scholarly journals Effect of sialylation and complexity of FSH oligosaccharides on inhibin production by granulosa cells

Reproduction ◽  
2013 ◽  
Vol 145 (2) ◽  
pp. 127-135 ◽  
Author(s):  
Nazareth Loreti ◽  
Verónica Ambao ◽  
Luz Andreone ◽  
Romina Trigo ◽  
Ursula Bussmann ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Concanavalin A ◽  
Inhibin B ◽  
Inhibin A ◽  
Biological Responses ◽  
Immature Rats ◽  
Fold Stimulation ◽  
Recombinant Human Fsh

Granulosa cell (GC) inhibin A and B production is regulated by FSH and gonadal factors. This gonadotrophin is released as a mixture of glycoforms, which induce different biological responses in vivo and in vitro. Our aim was to determine the effect of recombinant human FSH (rhFSH) glycosylation variants on inhibin A and B production by rat GCs. Preparative isoelectro focusing was used to isolate more acidic/sialylated (pH <4.00) and less acidic/sialylated (pH >5.00) rhFSH charge analogues. Concanavalin A was used to isolate unbound and firmly bound rhFSH glycoforms on the basis of their oligosaccharide complexity. GCs, obtained from oestrogen-primed immature rats, were cultured with either native rhFSH or its glycosylation variants. Inhibin A and B were determined using specific ELISAs. Results were expressed as mean±s.e.m. Under basal conditions, inhibin A was the predominant dimer produced (inhibin A: 673±55; inhibin B: 80±4 pg/ml). More acidic/sialylated charge analogues stimulated inhibin B production when compared to inhibin A at all doses studied; by contrast, less acidic/sialylated charge analogues stimulated inhibin A production and elicited no effect on inhibin B. Glycoforms bearing complex oligosaccharides showed a potent stimulatory effect on inhibin B when compared to inhibin A production (i.e. dose 1 ng/ml: 4.9±0.5 vs 0.9±0.1-fold stimulation, P<0.001). Glycoforms bearing hybrid-type oligosaccharides favoured inhibin A production (i.e. dose 4 ng/ml 2.9±0.1 vs 1.6±0.1-fold stimulation, P<0.05). These results show that the sialylation degree as well as the complexity of oligosaccharides present in the rhFSH molecule may be considered additional factors that differentially regulate dimeric inhibin production by rat GCs.

scholarly journals Inhibin B Is a More Potent Suppressor of Rat Follicle-Stimulating Hormone Release than Inhibin A in Vitro and in Vivo

Endocrinology ◽  
2009 ◽  
Vol 150 (10) ◽  
pp. 4784-4793 ◽  
Author(s):  
Yogeshwar Makanji ◽  
Peter D. Temple-Smith ◽  
Kelly L. Walton ◽  
Craig A. Harrison ◽  
David M. Robertson
Keyword(s):  
Follicle Stimulating Hormone ◽  
Inhibin B ◽  
Hormone Release ◽  
Inhibin A ◽  
Stimulating Hormone

Oestradiol synthesis by granulosa cells from immature rats treated with pregnant mare's serum gonadotrophin

1983 ◽  
Vol 104 (3) ◽  
pp. 372-380 ◽  
Author(s):  
Donald C. Johnson ◽  
Roger C. Hoversland
Keyword(s):  
Granulosa Cells ◽  
Steroid Synthesis ◽  
Bicarbonate Buffer ◽  
Immature Rats ◽  
Hypophysectomized Rats ◽  
System Inclusion ◽  
Ovulatory Follicles ◽  
Generating System

