Effect of lipopolysaccharide (LPS) on the production of progesterone (P4; a–c) and androstenedione (A4; d–f) in bovine
theca cells from post-selection follicles (>8.5 mm) during term 1 (white circles, 0–48 h) and term 2 (black triangles,
48–96 h). Theca cells were stimulated with 2.5 ng/ml luteinizing hormone (LH; a, d), 100 ng/ml estradiol (E2; b, e) or LH
and E2 (c, f). Data are expressed as the percentage of control (100%) steroid accumulation in the culture medium. All values
are means ± standard error of the mean of three independent experiments. Values with different letters (a, b) are
significantly different between groups (P < 0.05).

Effect of lipopolysaccharide (LPS) on the production of progesterone (P4; a–c) and androstenedione (A4; d–f) in bovine theca cells from post-selection follicles (>8.5 mm) during term 1 (white circles, 0–48 h) and term 2 (black triangles, 48–96 h). Theca cells were stimulated with 2.5 ng/ml luteinizing hormone (LH; a, d), 100 ng/ml estradiol (E2; b, e) or LH and E2 (c, f). Data are expressed as the percentage of control (100%) steroid accumulation in the culture medium. All values are means ± standard error of the mean of three independent experiments. Values with different letters (a, b) are significantly different between groups (P < 0.05).

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In postpartum dairy cows, lipopolysaccharide (LPS) derived from gram-negative bacteria such as Escherichia coli causes uterine inflammation and leads to ovarian dysfunction. The aim of this study was to determine the effect of LPS on steroid production in bovine theca cells at different stages of follicular development. Theca cells isolated from pr...

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... Moreover, luteinizing hormone (LH) secretion and pulsatility are also affected by LPS in cows and ewes [7][8][9]. LPS accumulates in ovarian follicular fluid and induces a local inflammatory response in the ovaries of animals affected by uterine or mammary gland infection [10][11][12]. As a result, inhibited follicular growth, reflected in disturbed ovarian cyclic activity, and lowered intrafollicular and circulating levels of estradiol (E2) are observed [3][4][5][6]. ...
... This is corroborated by both in vivo and in vitro observations demonstrating an LPS-dependent decrease in E2 production in bovine granulosa cells [13]. In LH-stimulated theca cells of rats, and in the bovine ovary, LPS inhibits progesterone and androstenedione synthesis [11,14]. Furthermore, administration of LPS in Endothelial Cell Growth Supplement (ECGS) (all from Chemie Brunschwig AG, Basel, Switzerland). ...
Article
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Escherichia coli (E. coli) is the most common Gram-negative bacterium causing infection of the uterus or mammary gland and is one of the major causes of infertility in livestock. In those animals affected by E. coli driven LPS-mediated infections, fertility problems occur in part due to disrupted follicular and luteal functionality. However, the molecular mechanisms by which LPS induces inflammation, and specifically, the role of LPS in the disruption of capillary morphogenesis and endothelial barrier function remain unclear. Here, we hypothesized that LPS may lead to alterations in luteal angiogenesis and vascular function by inducing inflammatory reactions in endothelial cells. Accordingly, OLENDO cells were treated with LPS followed by evaluation of the expression of selected representative proinflammatory cytokines: NF-kB, IL6, IL8, TNFα, and ICAM 1. While TNFα was not affected by treatment with LPS, transcripts of NF-kB, IL6, and IL8 were affected in a dosage-dependent manner. Additionally, the activity of TLR2 and TLR4 was blocked, resulting in suppression of the LPS-induced expression of ICAM 1, NF-kB, IL6, and IL8. Inhibition of the PKA or MAPK/ERK pathways suppressed the LPS-stimulated expression of NF-kB, IL6, and IL8, whereas blocking the PKC pathway had the opposite effect. Furthermore, LPS-induced phosphorylation of Erk1 and Erk2 was inhibited when the TLR4 or MAPK/ERK pathways were blocked. Finally, LPS seems to induce inflammatory processes in OLENDO cells via TLR2 and TLR4, utilizing different signaling pathways.
