In vitro embryo production is a well-known biotechnology tool to improve and sustain animal productivity. Therefore, optimization of this technique would enhance both animal productivity and farm profitability. The aim of the present study was to compare the mitochondrial activity and patterns of expression of genes that contribute to its regulation during the in vitro maturation of buffalo (Bubalus bubalis) and bovine (Bos indicus) oocytes. Ovaries were collected from local Egyptian abattoirs and cumulus-oocytes complexes (COCs)were aspirated from 2-8mm follicles diameter and were divided into four categories according to oocyte morphology. The grade A and grade B were cultured in TCM medium (supplemented with all required chemicals and hormones) for 22 hours at 38.5 o C and collected after their in vitro maturation (IVM). The total RNA of the oocytes was then extracted and target mitochondrial transcripts (TFAM and CPT2) were analyzed by real-time PCR. The results of this work revealed the intensity of mitochondria and lipids was reduced in good than bad matured bovine oocytes. However, there was no change of mitochondrial and lipid fluorescent intensities of bad quality oocytes before and after in vitro maturation. The expression profile of CPT2 gene was higher in immature compared to matured oocytes of bovine while, buffalo oocytes did not shown differences in the expression of this gene. Furthermore, the expression profile of CPT2 gene was lower in immature and matured buffalo oocytes than those of bovine. The transcript abundance of TFAM did not indicate any differences among in vitro maturation of both species. It was concluded that the patterns of the gene expression of CPT2 vary during in vitro maturation of bovine oocytes in reflecting their maturation competence than that of buffalo. Increased metabolic activity of oocytes during IVM is in line with CPT2 expression that is involved in lipid oxidation required for this process.

The oocyte is the female gamete that contributes not only half of the genetic material but also all of the cytoplasm to the zygote, supplying the transcripts, proteins, mitochondria and other components necessary for early embryonic development. The intrinsic oocyte quality is one of the main factors affecting the embryo yield, the implantation rate and the rate of healthy offspring. It is obvious that a fertilized oocyte must reach the blastocyst stage within 6–9 days in the proper culture conditions to have a significant chance of inducing a pregnancy and producing an offspring. The ability to sustain the first week of embryonic development is clearly influenced by the follicular status from which the oocyte is obtained indicating that this developmental potential is inherent within certain oocytes. Since most early embryos that do not reach the blastocyst stage are blocked at or close to the maternal to zygotic transition (MZT)-stage, which occurs at the eight-cell stage in cattle, one could speculate that incompetent oocytes fail to appropriately activate the embryonic genome. Oocyte selection based on glucose-6-phosphate dehydrogenase (G6PDH) activity has been successfully used to differentiate between competent and incompetent bovine oocytes. Recently, molecular regulation of genes regulating biological process of Brilliant Cresyl Blue staining (BCB) selected oocytes and embryos was investigated to explain their variation in quality and developmental potentiality. This short review will highlights some of these efforts that have been done in this interesting area of research.

The term "oocyte competence or quality" is the fi nal measurement of its ability to achieve all cytoplasmic, nuclear and molecular events either in vivo or in vitro to bring a healthy offspring. Sirard, et al. [1] have defi ned the levels of competence as the ability of oocyte to resume meiosis, cleave following fertilization, develop and differentiate into blastocyst stage, induce pregnancy and fi nally bring healthy offspring.

The production of in vitro produced embryos of good morphological quality and viability is a prerequisite for successful assisted reproduction biotechnologies in animal breeding and human. The co-culturing system has been applied to improve preimplantation development that could subsequently resulted in successful pregnancy. There are different types of reproductive and non-reproductive cells that have been used during preimplantation development. The most well-known reproductive cells are those recovered from ovaries (cumulus and granulosa cells), oviduct and endometrium cells. While, in last decade stem cells such as mesenchymal stem cells and murine embryonic fibroblasts that originated from different tissues have been used to support early embryonic development. The positive effect co-culturing system was suggested to be due to direct mechanical cell-to-cell contact that occurred between the dividing embryos embryo and helper cells in addition to secretions of various bioactive biological components like growth factors and scavenging the deleterious byproducts that resulted from embryo metabolism. In current review, we will highlight the effects of different couture systems on embryo development and their suggested mechanisms to exert the beneficial impacts.

It is documented that heat stress caused impairment on the reproductive performance of dairy animals. However, there are few reports that have focused on the molecular and intracellular responses of in vitro cultured buffalo granulosa cells during heat elevation. The present study was conducted to investigate the effect of heat elevation during in vitro culture of buffalo granulosa cells on their viability, quality, mitochondrial activity, and transcriptional activity. Granulosa cells were harvested after aspiration of cumulus-oocytes complexes that were collected from abattoir ovaries. The granulosa cells were cultured in vitro either at a normal physiological temperature suitable for oocyte maturation and embryo development (38.5°C) or exposed to the elevated temperature of 40.5°C on day 3 of culture (the first two days were for confluence) for two hours of culture then continued at 38.5°C up to day 7 of culture. The viability of granulosa cells was measured using trypan blue and quality was estimated by measuring the level of intracellular reactive oxygen species (ROS) on day 7. Moreover, metabolic activity was performed by measuring the fluorescent intensity of mitochondria. Moreover, transcriptional activity was done by profiling four selected candidate genes using quantitative real-time PCR. The results indicated that the granulosa cells viability rate significantly decreased in the heat stress group (25.1 ± 3.7), compared to the control group (36.6 ± 5.3) on confluence day (day 3). In addition, the viability rate on the last day of culture (day 7) decreased in heat stress, compared to control (83.7 ± 4.5 and 97.4 ± 0.4, respectively). On the other hand, there was a nonsignificant difference in ROS profile between the control (21.7*104 ± 1.3) and the heat-stressed group (15.7 ± 0.7) on day 7 of culture. However, the mitochondrial fluorescent intensity was higher in the control (21.9 ± 1.9) than in the heat-stressed group (15.4 ± 0.8) on day 7 of culture. The expression of cellular defense (HSF1) and apoptosis-inducing gene (P53) were significantly up-regulated in granulosa cells exposed to heat elevation, compared to the control group. On the other hand, the steroidogenesis-regulating gene (StAR) was down-regulated in granulosa cells cultured under heat shock, compared to the control group. In conclusion, heat stress reduced the viability of granulosa cells by inducing the expression of an apoptosis-related gene (P53) and compromised expression of genes regulating the steroid biosynthesis, which resulted in up-regulation of cell defense gene (HSF1) in an attempt to ameliorate the deleterious effect of heat stress on the biological activity of the granulosa cells.