Abstract. Granulosa cells harvested from follicles in hypophysectomized or intact immature rats treated with 20 IU of pregnant mare's serum gonadotrophin (PMS) produced immunoreactive oestradiol (E2) when incubated in Krebs Ringer bicarbonate buffer containing an NADPH generating system; inclusion of steroid substrates in the medium increased the rate of synthesis. Further, tritiated E2 was synthesized when labelled progesterone was used as substrate. Granulosa cells removed from pre-ovulatory follicles on the morning of prooestrus in adult females also produced E2 in vitro. Although E2 synthesis was apparent by cells from immature hypophysectomized rats within 12 h of PMS treatment, it inceased greatly with longer in vivo exposure to the gonadotrophin. Production was linear with the number of cells incubated and with time, at least through the first 30 min; the production rate decreased slightly with longer incubations. Exposure of the cells in vivo to hCG or ovine LH, before incubation, destroyed most of their ability to synthesize E2 even if progesterone or pregnenolone was added to the medium, but conversion of testosterone to E2 was reduced by only about 50%. Inhibitors of steroid synthesis, i.e. 4-OH-androstenedione, SU-10603, cyanoketone, or aminoglutethimide, greatly reduced the amount of E2 synthesized by the cells. The results indicate that granulosa cells exposed in vivo to gonadotrophins can synthesize E2 without the addition of androgenic substrate provided that cofactors are supplied. This finding has important implications for the current 'two cell' theory for oestrogen production by the ovary. A deficiency in steroidogenic enzymes within the granulosa cell appears to be an inadequate basis for the theory. However, the total synthesis of E2 in vivo by granulosa cells has not been shown.

scholarly journals Activation of yeast polymerase II transcription by herpesvirus VP16 and GAL4 derivatives in vitro.

1989 ◽  
Vol 9 (11) ◽  
pp. 4746-4749 ◽  
Author(s):  
D I Chasman ◽  
J Leatherwood ◽  
M Carey ◽  
M Ptashne ◽  
R D Kornberg
Keyword(s):  
Rna Polymerase Ii ◽  
Fusion Proteins ◽  
In Vitro System ◽  
Transcription System ◽  
Natural Mechanism ◽  
Transcription In Vitro ◽  
Rna Polymerase Ii Transcription ◽  
Fold Stimulation

Fusion proteins known to activate transcription in vivo were tested for the ability to stimulate transcription in vitro in a recently developed Saccharomyces cerevisiae RNA polymerase II transcription system. One fusion protein, whose activation domain was derived from the herpesvirus transcriptional activator VP16, gave more than 100-fold stimulation in the in vitro system. The order of effects of the various proteins was the same for transcription in vitro and in vivo, suggesting that the natural mechanism of activation is preserved in vitro.

Nippostrongylus brasiliensis: further properties of antibody-damaged worms and induction of comparable damage by maintaining worms in vitro

Parasitology ◽  
1975 ◽  
Vol 71 (2) ◽  
pp. 275-283 ◽  
Author(s):  
R. J. Love ◽  
Bridget M. Ogilvie ◽  
Diane J. McLaren
Keyword(s):  
Immune Response ◽  
Isoenzyme Pattern ◽  
Nippostrongylus Brasiliensis ◽  
Ultrastructural Morphology ◽  
Immature Rats ◽  
Normal Rat ◽  
Worm Expulsion ◽  
Anterior Migration

When adult Nippostrongylus brasiliensis were maintained in vitro they became damaged. Using the criteria of ultrastructural morphology, acetylcholinesterase isoenzyme pattern and the behaviour of the worms after transfer to a normal rat, this damage appeared to be similar to that produced by the in vivo action of antibodies.Antibodies were shown to be responsible for the anterior migration of adult worms which occurs during primary infections in mature rats and in the prolonged infections seen in lactating and immature rats.Antibody damaged worms and worms unaffected by antibodies were equally able to stimulate the immune response required for worm expulsion. Apparently antibody damage is not required for the initiation of the second immune component necessary for expulsion of this parasite.

scholarly journals MiRNA-143 mediates the proliferative signaling pathway of FSH and regulates estradiol production

2017 ◽  
Vol 234 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Li Zhang ◽  
XiaoXin Zhang ◽  
Xuejing Zhang ◽  
Yu Lu ◽  
Lei Li ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Loss Of Function ◽  
Cellular Processes ◽  
Cell Functions ◽  
Genes Expression ◽  
Underlying Mechanisms