... Escherichia coli and Trueperella pyogenes (TP) are the most widely known bacteria that cause postpartum endometritis [3,39,44]. Colonization of the upper reproductive tract by E. coli is reportedly associated with severe damage to the endometrium and disruption of ovarian cycle activity, followed by infertility [20,30,38,45]. In the early stages of intrauterine infection, E. coli is often the infectious agent, but the source of infection then shifts to bacteria such as TP, with persistent infection leading to chronic uterine disease [12]. ...
... In the early stages of intrauterine infection, E. coli is often the infectious agent, but the source of infection then shifts to bacteria such as TP, with persistent infection leading to chronic uterine disease [12]. Bacterial lipopolysaccharide (LPS) acts on ovarian luteinizing hormone-stimulated theca cells to inhibit steroid production [30]. The presence of E. coli and LPS in the lochia during the early postpartum period favors the development of uterine infections by TP and gram-negative anaerobes during the later postpartum period [12]. ...
Article
Although Escherichia coli is a commensal bacterium of the bovine vaginal microbiota, it is an important pathogenic bacterium that causes diseases of the reproductive tract and sub-fertility. Recent studies have focused on virulence factors (VFs) of intrauterine E. coli; however, actual endometrial VFs have not been clearly identified. The purpose of this study was to identify the VFs of E. coli associated with clinical metritis and endometritis. Thirty-two strains of E. coli and four mixed Trueperella pyogenes (TP) strains were detected in the uterus of 19 Holstein dairy cows with obvious clinical signs (between 8 and 66 days postpartum). The presence of six E. coli VFs (fimH, fyuA, kpsMTII, hra1, csgA, and astA) was examined by PCR, and clinical signs and reproductive performance (mixed TP, the percentage of polymorphonuclear neutrophils [PMN%], days to uterine involution, etc.) were evaluated. Four VFs (fimH, hra1, csgA, and astA) were detected in all E. coli strains, whereas fyuA and kpsMTII were detected in 94% and 50% of strains, respectively. Cows with E. coli strains harboring kpsMTII exhibited significantly severe clinical scores (vaginal discharge score, PMN%, uterine involution), suggesting that kpsMTII is a key VF for progression of clinical metritis and endometritis. In the present study, we clearly identified six VFs associated with clinical metritis and endometritis. In addition, E. coli strains with kpsMTII probably play a crucial role in the progression of clinical metritis and endometritis.
... Also, LPS inhibited estradiol production in A Iran. Biomed. J. 26 (1): 44-52 45 granulosa cells and progesterone production in theca cells, representing an endocrine-disrupting effect [1,4,5] . This endotoxin disrupted meiotic progression, mitochondrial distribution in the cytoplasm, and mitochondrial membrane potential, which caused the disruption of nuclear maturation of bovine oocytes [6] . ...
Article
Background: A mouse model of LPS-induced inflammation was used to investigate the effect of pharmacological inhibition of nuclear enzyme PARP-1 on oocyte maturation, apoptotic and necrotic death, as well as DNA integrity of follicular cells. Also, the relative expression of cumulus genes (HAS2, COX2, and GREM1) associated with oocyte developmental competence was assessed. Methods: Mice were treated with the PARP-1 inhibitor, 4-HQN, one hour before LPS administration. After 24 h, oocyte in vitro maturation was detected. Granulosa cell DNA damage was determined by the alkaline comet assay. Live, necrotic and apoptotic cells were identified using double vital staining by fluorescent dyes, Hoechst 33342 and propidium iodide. The expression levels of cumulus genes were assessed using reverse transcriptase PCR. Results: The administration of 4-HQN to LPS-treated mice ameliorated oocyte meiotic maturation and exerted a significant cytoprotective effect. 4-HQN attenuated LPS-induced DNA damage and favored cell survival by decreasing necrosis and apoptosis in granulosa cells. Exposure to 4-HQN increased mRNA expression levels for HAS2, COX2, and GREM1 in cumulus cells. Conclusion: The obtained results indicate the involvement of PARP-1 in the pathogenesis of ovarian dysfunction caused by LPS. We suppose that this enzyme can be an attractive target for the therapy of inflammatory disorders in ovary. The protective action of PARP-1 inhibition could at least partly be associated with the reduction of necrotic death of follicular cells and also in other cells. However, the detailed mechanisms of the favorable effect of PARP inhibitors on endotoxin-induced ovarian disorders need to be further explored.
