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Gad, A., U. Besenfelder, F. Rings, N. Ghanem, D. Salilew-Wondim, M. M. Hossain, D. Tesfaye, P. Lonergan, a Becker, U. Cinar, et al., "Effect of reproductive tract environment following controlled ovarian hyperstimulation treatment on embryo development and global transcriptome profile of blastocysts: implications for animal breeding and human assisted reproduction.", Human reproduction (Oxford, England), vol. 26, issue 7, pp. 1693-707, 2011. Abstract

In mammals, the reproductive tract plays a crucial role in the success of early reproductive events and provides an optimal microenvironment for early embryonic development. However, changes in the reproductive tract environment associated with controlled ovarian hyperstimulation and the influence on the embryo transcriptome profile have not been investigated. Therefore, we investigated differences in the development rate and the transcriptome profile of bovine blastocysts developing in the reproductive tract of unstimulated or superovulated heifers.

Forde, N., F. Carter, S. di Francesco, J. P. Mehta, M. Garcia-Herreros, A. Gad, D. Tesfaye, M. Hoelker, K. Schellander, and P. Lonergan, "Endometrial response of beef heifers on day 7 following insemination to supraphysiological concentrations of progesterone associated with superovulation.", Physiological genomics, vol. 44, issue 22, pp. 1107-15, 2012. Abstract

Ovarian stimulation is a routine procedure in assisted reproduction to stimulate the growth of multiple follicles in naturally single-ovulating species including cattle and humans. The aim of this study was to analyze the changes induced in the endometrial transcriptome associated with superovulation in cattle and place these observations in the context of our previous data on changes in the endometrial transcriptome associated with elevated progesterone (P4) concentrations within the physiological range and those changes induced in the embryo due to superovulation. Mean serum P4 concentrations were significantly higher from day 4 to day 7 in superovulated compared with unstimulated control heifers (P < 0.05). Between-group analysis revealed a clear separation in the overall transcriptional profile of endometria from unstimulated control heifers (n = 5) compared with superovulated heifers (n = 5). This was reflected in the number of differentially expressed genes (DEGs) identified between the two groups with 795 up- and 440 downregulated in superovulated endometria. Ten times more genes were altered by superovulation (n = 1,234) compared with the number altered due to elevated P4 within physiological ranges by insertion of a P4-releasing intravaginal device (n = 124) with only 22 DEGs common to both models of P4 manipulation. Fewer genes were affected by superovulation in the embryo compared with the endometrium, (443 vs. 1,234 DEGs, respectively), and the manner in which genes were altered was different with 64.5% of genes up- and 35.5% of genes downregulated in the endometrium, compared with the 98.9% of DEGs upregulated in the embryo. In conclusion, superovulation induces significant changes in the transcriptome of the endometrium which are distinct from those in the embryo.

Ashour, G., A. Gad, A. K. Fayed, N. A. Ashmawy, and A. El-Sayed, "Evaluation of Growth Performance, Blood Metabolites and Gene Expression Analysis in Egyptian Sheep Breeds, in Relation to Age", World’s Veterinary Journal, vol. 10, issue 4, pp. 18-29, 2020. Abstract


Abd El Naby, W. S., T. H. Hagos, M. M. Hossain, D. Salilew-Wondim, Y. a Gad, F. Rings, M. U. Cinar, E. Tholen, C. Looft, K. Schellander, et al., "Expression analysis of regulatory microRNAs in bovine cumulus oocyte complex and preimplantation embryos.", Zygote (Cambridge, England), pp. 1-21, 2011. Abstract

SummaryMicroRNAs (miRNAs) are small endogenous molecules that are involved in a diverse of cellular process. However, little is known about their abundance in bovine oocytes and their surrounding cumulus cells during oocyte development. To elucidate this situation, we investigated the relative expression pattern of sets of miRNAs between bovine oocyte and the surrounding cumulus cells during in vitro maturation using miRNA polymerase chain reaction (PCR) array. Results revealed that a total of 47 and 51 miRNAs were highly abundant in immature and matured oocytes, respectively, compared with their surrounding cumulus cells. Furthermore, expression analysis of six miRNAs enriched in oocyte miR-205, miR-150, miR-122, miR-96, miR-146a and miR-146b-5p at different maturation times showed a dramatic decrease in abundance from 0 h to 22 h of maturation. The expression of the same miRNAs in preimplantation stage embryos was found to be highly abundant in early stages of embryo development and decreased after the 8-cell stage to the blastocyst stage following a typical maternal transcript profile. Similar results were obtained by localization of miR-205 in preimplantation stage embryos, in which signals were higher up to the 4-cell stage and reduced thereafter. miR-205 and miR-210 were localized in situ in ovarian follicles and revealed a spatio-temporal expression during follicular development. Interestingly, the presence or absence of oocytes or cumulus cells during maturation was found to affect the expression of miRNAs in each of the two cell types. Hence, our results showed the presence of distinct sets of miRNAs in oocytes or cumulus cells and the presence of their dynamic degradation during bovine oocyte maturation.