MicroRNAs (MiRNAs) play important regulatory roles in many cellular processes. MiR-143 is highly enriched in the mouse ovary, but its roles and underlying mechanisms are not well understood. In the current study, we show that miR-143 is located in granulosa cells of primary, secondary and antral follicles. To explore the specific functions of miR-143, we transfected miR-143 inhibitor into primary cultured granulosa cells to study the loss of function of miR-143 and the results showed that miR-143 silencing significantly increased estradiol production and steroidogenesis-related gene expression. Moreover, our in vivo and in vitro studies showed that follicular stimulating hormone (FSH) significantly decreased miR-143 expression. This function of miR-143 is accomplished by its binding to the 3’-UTR of KRAS mRNA. Furthermore, our results demonstrated that miR-143 acts as a negative regulating molecule mediating the signaling pathway of FSH and affecting estradiol production by targeting KRAS. MiR-143 also negatively acts in regulating granulosa cells proliferation and cell cycle-related genes expression. These findings indicate that miR-143 plays vital roles in FSH-induced estradiol production and granulosa cell proliferation, providing a novel mechanism that involves miRNA in regulating granulosa cell functions.

scholarly journals Investigating the Intrafollicular Factors Affecting Oocyte Developmental Competency

2021 ◽  
Author(s):  
◽  
Zaramasina Clark
Keyword(s):  
Gene Expression ◽  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Embryo Quality ◽  
Cumulus Cells ◽  
Significant Finding ◽  
High Quality ◽  
Final Maturation

<p>The number of cycles of assisted reproductive technologies (ART) performed increased by ~9.5 % globally between 2008 and 2010. In spite of this, the success rate in terms of delivery was only ~19.0 % (Dyer et al., 2016). This discrepancy between the demand for, and success of, these technologies necessitates the development of tools to improve ART efficiency. To facilitate this, a better understanding of how the microenvironment changes within the developing follicle to culminate in a mature, developmentally-competent oocyte is required. This study employed an in vivo and in vitro ovine model to investigate the relationship between the surrounding microenvironment and oocyte maturation, and in particular, the attainment of oocyte developmental competency and high-quality embryos.  The first objective of this PhD study was to comprehensively investigate the changing microenvironment of in vivo matured, presumptive preovulatory (PPOV) follicles from wild-type (++) and high ovulation rate (OR; I+B+) ewes. The high OR ewes were heterozygous carriers of mutations in BMP15 (I+) and BMPRIB (B+). Functional differences in follicular somatic (granulosa and cumulus) cells between these genotypes, including differential gonadotropin responsiveness of granulosa cells, composition of follicular fluid and gene expression profiles in cumulus cells were evident. These differences emerged as part of a compensatory mechanism by which oocytes from smaller follicles, containing fewer granulosa cells, achieved developmental competency in I+B+ ewes.  The second objective of this PhD study was to develop new approaches for improving current in vitro maturation (IVM) strategies. The first approach utilised in this study focused on developing biomarkers that could be used to improve prediction of developmental competency in oocytes and in vitro produced embryos. This involved interrogating the hypothesis that a combination of molecular and morphokinetic biomarkers would better predict the developmental competency of oocytes and embryos compared to using these biomarkers alone. The second approach utilised in this PhD study tested the effects of modulating IVM conditions to better mimic the follicular microenvironment of a high, compared to a low, OR species on oocyte developmental competency and embryo quality. This involved supplementing IVM media with different ratios of two oocyte-secreted growth factors, i.e. GDF9:BMP15, that were representative of low or high OR species. These approaches demonstrated significant potential and warrant further investigation.  The most significant finding of this study was that despite variances in the surrounding microenvironment during in vivo and in vitro oocyte maturation that culminated in differential gene expression patterns in cumulus cells, and divergent gonadotropin-responsiveness of granulosa cells, the gene expression signatures of developmentally-competent oocytes and the morphokinetics of high-quality embryos were unaltered. This confirms the value of developing such biomarkers for oocyte development competency and embryo quality that remain unaltered despite a changing surrounding environment. Interestingly, simulating the ratio of GDF9:BMP15 that oocytes from high OR species are exposed to during maturation improved developmental competency in oocytes as demonstrated by increased blastocyst rates. Furthermore, this study has demonstrated that combinations of molecular (cumulus cell gene expression) and morphokinetic biomarkers improved the ability to predict developmental competency in oocytes and embryos. Overall, this study revealed novel information regarding the follicular microenvironment during final maturation and identified several novel approaches to improving the efficiency of ART.</p>

scholarly journals Targeting of the Pilosebaceous Follicle by Liquid Crystal Nanocarriers: In Vitro and In Vivo Effects of the Entrapped Minoxidil

