Research articleThe effect of energy balance on the transcriptome of bovine granulosa cells at 60 days postpartum
Introduction
Dairy production is an important aspect of the food industry in many countries. As the pregnancy and birth of a calf are essential to the onset of milk production in the cow, the dairy industry requires fertile animals. However, the past decades have seen a noticeable drop in the fertility of Canadian dairy cows, mainly for the Holstein breed. This trend is worrisome for producers, as it is associated with additional expenses and lower profits. The main problems observed are longer intervals between calving and first insemination [1], decreased conception rates [2], abnormal estrus [3], and increased pregnancy losses [4]. These problems considerably reduce the longevity of dairy cow herds: In 2013, 8.2% of dairy cows registered in Canada were culled for reproductive reasons (Canadian Dairy Information Centre). No single factor was proven to be the cause of the declining fertility in dairy cattle, but increased lactation demands was suggested to play a major role. Global conception rates have increased during the past few years, and the milk yield per animal also increased over the same period [5]. However, although previous studies highlighted relations between milk production and several aspects of cattle reproduction, they were not able to report a direct relationship between milk yield and fertility [6], [7].
During the postpartum period, dairy cows experience a period of negative energy balance (NEB) and need to mobilize their body reserves to cope with the energy demand of lactation [8]. The amount of energy required for milk production increases quickly after calving, whereas the increase in dry matter intake lags behind [9]. Animals selected for milk production tend to partition more energy toward milk rather than restoring body reserves [10], [11] and are therefore more susceptible to experience a prolonged period of NEB. Fertility in heifers and beef cows has not declined in a similar way as fertility in lactating cows [12], [13], [14], supporting the hypothesis of an excessive metabolic stress during lactation. However, energy balance status may affect reproduction [15] more than milk yield, as it induces metabolic stress.
During periods of fasting or of high energy demand such as lactation, body energy reserves are mobilized and lipid metabolism is activated. The released fatty acids are oxidized by the liver to produce ketone bodies, which can transport energy from the liver to other tissues. The concentration of these molecules in blood serum increases with lipid mobilization. The most abundant ketone body is beta-hydroxybutyrate (BHB), and its blood concentration is an indicator of ketosis in cattle, with subclinical ketosis ranging from 1200 to 1400 μmol/L [16]. Cows with a higher body condition score (BCS) at calving are more likely to experience ketosis [17] because they eat less than thin cows and will therefore need to mobilize more body reserves to cope with the energy demand of lactation [18], [19]. In dairy cows, high concentrations of BHB were linked to periparturient diseases [20].
The follicular environment greatly influences the competence of the oocyte [21] and, therefore, ultimately contributes to embryo quality. Granulosa cells' transcriptome is known to be altered by numerous aspects of follicular development, including growth stage [22], [23]. In vitro studies have shown that some blood metabolites are linked to energy balance, such as nonesterified fatty acids and BHB, and can influence bovine granulosa cell functions [24], [25], [26]. Our hypothesis is that energy balance has an impact on bovine granulosa cells' transcriptome. The objective of this study was to analyze the transcriptomic profiles of granulosa cells from dairy cows with more or less severe NEB at 60 days postpartum, to identify key genes and pathways connecting energy balance and follicular development.
Section snippets
Animals
Lactating Holstein cows were selected from 10 medium-sized dairy farms with 75 to 300 animals. Several small farms, with dietary and management differences, were preferred to a single large farm as a way to allow broader application of the conclusions. Animal diets and management of the farms were taken into consideration, and animals were paired accordingly for all contrasts. All samples were collected during the winter of 2013 to 2014. The farm managers were asked to provide milk recording
Microarray data
The data presented in this publication have been deposited in NCBI's Gene Expression Omnibus [35] and are accessible through GEO Series accession numbers GSE66116. The HCA graph (Supplementary Figure 1) revealed an absence of complete segregation between the groups. The BHB profile did not seem to have a similar effect in every animal and led to poor segregation. After normalization, a cutoff set at a symmetrical fold change of 1.5, an intensity of 8, and a P value of 0.05 was applied and it
Microarrays
This study used transcriptomic profile analysis of granulosa cells from dairy cows to identify genes and pathways connecting energy balance and follicular development, to provide suggestions to improve fertility in cattle. Animals were pooled to increase the number of animals per group (12 cows instead of four, as there are four arrays per slide). Pools reduce individual variations and background noise, but they also decrease the sensitivity of the analysis. However, the goal of this study was
Acknowledgments
The authors acknowledge Jean-Michel Guay and his assistants for the collection of samples. Annie Girard was supported by MAPAQ (Ministère de l'agriculture, des pêcheries et de l'alimentation du Québec), whereas Marc-André Sirard and Isabelle Dufort by NSERC (Natural Sciences and Engineering Research Council of Canada).