Koncicka, M., J. Cervenka, D. Jahn, R. Sucha, P. Vodicka, A. Gad, M. Alsheimer, and A. Susor, "Expression of lamin C2 in mammalian oocytes", PloS one, vol. 15, issue 4: Public Library of Science, pp. e0229781, 2020. AbstractWebsite


Gebremedhn, S., A. Ali, A. Gad, R. Prochazka, and D. Tesfaye, "Extracellular Vesicles as Mediators of Environmental and Metabolic Stress Coping Mechanisms During Mammalian Follicular Development", Frontiers in Veterinary Science, vol. 7, pp. 961, 2020. AbstractWebsite

Extracellular vesicles are evolutionarily conserved nano-sized phospholipid membraned structures and released from virtually all types of cells into the extracellular space. Their ability to carry various molecular cargos (mRNA, miRNA, proteins, and lipids) from one cell to the other to exert functional impact on the target cells enables them to play a significant role in cell to cell communication during follicular development. As the molecular signals carried by extracellular vesicles reflect the physiological status of the cells of origin, they are expected to mediate any effect of environmental or metabolic stress on the follicualr cells and the growing oocyte. Recent studies have evidenced that reproductive cells exposed to various environmental stressors (heat and oxidative stress) released extracellular vesicles enriched with mRNA and miRNA associated with stress response mechanisms. Moreover, the metabolic status of post-calving cows could be well-reflected in the follicular extracellular vesicle's miRNA profile, which signified the potential role of extracellular cellular vesicle molecular signals in mediating the effect of metabolic stress on follicular and oocyte development. In the present review, the potential role of extracellular vesicles in mediating the effect of environmental and metabolic stress in various reproductive cells and oocytes are thoroughly discussed Moreover, considering the importance of extracellular vesicles in shuttling protective or rescuing molecular signals during stress, their potential usage as means of targeted delivery of molecules to mitigate the effect of stress on oocytes are addressed as the focus of future research.

Gebremedhn, S., A. Gad, H. S. Aglan, J. Laurincik, R. Prochazka, D. Salilew-Wondim, M. Hoelker, K. Schellander, and D. Tesfaye, "Extracellular vesicles shuttle protective messages against heat stress in bovine granulosa cells", Scientific Reports, vol. 10, no. 1: Nature Publishing Group, pp. 1–19, sep, 2020. AbstractWebsite

Elevated summer temperature is reported to be the leading cause of stress in dairy and beef cows, which negatively affects various reproductive functions. Follicular cells respond to heat stress (HS) by activating the expression of heat shock family proteins (HSPs) and other antioxidants. HS is reported to negatively affect the bi-directional communication between the follicular cells and the oocyte, which is partly mediated by follicular fluid extracellular vesicles (EVs) released from surrounding cells. As carriers of bioactive molecules (DNA, RNA, protein, and lipids), the involvement of EVs in mediating the stress response in follicular cells is not fully understood. Here we used an in vitro model to decipher the cellular and EV-coupled miRNAs of bovine granulosa cells in response to HS. Moreover, the protective role of stress-related EVs against subsequent HS was assessed. For this, bovine granulosa cells from smaller follicles were cultured in vitro and after sub-confluency, cells were either kept at 37 °C or subjected to HS (42 °C). Results showed that granulosa cells exposed to HS increased the accumulation of ROS, total oxidized protein, apoptosis, and the expression of HSPs and antioxidants, while the viability of cells was reduced. Moreover, 14 and 6 miRNAs were differentially expressed in heat-stressed granulosa cells and the corresponding EVs, respectively. Supplementation of stress-related EVs in cultured granulosa cells has induced adaptive response to subsequent HS. However, this potential was not pronounced when the cells were kept under 37 °C. Taking together, EVs generated from granulosa cells exposed to HS has the potential to shuttle bioactive molecules to recipient cells and make them robust to subsequent HS.