... The etiology of endometritis is very complex; it is mostly caused by fungal, bacterial, and parasitic infections (46,47). LPSs are one of the important causes of endometritis in dairy cows; they can induce an inflammatory response by stimulating bEECs to secrete inflammatory cytokines (48). Therefore, we chose LPSs to establish an in vitro bovine endometritis model to study the anti-inflammatory effect of MSCs. ...
Article
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Endometritis is a disease that affects reproductive health in dairy cows and causes serious economic damage to the dairy industry world-wide. Although in recent years, the application of mesenchymal stem cell (MSC) therapy for the treatment of inflammatory diseases has attracted much attention, there are few reports of the use of MSCs in dairy cows. In the present study, our objective was to explore the inhibitory effects of bovine adipose-derived mesenchymal stem cells (bAD-MSCs) on lipopolysaccharide (LPS) induced inflammation in bovine endometrial epithelial cells (bEECs) along with the potential underlying molecular mechanisms. We characterized isolated bAD-MSCs using cell surface marker staining and adipogenic/osteogenic differentiation, and analyzed them using immunofluorescence, flow cytometry (surface marker staining), and adipogenic and osteogenic differentiation. Furthermore, to understand the anti-inflammatory effects of bAD-MSCs on LPS induced bEEC inflammation, we used a bAD-MSC/bEEC co-culture system. The results showed that bAD-MSC treatments could significantly decrease LPS induced bEEC apoptosis and pro-inflammatory cytokine expression levels, such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α). Furthermore, our results showed that bAD-MSC treatments could also significantly downregulate LPS induced p38, IkB-a, and JAK1 phosphorylation and Bax protein expression levels, which are closely related to inflammatory progress and cellular apoptosis in bEECs. Our findings demonstrate that bAD-MSCs play an inhibitory role in LPS induced bEEC inflammation and provide new insights for the clinical therapy of endometritis in dairy cows.
... Theca cells functions are modulated by steroid hormones, growth factors, and bone morphogenetic proteins (BMP) (Young and McNeilly, 2010). Additionally, they are highly sensitive to a variety of stimuli, including nutrition (Williams et al., 2001), stress , heat stress (Nteeba et al., 2015), and bacterial insult (Magata et al., 2014). Increasing evidence also suggests that endocrine disrupting chemicals (EDCs) can disrupt ovarian function (Patel et al., 2015;Craig and Ziv-Gal, 2018). ...
Article
Gap junction intercellular communication (GJIC) is necessary for ovarian function, and it is temporospatially regulated during follicular development and ovulation. At outermost layer of the antral follicle, theca cells provide structural, steroidogenic, and vascular support. Inter- and extra-thecal GJIC is required for intrafollicular trafficking of signaling molecules. Because GJIC can be altered by hormones and endocrine disrupting chemicals (EDCs), we tested if any of five common EDCs (bisphenol A (BPA), bisphenol S (BPS), bisphenol F (BPF), perfluorooctanesulfonic acid (PFOS), and triphenyltin chloride (TPT)) can interfere with theca cell GJIC. Since most chemicals are reported to repress GJIC, we hypothesized that all chemicals tested, within environmentally relevant human exposure concentrations, will inhibit theca cell GJICs. To evaluate this hypothesis, we used a scrape loading/dye transfer assay. BPS, but no other chemical tested, enhanced GJIC in a dose- and time-dependent manner in ovine primary theca cells. A signal-protein inhibitor approach was used to explore the GJIC-modulatory pathways involved. Phospholipase C and mitogen-activated protein kinase (MAPK) inhibitors significantly attenuated BPS-induced enhanced GJIC. Human theca cells were used to evaluate translational relevance of these findings. Human primary theca cells had a ∼40% increase in GJIC in response to BPS, which was attenuated with a MAPK inhibitor, suggestive of a conserved mechanism. Upregulation of GJIC could result in hyperplasia of the theca cell layer or prevent ovulation by holding the oocyte in meiotic arrest. Further studies are necessary to understand in vitro to in vivo translatability of these findings on follicle development and fertility outcomes.