Author contributions: Annie Girard and Marc-André Sirard conceived the study and wrote the article. Annie Girard supervised sample collections, performed the extractions,
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Embryonic response to high beta-hydroxybutyrate (BHB) levels in postpartum dairy cows
2020, Domestic Animal EndocrinologyCitation Excerpt :Although the cells are capable of adapting to variations in nutrient availability, such adaptation often comes at the cost of reduced embryo viability [13]. Granulosa cells evolving in the presence of elevated concentrations of β-hydroxybutyrate (BHB) exhibit a transcriptomic profile associated with slow cell proliferation and altered fatty acid metabolism [14]. Embryos obtained from oocytes that have matured in a medium supplemented with insulin exhibit major structural and functional alterations [15].
Invited review: Advances and challenges in application of feedomics to improve dairy cow production and health
2019, Journal of Dairy ScienceCitation Excerpt :The changes of diet and rumen microbiome (composition, function, and metabolites) in transition dairy cows will directly alter the nutrients provided for other tissues and organs, which may affect the corresponding functions. Several studies have revealed changes in metabolites and inflammatory responses in the liver (McCarthy et al., 2010; McCabe et al., 2012), uterus (Wathes et al., 2009), polymorphonuclear leukocytes (Agrawal et al., 2017), and granulosa cells (Girard et al., 2015) at the transcriptomic level when transition dairy cows were under negative energy balance. Another study using transcriptomics confirmed that the major hepatic functional changes in transition cows were related to fatty acid oxidation, cholesterol metabolism, and gluconeogenesis (Ha et al., 2017).
Association between pre-breeding metabolic profiles and reproductive performance in heifers and lactating dairy cows
2019, TheriogenologyCitation Excerpt :Higher levels of BHBA have been associated with a longer VWP [15] and pregnancy at first artificial insemination [39]. Increased BHBA was associated with altered expression of a number of genes involved in proliferation and fatty acid metabolism in follicular granulosa cells [40]. Elsewhere, BHBA treatment reduced estradiol and progesterone synthesis in cultured granulosa cells [28].
Reduced serum vitamin D concentrations in healthy early-lactation dairy cattle
2018, Journal of Dairy ScienceCitation Excerpt :Vitamin D signaling in immune cells improved some indices of innate immunity and suppressed some proinflammatory measures of adaptive immunity in cattle (Nelson et al., 2010, 2012). Moreover, vitamin D improved disease resistance in models of chronic inflammatory-based diseases and reproductive disorders in cattle (Téllez-Pérez et al., 2012; Girard et al., 2015). Infusion of vitamin D into infected mammary gland quarters enhanced local immune function, decreased colonization by mastitis-causing bacteria, and reduced SCC in milk compared with untreated cows (Lippolis et al., 2011).
Embryo development in dairy cattle
2016, TheriogenologyCitation Excerpt :Third, exposure of oocytes in vitro to NEFA at physiological concentrations consistent with those measured in the preovulatory follicle of postpartum lactating cows is detrimental to oocyte development [30–33] and oviduct cell function [34] and subsequent embryo development [35]. Furthermore, Girard et al. [36] reported that negative energy balance altered gene expression in granulosa cells of dairy cows at 60 days postpartum. Consistent with these data, the metabolomic profile of follicular fluid (FF) from heifers and postpartum nonlactating and lactating cows has highlighted differences in the microenvironment in which the oocyte develops which may contribute to compromised oocyte quality [37–39].