a-tissue-culture-model-estrogen-producing-primary-bovine-granulosa

A Tissue Culture Model of Estrogen-producing Primary Bovine Granulosa Cells

Ovarian granulosa cells (GC) are the major source of estradiol synthesis. Induced by the preovulatory luteinizing hormone (LH) surge, cells of the theca ...

a-modified-co-culture-system-for-understanding-granulosa-theca-cell

A Modified Co-Culture System for Understanding Granulosa-Theca Cell Interactions in the Bovine Ovary

A reliable co-culture of granulosa (GCs) and theca cells (TCs) has the potential to facilitate a more comprehensive understanding of the interactions, ...

Leibniz Institute for Farm Animal Biology (FBN)

Research Institute for Farm Animal Biology (FBN)

Cultured ovarian granulosa cells are essential models to study molecular mechanisms of gene regulation during folliculogenesis. Here, we characterize primary tissue culture models for bovine granulosa cells by morphological and physiological parameters and by novel molecular luteinization markers, as transcript abundance and DNA methylation levels. The data show that: (1) collagen substrate increased the number of attached, viable cells; (2) the expression of the key transcripts of estrogen synthesis, CYP19A1, could be induced and maintained in granulosa cells from small to medium but not from large follicles, whereas (3) only granulosa cells from large but not from smaller follicles were responsive to LH; (4) serum supplementation unfavorably transformed the cellular phenotype, induced proliferation and PCNA expression, reduced or abolished the transcript abundance of steroidogenic key genes and of gonadotropin receptor genes, CYP11A1, CYP19A1, FSHR and LHCGR but, however, did not increase the abundance of the luteinization-specific marker transcripts PTGS2, PTX3, RGS2 and VNN2; but (5) by increasing the plating density, estradiol production and the abundance of CYP19A1 transcripts, in particular those derived from the main ovarian promoter P2, were decreased concurrently leaving P2-specific DNA methylation levels unchanged, whereas progesterone secretion was stimulated and the expression of both luteinization-specific marker transcripts, RGS2 and VNN2, was significantly induced. From these data, we conclude that increasing the plating density induces a different, partly complementary, physiological and gene expression profile in cultured bovine granulosa cells and drives the cells towards an early post-LH stage of luteinization, even in the absence of luteinizing agents.

Increasing cell plating density mimics an early post-LH stage in cultured bovine granulosa cells.

The LH-induced folliculo-luteal transformation is connected with alterations of the gene expression profile in cells of the granulosa layer. It has been described that hypoxic conditions occur during luteinization, thus favoring the formation of L-lactate within the follicle. Despite being a product of anaerobic respiration, L-lactate has been shown to act as a signaling molecule affecting gene expression in neuronal cells. During the present study, we tested the hypothesis that L-lactate may influence differentiation of follicular granulosa cells (GC).

Lactate promotes specific differentiation in bovine granulosa cells depending on lactate uptake thus mimicking an early post-LH stage.

Previously, we could show that L-lactate affects cultured bovine granulosa cells (GC) in a specific manner driving the cells into an early pre-ovulatory phenotype. Here we studied genome wide effects in L-lactate-treated GC to further elucidate the underlying mechanisms that are responsible for the L-lactate induced transformation. Cultured estrogen producing GC treated either with L-lactate or vehicle control were subjected to mRNA microarray analysis.

L-lactate induces specific genome wide alterations of gene expression in cultured bovine granulosa cells.

L-lactate acts as a signaling molecule in bovine granulosa cells (GCs). The initiated alterations depend on the transport of L-lactate into the cells via monocarboxylate transporters. In the present study, we further elucidated the intracellular actions of L-lactate and tested whether the PKA signaling pathway is involved. Therefore, we treated cultured bovine GCs with L-lactate and PKA inhibitors H-89 and KT5720, and with an activator of PKA, 6-Bnz-cAMP. L-lactate treatment resulted in decreased estradiol production and downregulation of CYP19A1, FSHR, and LHCGR as well as in the upregulation of the markers of early luteinization PTX3, RGS2, and VNN2. These specific L-lactate effects were almost completely abolished by pre-treatment of the GCs with both inhibitors of PKA signaling. In addition, also the L-lactate-induced upregulation of LDHA and of the monocarboxylate transporters SLC16A1 and SLC16A7 was abolished after PKA inhibition. An activation of the PKA with 6-Bnz-cAMP revealed similar effects on the gene expression like L-lactate alone. In summary, the presented data demonstrate that L-lactate-induced effects on GCs are mediated via PKA signaling thus supporting the role of L-lactate as signaling molecule during the folliculo-luteal transition.

Lactate-induced effects on bovine granulosa cells are mediated via PKA signaling.

Pollution due to microplastics and nanoplastics is one of the major environmental issues of the last decade and represents a growing threat to human and animal health. In aquatic species, there is a large amount of information regarding the perturbation of marine organisms; instead, there are only a few studies focusing on the pathophysiological consequences of an acute and chronic exposure to micro- and nanoplastics in mammalian systems, especially on the reproductive system. There are several studies that have described the damage caused by plastic particles, including oxidative stress, apoptosis, inflammatory response, dysregulation of the endocrine system and accumulation in various organs. In addition to this, microplastics have recently been found to influence the evolution of microbial communities and increase the gene exchange, including antibiotic and metal resistance genes. Special attention must be paid to farm animals, because they produce food such as milk, eggs and meat, with the consequent risk of biological amplification along the food chain. The results of several studies indicate that there is an accumulation of microplastics and nanoplastics in human and animal tissues, with several negative effects, but all the effects in the body have not been ascertained, especially considering the long-term consequences. This review provides an overview of the possible adverse effects of the exposure of livestock to micro- and nanoplastics and assesses the potential risks for the disruption of reproductive physiological functions.

