Perturbations in Lineage Specification of Granulosa and Theca Cells May Alter Corpus Luteum Formation and Function

Anovulation is a major cause of infertility, and it is the major leading reproductive disorder in mammalian females. Without ovulation, an oocyte is not released from the ovarian follicle to be fertilized and a corpus luteum is not formed. The corpus luteum formed from the luteinized somatic follicular cells following ovulation, vasculature cells, and immune cells is critical for progesterone production and maintenance of pregnancy. Follicular theca cells differentiate into small luteal cells (SLCs) that produce progesterone in response to luteinizing hormone (LH), and granulosa cells luteinize to become large luteal cells (LLCs) that have a high rate of basal production of progesterone. The formation and function of the corpus luteum rely on the appropriate proliferation and differentiation of both granulosa and theca cells. If any aspect of granulosa or theca cell luteinization is perturbed, then the resulting luteal cell populations (SLC, LLC, vascular, and immune cells) may be reduced and compromise progesterone production. Thus, many factors that affect the differentiation/lineage of the somatic cells and their gene expression profiles can alter the ability of a corpus luteum to produce the progesterone critical for pregnancy. Our laboratory has identified genes that are enriched in somatic follicular cells and luteal cells through gene expression microarray. This work was the first to compare the gene expression profiles of the four somatic cell types involved in the follicle-to-luteal transition and to support previous immunofluorescence data indicating theca cells differentiate into SLCs while granulosa cells become LLCs. Using these data and incorporating knowledge about the ways in which luteinization can go awry, we can extrapolate the impact that alterations in the theca and granulosa cell gene expression profiles and lineages could have on the formation and function of the corpus luteum. While interactions with other cell types such as vascular and immune cells are critical for appropriate corpus luteum function, we are restricting this review to focus on granulosa, theca, and luteal cells and how perturbations such as androgen excess and inflammation may affect their function and fertility

Livestock animals to study infertility in women

Anovulation is a major cause of infertility in mammalian females. Ovarian research in humans is challenging due to the shortage of “normal tissue” that can be obtained from women that are trying to get pregnant. Livestock animals provide valuable resources to investigate the optimal intra-follicular environment required to facilitate folliculogenesis and translate to humans. Different livestock models may be more useful depending on the specific biological process studied

TRIENNIAL REPRODUCTION SYMPOSIUM: Looking back and moving forward—how reproductive physiology has evolved

In honor of the 50th anniversary of the Society for the Study of Reproduction (SSR), a Triennial Reproduction Symposium was cosponsored by SSR and the American Society of Animal Science (ASAS). The Society for the Study of Reproduction was formed in 1967 when a group of reproductive biologists that were members of ASAS met with physician scientists and decided to organize a new scientific society. The goal of SSR was to promote the study of reproduction by fostering interdisciplinary communication among scientists, holding conferences, and publishing meritorious studies. Today after its 50th anniversary, the mission for SSR is to harness the science of reproduction, fertility, and development for a healthy world. The American Society of Animal Science began as the American Society of Animal Nutrition in 1908 and was broadened to include additional disciplines in 1912. The society changed its name to the American Society of Animal Production in 1915 and in 1961 the name of the society was changed to ASAS. Today, ASAS is a membership society that supports the careers of scientists and animal producers in the United States and internationally. The Society fosters the discovery, sharing, and application of scientific knowledge concerning the responsible use of animals to enhance human life and well-being

TRIENNIAL REPRODUCTION SYMPOSIUM: Looking back and moving forward—how reproductive physiology has evolved