... Although several studies investigated the effects of LPS on the reproductive performance of dairy cows (Bromfield and Sheldon, 2013;Magata et al., 2014;Zhao et al., 2019), the response of ovine oocytes to increasing concentrations of LPS is not well defined yet. Ewes do not experience negative energy balance (NEB) and metabolic stress after parturition as much as dairy cows, however severe feed shortage and high numbers of suckling lambs can predispose milking ewes to NEB, weaken the immune system and consequently their susceptibility towards infectious diseases such as metritis and mastitis. ...
Article
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Although a considerable number of studies have investigated the effects of lipopolysaccharide (LPS) on the reproductive performance of dairy cows, the response of ovine oocytes to LPS during their in vitro maturation and development is not well defined yet. Ewe’s ovaries were obtained from a slaughterhouse, the oocytes were collected and matured in the presence of increasing concentrations (0, 0.01, 0.1, 1 and 10 µg/mL) of LPS in order to evaluate the meiotic maturation by measuring the proportion of oocytes reaching the MII stage. The concentration of intracellular glutathione (GSH) was measured in oocytes following maturation in vitro. In addition, concentrations of selected metabolites including glucose, pyruvate, lactate and glutamine were quantified in the medium following maturation. A number of treated matured oocytes along with the control group were subsequently fertilized using frozen semen and assessed for the rate of cleavage and for the proportion reaching the blastocyst stage. The number of oocytes in MII stage was significantly reduced in response to the increasing concentrations of LPS (77.83%, 70.64%, 68.86%, 66.32%, respectively, in case of 0.01, 0.1, 1 and 10 µg/mL LPS when compared to the control group, 76.34%; P<0.05). There were no differences neither in the intracellular concentration of GSH in the oocytes nor in case of the metabolites in the maturation medium. Although the rate of cleaved oocytes was not affected by increasing levels of LPS, the blastocyst rate was reduced in a dose dependent manner (36.69%, 34.21%, 30.35%, 17.27% and 14.03% for the control, 0.01, 0.1, 1 and 10 µg/mL LPS, respectively (P<0.05). These results demonstrate that the developmental competence of ovine oocytes may be affected detrimentally by LPS and such deleterious effects could be related to the maturation process.
... Escherichia coli is one of the most relevant bacteria implicated in the establishment of postpartum uterine diseases; also increases the uterus susceptibility to subsequent infections by Arcanobacterium pyogenes and other bacterial species [12][13][14][15]. It has been reported that E. coli colonization of the upper reproductive tract is associated with severe damage to the endometrium and disruption of the ovarian cyclic activity followed by infertility [16][17][18][19]. Moreover, E. coli lipopolysaccharide (LPS) disrupts endocrine function by decreasing peripheral plasma concentrations of estradiol and progesterone, as well as suppressing GnRH and LH secretions by the hypothalamus and pituitary gland, respectively [18]. ...
... Although to date there is no etiological agent associated with the RB syndrome; taking into consideration that E. coli is capable of altering the endocrine function of the BRT by disturbing ovulation, as well as the embryo viability and that cows with subclinical endometritis have high serum concentrations of LPS 40 to 60 days after parturition [13,18,19], it is likely that this microorganism may participate in the pathogenesis of the RB syndrome. Complementary studies should be done to evaluate the implication of E. coli in this syndrome. ...
