To discuss dexamethasone on proliferation of mouse thyroid follicular cells and apoptosis. Methods:Taken BALB/c mice thyroid tissue to trypsin+Ⅱcollagenase digestion organizations get thyroid follicular cells,and expression of thyro-globulin determined whether or not the target cell. Then different concentrations of dexamethasone to stimulate target cells,and the use of MTT,flow cytometry cell proliferation rate,apoptosis rate and the apoptosis-related gene expression analysis. Results: Trypsin joint type Ⅱ collagenase treatment of thyroid tissue to obtain a stable passage of thyroid follicular epithelial cells,and cells stably expressing thyroglobulin. At the same time, different concentrations of dexamethasone on cell proliferation difference was statistically significant (F=8. 544, P<0. 05 ), and the suppression of drug action have interaction ( F = 4. 532, P<0. 05 );in addition, differently dexamethasone concentration 10-6 mol/L, 10-5 mol/L, 10-4 mol/L, the apoptosis rates were 13. 39% ± 0. 79%, 17. 43% ± 1. 38%, 26. 42%±1. 74%,both with 0 mol/L to plug betamethasone 4. 51%± 0. 06% apoptosis rate differences were statistically significant (P<0. 05,P<0. 01),while the difference in the expression of apoptotic genes trend still showed a dose-dependent manner. Conclusion:Dexamethasone can effectively inhibit thyroid follicular cell proliferation and induce apoptosis through a variety of apoptotic pathways.

A spatial model showing differences between juxtacrine and paracrine mutual oocyte-granulosa cells interactions.

The bidirectional communication between oocytes and granulosa cells are mediated by several factors via a local feedback loop(s). The current model was carried out to study the spatial mutual interaction of porcine denuded oocytes and granulosa cells either in direct contact (juxtacrine) or paracrine co-culture using transwell system. Transwell 0.4 µm polyester membrane inserts were used to permit oocytes-granulosa cells paracrine communication with a distance of 2 mm between them in co-culture. Oocytes were cultured with granulosa cells in a defined basic maturation medium for 44 h. In results, oocyte secreted factors (OSFs; GDF9 and BMP15) temporal expression showed progressive decrement by the end of culture in case of direct contact with granulosa cells while it was increased progressively in the paracrine co-culture groups. However, oocytes that were cultured in direct contact showed a significant increase in blastocyst development after parthenogenetic activation than the paracrine co-cultured ones (20% vs. 11.5%, respectively). By the end of culture, granulosa cell count in direct contact showed a significant decrease than the indirect co-culture group (1.2 × 105 cell/mL vs. 2.1 × 105 cell/mL, respectively). Steroids (P4 and E2) and steriodogenesis enzymes mRNA levels showed significant temporal alterations either after 22 h and 44 h of IVM in both juxtacrine and paracrine co-culture systems (P ≤ 0.05). CX43 was much more highly expressed in the granulosa of the direct contact group than the indirect co-culture group. These results indicate the difference in mutual communication between oocytes and granulosa cells that were cocultured either in direct contact (juxtacrine) or with a short distance (paracrine) and propose a new paradigm to study different ovarian follicular cells interaction.

Autologous somatic cell nuclear transfer in pigs using recipient oocytes and donor cells from the same animal

The objective of the present study was to examine the feasibility of the production of autologous porcine somatic cell nuclear transfer (SCNT) blastocysts using oocytes and donor cells from slaughtered ovaries. Therefore, we attempted to optimize autologous SCNT by examining the effects of electrical fusion conditions and donor cell type on cell fusion and the development of SCNT embryos. Four types of donor cells were used: 1) denuded cumulus cells (DCCs) collected from in vitro-matured (IVM) oocytes; 2) cumulus cells collected from oocytes after 22 h of IVM and cultured for 18 h (CCCs); 3) follicular cells obtained from follicular contents and cultured for 40 h (CFCs); and 4) adult skin fibroblasts. The DCCs showed a significantly (p > 0.01) lower rate of fusion than the CCCs when two pulses of 170 V/mm DC were applied for 50 microsec (19 +/- 2% vs. 77 +/- 3%). The rate of DCC fusion with oocytes was increased by the application of two DC pulses of 190 V/mm for 30 microsec, although this was still lower than the rate of fusion in the CCCs (33 +/- 1% vs. 80 +/- 2%). The rates of cleavage (57 +/- 5%) and blastocyst formation (1 +/- 1%) in the DCC-derived embryos did not differ from those (55 +/- 6% and 3 +/- 1%, respectively) in the CCC-derived SCNT embryos. Autologous SCNT embryos derived from CFCs (5 +/- 2%) showed higher levels of blastocyst formation (p > 0.01) than CCC-derived autologous SCNT embryos (1 +/- 0%). In conclusion, the results of the present study show that culturing cumulus and follicular cells before SCNT enhances cell fusion with oocytes and that CFCs are superior to CCCs in the production of higher numbers of autologous SCNT blastocysts.