Pharmaceutics ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 1127
Author(s):  
Massimo Fresta ◽  
Antonia Mancuso ◽  
Maria Chiara Cristiano ◽  
Konrad Urbanek ◽  
Felisa Cilurzo ◽  
...  
Keyword(s):  
Liquid Crystal ◽  
Biological Effects ◽  
Biological Response ◽  
Response Duration ◽  
Topical Administration ◽  
Active Compounds ◽  
Biological Responses ◽  
Negative Surface

The topical administration of active compounds represents an advantageous strategy to reach the various skin components as well as its appendages. Pilosebaceous follicles are skin appendages originating in the deeper skin layers. They are very difficult to target, and hence higher active dosages are generally required to achieve effective biological responses, thus favoring the rise of side effects. The aim of this work was to design a supramolecular colloidal carrier, i.e., a liquid crystal nanocarrier, for the selective delivery of active compounds into the pilosebaceous follicle. This nanocarrier showed mean sizes of ~80 nm, a good stability, a negative surface charge, and great safety properties. In vitro studies highlighted its ability to contain and release different substances and to successfully permeate the skin. Minoxidil was encapsulated in the nanocarriers and the in vivo biological effect was compared with a conventional dosage form. Minoxidil-loaded liquid crystal nanocarrier was able to selectively reach the pilosebaceous follicle, thus allowing an increased biological effectiveness of the delivered active in terms of biological response, duration of the biological effects, and reduction of collaterals. Our investigation showed that liquid crystal nanocarriers represent a promising device for the treatment of different pilosebaceous follicular impairments/diseases.

scholarly journals Inhibition of oocyte growth factors in vivo modulates ovarian folliculogenesis in neonatal and immature mice

Reproduction ◽  
2010 ◽  
Vol 139 (3) ◽  
pp. 587-598 ◽  
Author(s):  
Samu Myllymaa ◽  
Arja Pasternack ◽  
David G Mottershead ◽  
Matti Poutanen ◽  
Minna M Pulkki ◽  
...  
Keyword(s):  
Growth Factors ◽  
Granulosa Cells ◽  
Microscopic Analysis ◽  
Corpora Lutea ◽  
Electron Microscopic ◽  
Oocyte Growth ◽  
Long Term Study ◽  
Ovarian Folliculogenesis

Growth differentiation factor-9 (GDF9) and bone morphogenetic protein-15 (BMP15) are among the key regulators transmitting the signaling between the oocyte and the surrounding granulosa cells. Previously, it has been shown that a recombinant BMP type II receptor ectodomain–Fc fusion protein (BMPR2ecd–Fc) is able to inhibit the actions of GDF9 and BMP15 in vitro. Here, we have produced bioactive BMPR2ecd–Fc, which was injected i.p. into neonatal mice. Early folliculogenesis was first studied by injecting mice five times with various doses of BMPR2ecd–Fc during the postnatal days 4–12. Folliculogenesis was affected dose dependently, as evidenced by a decreased mitogenesis of granulosa cells of the growing follicles. Furthermore, we also noticed a decrease in the number of secondary and tertiary follicles as well as an increase in the oocyte size. Electron microscopic analysis revealed that the ultrastructure of the granulosa cells of the primary follicles was not affected by the BMPR2ecd–Fc treatment. A second study was conducted to investigate whether a longer treatment with 12 injections during postnatal days 4–28 would inhibit folliculogenesis. Similar effects were observed in the two studies on the early follicular developmental stages. However, in the long-term study, later stages of folliculogenesis were not blocked but rather increased numbers of antral follicles, preovulatory follicles, and corpora lutea were found. We conclude that BMPR2ecd–Fc is a potent modulator of ovarian folliculogenesis in vivo, and thus, is a valuable tool for studying the physiology and downstream effects of oocyte-derived growth factors in vivo.

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