Article
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Even though Escherichia coli are common bacteria of the bovine vaginal microbiota, they represent an important pathogen that causes diseases in the reproductive tract and subfertility. However, the actual endometrial virulence profile of E. coli is poorly understood. The present study aims to characterize the phylogenetic structure and virulence potential of native vaginal populations of E. coli from healthy heifers (H), and cows with postpartum uterine diseases (PUD), such as metritis/endometritis (MT) or repeat breeder cows (RB). To this end, the virulence repertoire of 97 E. coli isolates was genotypically and phenotypically assessed. Most of them were assigned to phylogenetic group A (74%), followed by B1 (17%) and D (9%); RB strains were significantly (p < 0.05) more represented by B1. Seven of the 15 evaluated virulence genes (VFG) were detected and the most prevalent were fimH (87%), agn43 (41%) and csgA (35%); while traT (27%), fyuA (11%), hlyA (5%) and kpsMT II (5%) were observed in a lower proportion. Particularly, fyuA was significantly higher (p < 0.05) in MT cows whereas csgA showed the same behavior in PUD animals (p < 0.05). When comparing H and PUD strains, these last ones were associated to positive expression of biofilm, fimbriae curli/cellulose and motility; yet RB strains did not show motility. Vaginal B1 E. coli populations, that possess VFG (fyuA and csgA) as well as the expression of motility, curli fimbriae/cellulose and biofilm, may represent risk factors for endometrial disorders; specifically, those that also, have kpsMT II may have a pathogenic potential for causing the RB syndrome. Future research focusing on the detection of these strains in the vaginal microbiota of cows with postpartum uterine diseases should be done since the control of their presence in vagina could reduce the risk that they access the uterus during the postpartum period.
... Binding of LPS to this receptor complex results in the nuclear translocation of nuclear factor kappa-B components, which leads to the production of pro-inflammatory cytokines and chemokines [23,37]. In bovine follicles, granulosa cells [31,38] and theca cells [39] express TLR4, CD14, and MD2, suggesting that follicular cells can respond to LPS. Indeed, bovine ovarian granulosa cells can initiate an inflammatory response to LPS via the TLR4 pathway. ...
... Moreover, treatment of theca cells with LPS impaired P4 and A4 production and reduced the mRNA expression of steroidogenesis-related genes (e.g. StAR and CYP17) [39]. ...
Article
Full-text available
Uterine inflammatory diseases commonly occur in postpartum dairy cows, resulting in reduced reproductive performance due to aberrant uterine and ovarian activity. Infection of the uterus with gram-negative bacteria results in the detection of lipopolysaccharide (LPS) in the plasma and follicular fluid of cows along with uterine inflammation. LPS acts on follicular components such as theca cells, granulosa cells, and follicle-enclosed oocytes, leading to impaired follicular activity. Follicles with a high LPS environment exhibit reduced follicular steroidogenesis due to the inhibition of steroidogenic enzyme transcription. Primary cell cultures of bovine granulosa and theca cells have shown that LPS acts on follicular cells to impair steroid production, which may disturb follicle growth and/or reduce their ability to ovulate. Even if ovulation occurs, cows with uterine inflammation are less likely to conceive because in addition to uterine damage, LPS also impairs the developmental competence of oocytes. LPS perturbs the nuclear and cytoplasmic maturation of bovine oocytes. Moreover, oocytes matured in an LPS treatment are less likely to develop into the blastocyst stage. Such oocytes also have a reduced number of trophoblast cells in blastocysts. Therefore, the detrimental effects of LPS on ovarian activity may be partly responsible for infertility in cows with uterine inflammation. Novel treatment and prevention strategies for uterine inflammatory diseases can be developed by advancing our knowledge of the pathophysiology underlying ovarian dysfunction, and this can only be achieved by further research. The present review outlines the molecular pathogenesis of LPS-induced ovarian dysfunction. Graphical Abstract Fullsize Image
... LPS supressed bovine granulosa cell production of oestradiol , while progesterone production did not change in response to LPS. In contrast, LH-stimulated or luteinising bovine theca cells responded to LPS challenge with reduced progesterone production (Magata et al. 2014, Shimizu et al. 2016, suggesting a greater sensitivity of theca cells to LPS during luteinisation. Conversely, LPS stimulated progesterone production by bovine luteal cells in vitro, although there was no additive effect of LH and LPS (Grant et al. 2007). ...
... LPS has been shown to inhibit neural progenitor cell proliferation (Ekdahl et al. 2003) and to suppress critical mediators of cell cycle progression (Cohn et al. 2010). The number of viable bovine luteal cells was reduced in response to short-term LPS treatment (Grant et al. 2007), while in contrast, LPS did not alter the survival of developing or luteinising bovine theca or granulosa cells over 96 h of culture , Magata et al. 2014. ...
