Assessment of cGMP level in medium during in vitro growth period of murine preantral follicles with and without supplementation of C-type natriuretic peptide

Zygote ◽  
2021 ◽  
pp. 1-5
Author(s):  
Li Ang ◽  
Guo Xingping ◽  
Cao Haixia ◽  
Wang Zhulin ◽  
Wang Huaixiu
Keyword(s):  
Natriuretic Peptide ◽  
Ovarian Follicle ◽  
Ovarian Follicles ◽  
Growth Patterns ◽  
Guanosine Monophosphate ◽  
Cgmp Level ◽  
Control Group ◽  
Oocyte Maturity ◽  
Experimental Group

Summary To enhance the developmental competency of murine ovarian follicles cultured in vitro, C-type natriuretic peptide (CNP) was supplemented in the culture system. Although the mechanism is not fully elucidated, it was reported that the effect of CNP supplementation was mediated by increased cyclic guanosine monophosphate (cGMP). In the present study, cGMP levels in media for murine preantral follicle culture were compared both between a control group without CNP supplementation and an experimental group with CNP supplementation and between days in each group. In addition, follicle growth patterns and oocyte maturity were assessed and compared between the two groups. Results demonstrated that along with in vitro culture, cGMP levels increased (P < 0.05) both in the control group and the experimental group, whereas cGMP levels were not significantly different between the two groups on the same day of in vitro culture (P > 0.05). The oocyte’s maturity was superior in the experimental group compared with the control group (P < 0.05). As ovarian follicles grew three-dimensionally in the experimental group but were flattened in the control group, CNP might improve oocyte maturity through maintaining the three-dimensional architecture of the ovarian follicle because of increased transzonal projections (TZP) and functional gap junctions between oocyte and surrounding granulosa cells.

Supplementation of c-type natriuretic peptide during in vitro growth period benefits the development of murine preantral follicles

Zygote ◽  
2020 ◽  
pp. 1-5
Author(s):  
Li Ang ◽  
Cao Haixia ◽  
Li Hongxia ◽  
Li Ruijiao ◽  
Guo Xingping ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Granulosa Cells ◽  
Culture System ◽  
Early Stage ◽  
Developmental Competence ◽  
Control Group ◽  
Follicle Development ◽  
Control Groups ◽  
Preantral Follicles

Summary The present study investigated the effects of c-type natriuretic peptide (CNP) on the development of murine preantral follicles during in vitro growth (IVG). Preantral follicles isolated from ovaries of Kunming mice were cultured in vitro. In the culture system, CNP was supplemented in the experimental groups and omitted in the control groups. In Experiment 1, CNP was only supplemented at the early stage and follicle development was evaluated. In Experiments 2 and 3, CNP was supplemented during the whole period of in vitro culture. In Experiment 2, follicle development and oocyte maturity were evaluated. In Experiment 3, follicle development and embryo cleavage after in vitro fertilization (IVF) were assessed. The results showed that in the control groups in all three experiments, granulosa cells migrated from within the follicle and the follicles could not reach the antral stage. In the experimental groups in all three experiments, no migration of granulosa cells was observed and follicle development was assessed as attaining the antral stage, which was significantly superior to that of the control group (P < 0.0001). Oocyte meiotic arrest was effectively maintained, hence giving good developmental competence. In conclusion, CNP supplementation in the culture system during IVG benefited the development of murine preantral follicles.

scholarly journals Signalling pathways and mechanistic cues highlighted by transcriptomic analysis of primordial, primary, and secondary ovarian follicles in domestic cat

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shauna Kehoe ◽  
Katarina Jewgenow ◽  
Paul R. Johnston ◽  
Susan Mbedi ◽  
Beate C. Braun
Keyword(s):  
Gene Expression ◽  
Transforming Growth Factor ◽  
Ovarian Follicle ◽  
Mammalian Species ◽  
Expression Patterns ◽  
Ovarian Follicles ◽  
Domestic Cat ◽  
Transcriptional Analysis ◽  
Ovarian Folliculogenesis