In honor of the 50th anniversary of the Society for the Study of Reproduction (SSR), a Triennial Reproduction Symposium was cosponsored by SSR and the American Society of Animal Science (ASAS). The Society for the Study of Reproduction was formed in 1967 when a group of reproductive biologists that were members of ASAS met with physician scientists and decided to organize a new scientific society. The goal of SSR was to promote the study of reproduction by fostering interdisciplinary communication among scientists, holding conferences, and publishing meritorious studies. Today after its 50th anniversary, the mission for SSR is to harness the science of reproduction, fertility, and development for a healthy world. The American Society of Animal Science began as the American Society of Animal Nutrition in 1908 and was broadened to include additional disciplines in 1912. The society changed its name to the American Society of Animal Production in 1915 and in 1961 the name of the society was changed to ASAS. Today, ASAS is a membership society that supports the careers of scientists and animal producers in the United States and internationally. The Society fosters the discovery, sharing, and application of scientific knowledge concerning the responsible use of animals to enhance human life and well-being

Expression and Action of Transforming Growth Factor Beta (TGFβ1, TGFβ2, and TGFβ3) during Embryonic Rat Testis Development1

The objective of the current study was to determine the role of transforming growth factor beta (TGF) during seminiferous cord formation and embryonic testis development. The expression pattern of mRNA for TGF isoforms was evaluated during testis development through a quantitative reverse transcription-polymerase chain reaction (QRT-PCR) procedure. Expression of mRNA for TGF1 was highest at postnatal day 0 (P0) and P10. In contrast, TGF2 was high at embryonic day 15 (E15), declined at E16, and showed a transient increase at P0 through P3 of testis development. Interestingly, expression of mRNA for TGF3 was high during embryonic development and then declined after P3. Immunohistochemical localization of TGF1 and TGF2 demonstrated expression in Sertoli cells at E14 and in the seminiferous cords at P0. Selective interstitial cells expressed high concentrations of TGF1 and TGF2 in P0 testis. TGF3 was expressed in selective cells at the junction of the E14 testis and mesonephros. The cells expressing TGF3 in the testis appeared to be preperitubular cells that resided around the seminiferous cords. TGF3 was localized to gonocytes in P0 testis. TGF1 was found to have no influence on seminiferous cord formation in embryonic organ cultures of E13 testis. In contrast, growth of both E13 and E14 embryonic organ cultures was inhibited by TGF1 and resulted in reduced testis size (40% of controls) with fewer cords present. A P0 testis cell culture and thymidine incorporation assay were used to directly examine the effects of recombinant TGF1. TGF1 alone had no influence on thymidine incorporation in P0 testis cell cultures when compared to controls. Interestingly, TGF1 inhibited epi-dermal growth factor (EGF), and 10% calf serum stimulated P0 testis cell growth but not FSH-stimulated growth. Therefore, TGF1 appears to inhibit testis growth in both the embryonic and early postnatal periods. The hormonal regulation of TGF expression was measured using P0 testis cell cultures and a QRT-PCR procedure for each TGF isoform. High concentrations of EGF stimulated expression of mRNA for TGF1 after 24 h but suppressed expression of TGF3. In contrast, there was no effect of FSH on TGF isoform expression. In summary, TGF regulates embryonic and P0 testis growth through inhibiting the actions of positive growth factors such as EGF. In addition, EGF but not FSH appears to regulate TGF isoform expression. Combined observations from the present study demonstrate that TGF iso-forms are differentially expressed and appear to be regulators of testis growth during the embryonic and early postnatal periods

Extending Grazing In Heifer Development Systems Decreases Cost Without Compromising Production