Article
Full-text available
Reproductive tract inflammatory disease (RTID) commonly occurs after the traumatic events of parturition and adversely affects follicular function. This study is the first to describe the cellular and steroidogenic characteristics of corpora lutea from cattle with RTID and the effects of pathogen-associated molecular patterns (PAMPs) on luteal angiogenesis and function in vitro. Luteal weight (P<0.05) and progesterone content (P<0.05) were reduced (1.2-fold) in cows with RTID, accompanied by reduced CYP11A (P<0.05), HSD3B (P<0.01) and STAR (P<0.01) protein expression. Immunohistochemistry revealed that luteal vascularity (VWF) and pericyte (ACTA2) coverage were >3-fold lower in RTID cows (P<0.05). To link these observations to bacterial infection and determine specificity of action, a physiologically-relevant luteal angiogenesis culture system examined the effects of PAMPs on endothelial cell (EC) network formation and progesterone production, in the presence of pro-angiogenic factors. Luteal EC networks were reduced ≤95% (P<0.05) by lipopolysaccharide (LPS, toll-like receptor (TLR) 4 agonist) but not by TLR2 agonists lipoteichoic acid or peptidoglycan. Conversely, progesterone production and steroidogenic protein expression were unaffected by PAMPs (P>0.05). Moreover, the adverse effect of LPS on luteal EC networks was dose-dependent and effective from 1ng/ml (P<0.05), while few EC networks were present above 10ng/ml LPS (P<0.001). LPS reduced proliferation (P<0.05) and increased apoptosis of EC (P<0.001). The specific TLR4 inhibitor TAK242 reversed the effects of LPS on EC networks. In conclusion, luteal vasculature is adversely sensitive to LPS acting via TLR4, therefore ovarian exposure to LPS from any Gram-negative bacterial infection will profoundly influence subsequent reproductive potential.
... The expression of Toll-like receptors (TLR) by granulosa and theca cells makes them responsive to LPS and other bacterial products [31][32][33]. The activation of TLRs on granulosa and theca cells disturbs the communication between oocyte and cumulus cells that is needed for the development of competent oocyte [34]. ...
... Besides, the likely reason behind delayed ovulation is reduced estradiol production which is due to the stimulation of TLR receptors on theca cells by inflammatory products and mediators [31,33,43]. The dropped amount of estrogen in circulation results in dim and delayed preovulatory LH surge which subsequently causes delayed ovulation. ...
... The dropped amount of estrogen in circulation results in dim and delayed preovulatory LH surge which subsequently causes delayed ovulation. In their study, Magata, et al. [33] cultured bovine theca cells in vitro in a media containing peptidoglycan and/or LPS to investigate the effects of these bacterial toxins on steroidogenesis. Theca cells presented TLR receptors and nucleotide-binding oligomerization domain 1 and 2. The bacterial toxins caused decreased prostaglandin and androstenedione production along with decreased mRNA expression of steroidogenic enzymes. ...
Article
Full-text available
According to the international dairy federation, the dairy sector provides up to one billion livelihoods around the globe. Albeit, it faces many challenges including the two potentially threatening diseases which are mastitis and infertility. Both are complex multifactorial diseases, and mastitis is associated with causing infertility. Mastitis is an intramammary infection (IMI) and it occurs in two forms, clinical and subclinical. Mastitis being the infection of mammary glands, directly affects milk production to reduce its quality and quantity and indirectly hinders the reproductive performance of dairy animals. It negatively affects the reproduction parameters of dairy animals including, an increase in days open, a decrease in pregnancy rates, and increases chances of early embryonic losses and abortion. The chronic cases of mastitis lead toward the infertility of dairy animals and both of these diseases are responsible for increasing the culling rate and decreasing the profitability of a dairy farm. The objective of this study is to illustrate the influence of subclinical mastitis on reproduction parameters of dairy animals including, days open, days to the first service after calving, pregnancy rates, abortion, and embryonic losses. The relation between mastitis and infertility will be explained mechanistically.