AbstractIn vitro growth (IVG) of dormant primordial ovarian follicles aims to produce mature competent oocytes for assisted reproduction. Success is dependent on optimal in vitro conditions complemented with an understanding of oocyte and ovarian follicle development in vivo. Complete IVG has not been achieved in any other mammalian species besides mice. Furthermore, ovarian folliculogenesis remains sparsely understood overall. Here, gene expression patterns were characterised by RNA-sequencing in primordial (PrF), primary (PF), and secondary (SF) ovarian follicles from Felis catus (domestic cat) ovaries. Two major transitions were investigated: PrF-PF and PF-SF. Transcriptional analysis revealed a higher proportion in gene expression changes during the PrF-PF transition. Key influencing factors during this transition included the interaction between the extracellular matrix (ECM) and matrix metalloproteinase (MMPs) along with nuclear components such as, histone HIST1H1T (H1.6). Conserved signalling factors and expression patterns previously described during mammalian ovarian folliculogenesis were observed. Species-specific features during domestic cat ovarian folliculogenesis were also found. The signalling pathway terms “PI3K-Akt”, “transforming growth factor-β receptor”, “ErbB”, and “HIF-1” from the functional annotation analysis were studied. Some results highlighted mechanistic cues potentially involved in PrF development in the domestic cat. Overall, this study provides an insight into regulatory factors and pathways during preantral ovarian folliculogenesis in domestic cat.

scholarly journals 5-Aminolevulinic Acid-Based Photodynamic Therapy Pretreatment Mitigates Ultraviolet A-Induced Oxidative Photodamage

2018 ◽  
Vol 2018 ◽  
pp. 1-11 ◽  
Author(s):  
Hui Hua ◽  
Jiawei Cheng ◽  
Wenbo Bu ◽  
Juan Liu ◽  
Weiwei Ma ◽  
...  
Keyword(s):  
Aminolevulinic Acid ◽  
Epidermal Keratinocytes ◽  
Control Group ◽  
Hacat Cells ◽  
Ultraviolet A ◽  
Hairless Mice ◽  
Human Epidermal Keratinocytes ◽  
Experimental Group

Aim. To determine whether 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT) is effective in combating ultraviolet A- (UVA-) induced oxidative photodamage of hairless mice skin in vivo and human epidermal keratinocytes in vitro. Methods. In in vitro experiments, the human keratinocyte cell line (HaCaT cells) was divided into two groups: the experimental group was treated with ALA-PDT and the control group was left untreated. Then, the experimental group and the control group of cells were exposed to 10 J/m2 of UVA radiation. ROS, O2− species, and MMP were determined by fluorescence microscopy; p53, OGG1, and XPC were determined by Western blot analysis; apoptosis was determined by flow cytometry; and 8-oxo-dG was determined by immunofluorescence. Moreover, HaCaT cells were also treated with ALA-PDT. Then, SOD1 and SOD2 were examined by Western blot analysis. In in vivo experiments, the dorsal skin of hairless mice was treated with ALA-PDT or saline-PDT, and then, they were exposed to 20 J/m2 UVA light. The compound 8-oxo-dG was detected by immunofluorescence. Conclusion. In human epidermal keratinocytes and hairless mice skin, UVA-induced oxidative damage can be prevented effectively with ALA-PDT pretreatment.

scholarly journals Piglets produced by transfer of embryos obtained by in vitro fertilization of oocytes matured in vitro with thymosin: A case report

2020 ◽  
Vol 76 (03) ◽  
pp. 6356-2020 ◽  
Author(s):  
KATARZYNA PONIEDZIAŁEK-KEMPNY ◽  
BARBARA GAJDA ◽  
IWONA RAJSKA ◽  
LECHOSŁAW GAJDA ◽  
ZDZISŁAW SMORĄG
Keyword(s):  
Case Report ◽  
In Vitro Fertilization ◽  
Control Group ◽  
First Birth ◽  
Vitro Fertilization ◽  
Research Material ◽  
Preliminary Study ◽  
Experimental Group

The aim of the study was to examine the in vivo viability of in vitro-produced (IVP) porcine embryos obtained from oocytes matured with thymosin. The research material for this study consisted of immature pig oocytes obtained from ovaries after slaughter and ejaculated semen obtained from one boar. The immature oocytes were cultured in vitro until the metaphase II stage in a medium supplemented with thymosin (TMS). The presumptive zygotes obtained were cultured in vitro for 4-40 hours. The presumptive zygotes and 2-4-cell embryos were evaluated in vivo after transferring them to synchronized recipients. After the transfer of embryos from the experimental group into 2 recipients (50 embryos into each gilt) and the transfer of 50 embryos from the control group into 1 recipient, both gilts that had received embryos obtained by in vitro fertilization of oocytes matured with TMS became pregnant and delivered a total of 16 live piglets. After the transfer of embryos from the control group, no pregnancy was achieved. In conclusion, the results of our preliminary study suggest that the maturation of pig oocytes with thymosin supports the in vivo survival of in vitro produced embryos. It is important to note, that this was the first birth of piglets obtained after transfer of IVP embryos in Poland.