Three experiments compared heifer development systems. In Exp. 1, 299 heifers (253 ± 2 kg) from 3 yr were used to compare dry lot (DL) to grazing corn residue (CR) post weaning. Heifers in the DL consumed a common diet after weaning for 187 d until breeding. The CR heifers grazed for 145 d with a supplement (0.45 kg/d; 28% CP) and were then fed in the DL until breeding. In Exp. 2, 270 heifers (225 ± 2 kg) in 3 yr grazed Sandhills winter range (WR) or CR with a supplement (0.45 kg/d; 28% CP) post weaning. In Exp. 3, 180 heifers (262 ± 3 kg) in 2 yr grazed Eastern Nebraska WR or CR with a supplement (0.45 – 0.90 kg/d; 29% CP) post weaning. The CR heifers had lower (P \u3c 0.001) ADG before breeding compared to DL or WR heifers in Exp. 1 and 2, but WR and CR were similar (P = 0.66) in Exp. 3. The DL and WR heifers were heavier (P \u3c 0.003) than CR at breeding and pregnancy diagnosis in Exp. 1 and 2, but similar (P = 0.62) in Exp. 3. The percentage of heifers pubertal at breeding was greater (P \u3c 0.001) for DL than CR in Exp. 1, for WR than CR in yr 1 and 2 of Exp. 2 (P \u3c 0.01), but similar (P = 0.36) in Exp. 3. Pregnancy rate to AI was lower (P = 0.08) for CR than DL heifers in Exp. 1, but not different (P = 0.89) in Exp. 3. Final pregnancy rate was not affected (P ≥ 0.27) in Exp. 1, 2 or 3. In Exp. 2,, yr 2, CR heifers required (P = 0.01) more calving assistance than WR. Milk production of WR heifers was greater (P = 0.04) than CR in Exp. 3. Calf weaning BW, two-year old AI (Exp. 1 and 3) and final pregnancy rates (Exp. 1, 2 and 3) were not different (P \u3e 0.10). Development grazing CR reduced cost by $45/pregnancy compared to DL, but cost of WR was similar to CR. Development grazing CR reduces ADG before breeding without sacrificing final pregnancy rate. Development grazing WR increases milk production, but does not increase weaning BW. Grazing CR during heifer development reduces cost compared to DL. Grazing CR or WR is suitable for heifer development at similar cost

Expression and Action of Neurotropin-3 and Nerve Growth Factor in Embryonic and Early Postnatal Rat Testis Development

The current study examines the expression and potential actions of neurotropin-3 (NT3), nerve growth factor (NGF), and their receptors during morphological sex determination (seminiferous cord formation) and perinatal rat testis development. The expression of neurotropins and their receptors was analyzed with immunohistochemistry. Cellular localization of neurotropin ligand and receptor proteins changed during embryonic testis development. Neurotropin-3 was localized to Sertoli cells at Embryonic Day 14 (E14), was present in gonocytes at Postnatal Day 0 (P0), and after birth became localized to the interstitium and Sertoli cells (P3–P5). The expression of trk C (the high affinity receptor for NT3) was localized to mesonephric ducts and cells surrounding the cords (E14–E18). In addition, Sertoli cells and preperitubular cells surrounding the cords at E14 also stained for trk C. Neurotropin-3 was expressed in gonocytes and Sertoli cells at P0–P5. Nerve growth factor was detected in Sertoli cells at E14, was clearly in Sertoli and interstitial cells at E16 and E18, and in Sertoli, germ, and interstitial cells from P0–P5. The expression of trk A (the high affinity receptor for NGF) was located in Sertoli and interstitial cells at E16–P5. To determine the actions of neurotropins during embryonic and perinatal testis development, experiments were conducted on E13 and P0 testis. Antisense oligonucleotide experiments with NT3 were used on E13 testis organ cultures to determine effects on seminiferous cord formation. Cord formation was inhibited in 40% of the organ cultures treated with the antisense NT3 oligonucleotides, while no inhibition was observed with sense oligonucleotides. In P0 testis cultures, both NT3 and NGF alone and in combination stimulated thymidine incorporation into DNA. Therefore, the neurotropins are involved in embryonic morphological events (cord formation; NT3) and in growth of the perinatal testis (P0; NT3 and NGF). To define further the growth effects of neurotropins on testis development, expression of transforming growth factor alpha and beta (TGFα and TGFβ) were examined in response to neurotropins. The P0 testis cultures were treated with neurotropins, and expression of mRNA for TGFα and TGFβ was analyzed utilizing a quantitative reverse transcription-polymerase chain reaction assay. Nerve growth factor and NT3 alone or in combination inhibited expression of mRNA for TGFα while NT3 increased mRNA expression of epidermal growth factor receptor. The combination treatment of neurotropins inhibited expression of TGFβ1 and increased expression of TGFβ3. In summary, observations suggest that NT3, NGF, trk A, and trk C are localized to cells critical to seminiferous cord formation and appear to be important regulators of morphological sex determination. In addition to these morphological effects, both NT3 and NGF stimulate P0 testis growth and may elicit their action through altering the expression of locally produced growth factors such as TGFα and TGFβ. Taken together these results suggest that neurotropins are regulators of paracrine cell-cell interactions that result in morphological sex determination and perinatal testis growth