Impact of Microplastics and Nanoplastics on Livestock Health: An Emerging Risk for Reproductive Efficiency.

Nutrition has not only an impact on the general wellbeing of an animal but can also affect reproductive processes. In cattle, feeding regimes can influence the age of puberty onset and alter gonadal development. We analyzed effects of different milk replacer (MR) feeding regimes during rearing on ovarian physiology with specific emphasis on the numbers as well as gene expression characteristics of granulosa cells (GCs) at the age of puberty onset. Two groups of calves received either 10% or 20% of bodyweight MR per day during their first 8 weeks. After weaning, both groups were fed the same mixed ration ad libitum until slaughter at 8 months.

Early milk-feeding regimes in calves exert long-term effects on the development of ovarian granulosa cells.

Plastic pollution is a global diffuse threat, especially considering its fragmentation into microplastics (MPs) and nanoplastics (NPs). Since the contamination of the aquatic environment is already well studied, most studies have now focused on the soil. Moreover, the number of studies on the exposure routes and toxic effects of MNPs in humans is continuously increasing. Although MNPs can cause inflammation, cytotoxicity, genotoxicity and immune toxicity in livestock animals, which can accumulate ingested/inhaled plastic particles and transfer them to humans through the food chain, research on this topic is still lacking. In considering farm animals as the missing link between soil/plant contamination and human health effects, this paper aims to describe their importance as carriers and vectors of MNP contamination. As research on this topic is in its early stages, there is no standard method to quantify the amount and the characteristics of MNPs in different matrices. Therefore, the creation of a common database where researchers can report data on MNP characteristics and quantification methods could be helpful for both method standardization and the future training of an AI tool for predicting the most abundant/dangerous polymer(s), thus supporting policy decisions to reduce plastic pollution and perfectly fitting with One Health principles.

Connecting the Dots: Livestock Animals as Missing Links in the Chain of Microplastic Contamination and Human Health.

The harmful effects of micro- and nanoplastics (MNPs) on the aquatic ecosystem are already well established, and several studies have demonstrated that MNPs can contaminate the soil. However, the impact of MNPs on farm animals, whose products are intended for human consumption, as well as the accumulation and translocation of these particles in their bodies, is less investigated and not well understood. To address this issue, we evaluated the cellular uptake and the effects of three different concentrations (5, 25, and 75 μg/mL) of 100 nm polystyrene nanoplastics (PS-NPs) on ovarian bovine granulosa cells (GCs) and porcine myoblasts derived from skeletal muscle satellite cells as in vitro primary cell culture models. The uptake of PS-NPs was shown for all the concentrations tested, both for GCs and for myoblasts. The results for GCs reported a significant decrease in cell viability (P < 0.05) for all concentrations of nanoplastics tested compared to the control. However, steroid hormone production and the mRNA expression of GC physiology marker genes were not affected. The results for myoblasts showed a significant decrease in the mean confluence (P < 0.05) after exposure to a concentration of 75 μg/mL of nanoplastics compared to the control. This may be indicative of an initial inhibition of muscle fibre formation. However, cell viability, proliferative capacity, and the mRNA expression of myogenesis-associated genes were not affected. As there is currently no standard method for assessing the quantity of particles that overcome the anatomical barriers and accumulate in various parts of the body, recognizing the implications of exposure to MNPs in farm animals can help us to better comprehend the potential risks to human health. This knowledge is critical for developing informed treatment and avoidance strategies, ensuring the safety of both the food we consume and the environment in which it is produced.

ArticleTotal : 2	Year
Publication title Cited by 23	2018
Publication title	2025

Article	Year
Increasing cell plating density mimics an early post-LH stage in cultured bovine granulosa cells. Cell and tissue research\| PubMed ID: 24026437	2013
Lactate promotes specific differentiation in bovine granulosa cells depending on lactate uptake thus mimicking an early post-LH stage. Reproductive biology and endocrinology : RB&E\| PubMed ID: 29463248	2018
L-lactate induces specific genome wide alterations of gene expression in cultured bovine granulosa cells. BMC genomics\| PubMed ID: 30953450	2019
Lactate-induced effects on bovine granulosa cells are mediated via PKA signaling. Cell and tissue research\| PubMed ID: 34985545	2022
Impact of Microplastics and Nanoplastics on Livestock Health: An Emerging Risk for Reproductive Efficiency. Animals : an open access journal from MDPI\| PubMed ID: 37048387	2023
Early milk-feeding regimes in calves exert long-term effects on the development of ovarian granulosa cells. BMC genomics\| PubMed ID: 37626314	2023
Connecting the Dots: Livestock Animals as Missing Links in the Chain of Microplastic Contamination and Human Health. Animals : an open access journal from MDPI\| PubMed ID: 38275809	2024
Exploring the influence of polystyrene-nanoplastics on two distinct in vitro systems in farm animals: A pilot study. The Science of the total environment\| PubMed ID: 40209587	2025

Anja Baufeld

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