scholarly journals 81 EFFECT OF SUCROSE CONCENTRATION IN WARMING MEDIUM ON THE DEVELOPMENT POTENTIAL OF VITRIFIED BOVINE OOCYTES

2005 ◽  
Vol 17 (2) ◽  
pp. 190
Author(s):  
W.C. Chang ◽  
J. Xu ◽  
S. Jiang ◽  
X.C. Tian ◽  
X. Yang ◽  
...  
Keyword(s):  
Sucrose Concentration ◽  
Control Group ◽  
Bovine Oocyte ◽  
Cell Stage ◽  
Significant Difference ◽  
Humidified Air ◽  
Bovine Oocytes ◽  
Experimental Group ◽  
One Way Anova

The aim of this experiment was to determine the effect of the sucrose concentration (0 to 0.33 M) in the dilution medium on the viability, fertilizability, and development of vitrified bovine oocytes. Bovine oocyte-cumulus complexes were collected from slaughterhouse ovaries and in vitro-matured as reported previously. After 24-h maturation in TCM199-based medium under 5% CO2 humidified air at 39°C, these were exposed to hyaluronidase and carefully pipetted to remove all except the 3–5 innermost layers of cumulus. Oocytes were put into the pre-equilibration medium for 3 min and then into vitrification solution containing HEPES-buffered TCM199 supplemented with 20% FBS, ethylene glycol, and dimethylsulphoxide for 25–30 s; they were then vitrified by modified solid surface vitrification (Dinnyes et al. 2000 Biol. Reprod. 63, 513–518).The oocytes were warmed at 39°C by placing them in holding medium with 0, 0.08, 0.17, 0.25, or 0.33 M sucrose. Non-vitrified oocytes were used as controls. Oocytes were inseminated 30 min after warming, and the presumptive zygotes were cultured in CR1-aa medium supplemented with 6 mg/mL BSA at 39°C in a humidified atmosphere of 5% CO2, 5% O2, and 90% N2 for eight days. Data were analyzed by one-way ANOVA. As shown in Table 1, there was no significant difference in survival rate (P > 0.05) of the vitrified oocytes that were placed in dilution solution containing 0.17, 0.25, or 0.33 M sucrose and the non-treated controls. On Day 2 (fertilized on Day 0), cleavage to the 8-cell stage was similar for the 0.17, 0.25, and 0.33 M dilution groups, but the rates for all three were significantly lower (P < 0.05) than for the control group. The blastocyst rate on Day 8 was significantly higher for the 0.25 M group than for any other experimental group but still significantly lower than for the control. In conclusion, this study suggests that with this vitrification/warming procedure the optimum concentration of sucrose in the dilution solution is 0.25 M. Table 1. Oocyte survival after vitrification/warming and subsequent embryo development The authors would like to thank Ms Colleen Shaffer for the preparation of bovine oocytes.

scholarly journals Serum deprivation affects secretory activity of cultured porcine ovarian follicles and granulosa cells and their response to hormones

2009 ◽  
Vol 54 (No. 10) ◽  
pp. 455-460
Author(s):  
A.V. Sirotkin
Keyword(s):  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Secretory Activity ◽  
Ovarian Follicles ◽  
Serum Deprivation ◽  
Ovarian Cells ◽  
Igf I ◽  
Granulose Cells ◽  
Vitro Model

The aim of the present study is to understand the hormonal mechanisms of the effect of malnutrition on ovarian follicle functions. For this purpose, we examined the effect of malnutrition/serum deprivation, addition of metabolic hormones and gonadotropin (IGF-I, leptin and FSH) and their combination on the release of progesterone (P<sub>4</sub>), testosterone (T), estradiol (E<sub>2</sub>) and insulin-like growth factor I (IGF-I) by cultured whole ovarian follicles and on P<sub>4</sub> and IGF-I output by cultured granulosa cells isolated from porcine ovaries. It was observed that in ovarian follicles cultured with nutrients/serum addition of IGF-I reduced release of P<sub>4</sub>, but not of T or E<sub>2</sub>. Exogenous leptin reduced output of E<sub>2</sub>, but not of P<sub>4</sub> or T, and increased IGF-I output. No significant effect of FSH on release of steroid hormones by isolated follicles was found. Serum deprivation did not affect release of P<sub>4</sub>, but reduced output of T and E<sub>2</sub>, and promoted IGF-I release by cultured ovarian follicles. Addition of hormones failed to prevent the effect of malnutrition on the secretory activity of cultured ovarian follicles. In cultured granulose cells, all the tested hormones promoted release of both P<sub>4</sub> and IGF-I. Food restriction/serum deprivation reduced both P<sub>4</sub> and IGF-I output. Additions of either IGF-I, leptin and FSH prevented the inhibitory action of malnutrition on both P<sub>4</sub> and IGF-I release. The present observations (1) confirm the involvement of the hormones IGF-I, leptin and FSH in the control of secretory activity of ovarian cells, (2) demonstrate, that both isolated ovarian granulosa cells and whole follicles cultured in the absence of serum nutrients could be an adequate in-vitro model for studying the effect of malnutrition on ovarian secretory functions, and (3) suggest, that malnutrition could affect ovarian functions through changes in the release of ovarian hormones.