Inhibition of Vascular Endothelial Growth Factor Manipulates Follicles in Beef Females

Vascular Endothelial Growth Factor (VEGF) is produced by cells surrounding the egg in the follicle. If VEGF is inhibited, ovulation does not occur. Understanding how VEGF regulates follicle development may allow for manipulation of estrous cycles. In previous studies in our laboratory, blocking the actions of VEGF decreased activation of early stage follicles in neonatal rat ovary cultures. Therefore, we hypothesized inhibition of VEGF actions would also inhibit follicle activation in bovine ovarian cortical cultures. Inhibition of VEGF did inhibit follicle progression, thus regulation of VEGF may be a way to manipulate follicle development and more accurately time ovulation

Inhibition of Vascular Endothelial Growth Factor Manipulates Follicles in Beef Females

Vascular Endothelial Growth Factor (VEGF) is produced by cells surrounding the egg in the follicle. If VEGF is inhibited, ovulation does not occur. Understanding how VEGF regulates follicle development may allow for manipulation of estrous cycles. In previous studies in our laboratory, blocking the actions of VEGF decreased activation of early stage follicles in neonatal rat ovary cultures. Therefore, we hypothesized inhibition of VEGF actions would also inhibit follicle activation in bovine ovarian cortical cultures. Inhibition of VEGF did inhibit follicle progression, thus regulation of VEGF may be a way to manipulate follicle development and more accurately time ovulation

Vascular Endothelial Growth Factor and Kinase Domain Region Receptor Are Involved in Both Seminiferous Cord Formation and Vascular Development During Testis Morphogenesis in the Rat

Morphological male sex determination is dependent on migration of endothelial and preperitubular cells from the adjacent mesonephros into the developing testis. Our hypothesis is that VEGFA and its receptor KDR are necessary for both testicular cord formation and neovascularization. The Vegfa gene has 8 exons with many splice variants. Vegfa120, Vegfa164, and Vegfa188 mRNA isoforms were detected on Embryonic Day (E) 13.5 (plug date = E0) in the rat. Vegfa120, Vegfa144, Vegfa164, Vegfa188, and Vegfa205 mRNA were detected at E18 and Postnatal Day 3 (P3). Kdr mRNA was present on E13.5, whereas Fms-like tyrosine kinase 1 receptor (Flt1) mRNA was not detected until E18. VEGFA protein was localized to Sertoli cells at cord formation and KDR to germ and interstitial cells. The VEGFA signaling inhibitors SU1498 (40 μM) and VEGFR-TKI (8 μM) inhibited cord formation in E13 testis cultures with 90% reduced vascular density (P \u3c 0.01) in VEGFR-TKI-treated organs. Furthermore, Je-11 (10 μM), an antagonist to VEGFA, also perturbed cord formation and inhibited vascular density by more than 50% (P \u3c 0.01). To determine signal transduction pathways involved in VEGFA’s regulation of testis morphogenesis, E13 testis were treated with LY 294002 (15 μM), a phosphoinositide 3-kinase (PI3K) pathway inhibitor, resulting in inhibition of both vascular density (46%) and cord formation. Thus, we support our hypothesis and conclude that VEGFA, secreted by the Sertoli cell, is involved in both neovascularization and cord formation and potentially acts through the PI3K pathway during testis morphogenesis to elicit its effects