Bovine ovarian stem cells differentiate into germ cells and oocyte-like structures after culture in vitro.

Stem cells have been isolated from ovaries, and their ability to differentiate into oocytes in vitro has been demonstrated for mice and human, but not for bovine species. The aims of this study were to isolate germline stem cells from bovine ovaries and to evaluate the effects of bone morphogenetic proteins (BMPs) 2 and 4, and follicular fluid on the differentiation of these stem cells into oocyte-like structures. The ovarian stem cells were isolated and cultured in α-MEM+ supplemented with BMP2, BMP4 or follicular fluid. On days 0 and 14, cells were evaluated for their morphological appearance, viability, expression of alkaline phosphatase and for markers of germ cell formation (VASA and DAZL) and oocyte development (GDF9, ZPA and SCP3) by qPCR. Levels of mRNA were analysed using ANOVA and Bonferroni test (p < .05). The results showed that at day 0, ovarian stem cells expressed specific markers of pluripotency (OCT4, SOX). In addition, these cells were positive for alkaline phosphatase, which is a marker commonly used to identify primordial germ cells (PGCs). After the period of differentiation, cells had morphological features that resemble PGCs and oocyte-like cells (OLCs). An increase, ranging from five to 14 times, in the expression of VASA was observed in cells cultured in medium supplemented with BMPs and follicular fluid, while the increase in DAZL expression ranged from four to six times. In addition, OLCs had an increase in expression of mRNAs for GDF9, ZPA and SCP3 that ranged from two to eight times. In conclusion, OLCs can be differentiated in vitro from ovarian stem cells and BMPs and follicular fluid are effective in stimulating the expression of mRNAs for germ cell and oocyte markers.

Digital Smile Design for Computer-assisted Esthetic Rehabilitation: Two-year Follow-up.

OBJECTIVE: The esthetics of the smile are related to the color, shape, texture, dental alignment, gingival contour, and the relationship of these with the face. PURPOSE: To present a two-year follow-up for an esthetic rehabilitation clinical case in which the method of digital smile design (DSD) was used to assist and improve diagnosis, communication, and predictability of treatment through an esthetic analysis of the assembly: face, smile, periodontal tissue, and teeth. CLINICAL PROCEDURE: The smile's esthetics were improved through gingival recontouring, dental home bleaching, and a restorative procedure with thin porcelain laminate veneers using lithium disilicate glass-ceramic (e.max Ceram, Ivoclar-Vivadent) laminates on teeth 4 through 13. DISCUSSION: The proposed technique had an acceptable clinical performance at the end of a two-year follow-up. SIGNIFICANCE: DSD can be used to increase professional/patient communication and to provide greater predictability for the smile's esthetic rehabilitation.

Protein and messenger RNA expression of interleukin 1 system members in bovine ovarian follicles and effects of interleukin 1ß on primordial follicle activation and survival in vitro.

This study aimed to investigate the expression of interleukin 1 (IL-1) system members (proteins and messenger RNA of ligands and receptors) and its distribution in ovarian follicles of cyclic cows and to evaluate the effects of IL-1ß on the survival and activation of primordial follicles in vitro. The ovaries were processed for localization of IL-1 system in preantral and antral follicles by immunohistochemical, real-time polymerase chain reaction, and Western blot analysis. For in vitro studies, ovarian fragments were cultured in α-MEM(+) supplemented with IL-1ß (0, 1, 10, 50, or 100 ng/mL), and after 6 d, the cultured tissues were processed for histologic analysis. Immunohistochemical results showed that the IL-1 system proteins IL-1ß, IL-1RA, IL-1RI, and IL-1RII were detected in the cytoplasm of oocytes and granulosa cells from all follicular categories and theca cells of antral follicles. Variable levels of messenger RNA for the IL-1 system members were observed at different stages of development. After 6 d of culture, the presence of IL-1ß (10 or 50 ng/mL) was effective in maintaining the percentage of normal follicles and in promoting primordial follicle activation. In conclusion, IL-1 system members are differentially expressed in ovarian follicles according to their stage of development. Moreover, IL-1ß promotes the development of primordial follicles. These results indicate an important role of the IL-1 system in the regulation of bovine folliculogenesis.

An attempt to correlate fat and protein content of biological samples with residual carbon after microwave-assisted digestion.

The residual carbon content of a variety of bovine-derived samples and forage was determined by inductively coupled plasma optical emission spectrometry with radial view configuration (ICP-OES) after microwave-assisted digestion under high pressure in a closed vessel. The original carbon concentration in the samples was determined by elemental analysis. The highest amount of original carbon content (64%) was found in viscera. After digestion, up to 75% of it was destroyed. Viscera presented the highest ether extract and blood exhibited a high crude protein content of up to 99%. The efficiency in destroying the organic matter in biological materials seemed to be related to their fat content and showed no significant difficulty for protein-rich samples. The correlation coefficient between the fat content of the samples and the residual carbon after acid decomposition was 0.9173 indicating a fair fit. However, no correlation was observed between % RC and the protein content.