scholarly journals In vitro ovarian follicle growth: a comprehensive analysis of key protocol variables†

2020 ◽  
Vol 103 (3) ◽  
pp. 455-470
Author(s):  
Leah E Simon ◽  
T Rajendra Kumar ◽  
Francesca E Duncan
Keyword(s):  
Wildlife Conservation ◽  
Culture Media ◽  
Ovarian Follicle ◽  
Three Dimensional ◽  
Ovarian Follicles ◽  
Culture Methods ◽  
Growth And Survival ◽  
Follicle Culture ◽  
Culture Techniques

Abstract Folliculogenesis is a complex process that requires integration of autocrine, paracrine, and endocrine factors together with tightly regulated interactions between granulosa cells and oocytes for the growth and survival of healthy follicles. Culture of ovarian follicles is a powerful approach for investigating folliculogenesis and oogenesis in a tightly controlled environment. This method has not only enabled unprecedented insight into the fundamental biology of follicle development but also has far-reaching translational applications, including in fertility preservation for women whose ovarian follicles may be damaged by disease or its treatment or in wildlife conservation. Two- and three-dimensional follicle culture systems have been developed and are rapidly evolving. It is clear from a review of the literature on isolated follicle culture methods published over the past two decades (1980–2018) that protocols vary with respect to species examined, follicle isolation methods, culture techniques, culture media and nutrient and hormone supplementation, and experimental endpoints. Here we review the heterogeneity among these major variables of follicle culture protocols.

scholarly journals Experimental Application of Sage in Rabbit Husbandry

2008 ◽  
Vol 77 (4) ◽  
pp. 581-588 ◽  
Author(s):  
R. Szabóová ◽  
A. Lauková ◽  
Ľ. Chrastinová ◽  
M. Simonová ◽  
V. Strompfová ◽  
...  
Keyword(s):  
Inhibitory Effect ◽  
Negative Influence ◽  
Feed Consumption ◽  
Control Group ◽  
Coagulase Negative Staphylococci ◽  
E Coli ◽  
Commercial Feed ◽  
Experimental Group

Salvia spp. belongs to the Labiatae family and is characterized by antimicrobial and antiinflammatory effect. The aim of this study was to test its in vitro and in vivo inhibitory effect against bacteria as well as to find an alternative possibility to use sage in the rabbit ecosystem examining biochemical, zootechnical and inmunological indicators, compared to the commercial feed mixture Xtract. Using the sage extract in in vitro tests, its inhibitory effect was noted. Under in vivo conditions, in the experimental group with sage (EG1), reduction of Pseudomonas-like sp. (p < 0.01) and E. coli (p < 0.01) was noted after 7 days of sage application compared to the control group CG2 (with Robenidin) as well as after 21 days of sage extract application, when the reduction of coagulase-negative staphylococci (p < 0.01) was detected (in comparison with the experimental group-EG2, Xtract group). In the caecum of rabbits from EG1, higher values of lactic, acetic and butyric acids were noted. The values of propionic acid were not influenced. Biochemical indicators were not influenced; however, the values of GSH Px were lower in EG1 compared to EG2. Higher phagocytic activity (18%) was noted in EG1 than in EG2 (13%) after 21 days of additives application. The reduction of Eimeria sp. oocysts was demonstrated in EG1 (sage group) after 7 days of sage application comparing to CG2 (217 OPG to 566 OPG). The animals in both experimental groups achieved higher feed consumption and weight gain, lower mortality compared to both controls. Neither of the additives had a negative influence on the health status and growth performance of rabbits.

scholarly journals Supplementation of c-type natriuretic peptide during the whole in vitro growth period benefits the development of murine secondary follicles

2020 ◽  
Author(s):  
Ang Li ◽  
Haixia Cao ◽  
Hongxia Li ◽  
Ruijiao Li ◽  
Huaixiu Wang ◽  
...  
Keyword(s):  
Natriuretic Peptide ◽  
Ovulation Induction ◽  
Germinal Vesicle ◽  
Antral Follicle ◽  
Control Group ◽  
Follicle Development ◽  
In Vitro Growth ◽  
Germinal Vesicle Breakdown ◽  
Preovulatory Follicles

Abstract Background Supplementation of c-type natriuretic peptide (CNP) in the culture medium shortly before in vitro maturation (IVM) has been reported to be effective in delaying meiotic resumption of murine oocyte. The present study investigated the effect of CNP supplementation during the whole period of in vitro growth (IVG) on the development of murine secondary ovarian follicles.Methods Late secondary ovarian follicles isolated from ovaries of Kunming mice were cultured in vitro with and without supplementation of CNP. In experiment 1, CNP was supplemented at the early stage and the follicle development was evaluated. In experiment 2 and 3, CNP was supplemented during the whole period of IVG. In experiment 2, follicle development and oocyte maturity were evaluated. In group 3, follicle development and rate of cleaved embryos after in vitro fertilization (IVF) was assessed.Results In control group in all 3 experiments, granulosa cells migrated from within follicle and adhered to the plate at different degrees. The follicles flattened and could not reach antral stage. About 39.8% (39/98) of the oocytes ovulated nakedly. As no antral follicle was obtained, IVF was not performed in control group in experiment 3. In experiment group in all 3 experiments, no migration of guanulosa cells was observed and the follicles grew three-dimensionally. Ovulation of naked oocyte decreased substantially. The rate of antral stage follicle were 45% (18/40) in experiment 1. This parameter was 75.9% (44/58) in experiment 2 and 3 combined. In experiment 2, in preovulatory follicles without ovulation induction, oocytes at germinal vesicle (GV) stage and germinal vesicle breakdown (GVBD) stage were 87.5% (14/16) and 12.5% (2/16), respectively. In preovulatory follicles with ovulation induction, no GV stage oocyte was retrieved, oocytes at GVBD and metaphase II (MII) stage were 50% (8/16), respectively. In experiment 3, among 18 follicles cultured, 12 cumulus-oocyte complexes (COC) ovulated automatically after ovulation induction. Eleven oocytes were fertilized and cleaved. Compared with control groups, the follicle development assessed by naked oocyte ovulation and follicle stage (preantral follicle and antral follicle) in experiment groups were significantly superior (p<0.0001). CNP effectively maintained oocytes’ meiotic arrest and enhanced fertilization competency.Conclusions The supplementation of CNP in culture system of murine late secondary follicle during the whole period of IVG could sustain the 3-dimensional structure of follicle, increase the antral formation rate. As a result, the oocyte’s competency to be fertilized was greatly improved.

scholarly journals Cortical Piezo-Puncture as a Minimally Invasive Method for Reducing MiniScrew Implant Insertion Torque: A Preliminary in vitro Study

2018 ◽  
Vol 8 ◽  
pp. 124-130
Author(s):  
Juan Pablo Gomez ◽  
Lisbetd Giraldo ◽  
Daniela Montoya ◽  
Miguel Urrea
Keyword(s):  
In Vitro Study ◽  
Control Group ◽  
Insertion Torque ◽  
Miniscrew Implants ◽  
Mini Implants ◽  
Intact Bone ◽  
Invasive Method ◽  
Miniscrew Implant ◽  
Experimental Group

Objective The objective of this study was to determine the effect of cortical piezo-puncture (CPP) on maximum insertion torque (MIT), maximum removal torque (MRT), and maximum axial load (MAL) during the insertion of self-drilling miniscrew implants (MSI), in an experimental model with proximal epiphysis of bovine tibia. Materials and Methods A comparative study was conducted using two groups of 20 self-drilling MSI inserted in intact bone (control group) and in bone with previous CPP (experimental group). MIT, MRT, and MAL of the 20 mini implants of each group were measured. Using SPSS software, Student’s t-test was applied to compare MIT and MRT and the U-test Mann–Whitney test was applied to compare MAL in both groups as well as Pearson and Spearman correlation. Results In the experimental group, average values of 12.85 (±4,32) Newton x centimeters (Ncm), 13.7 (±4,54) Ncm, and 22,474 (±895,95) gF for MIT, MRT, and MAL were found, respectively. In the control group, average values found for MIT, MRT, and MAL were 20.2 (±4,7) Ncm, 22.3 (±5,17) Ncm, and 4688,7 (±320,18) gF, respectively. Statistically significant differences were observed in MIT, MRT, and MAL between control and experimental groups (P < 0,001). Conclusions CPP before insertion of orthodontic MSI in bovine tibia significantly reduces MIT, MRT, and MAL.

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