Choice of protocol for the in vivo bioassay of equine Chorionic Gonadotropin (eCG / PMSG) in immature female rats

Combarnous, Yves; Mariot, J.; Relav, Lauriane; Nguyen, Thi M.‐D.; Klett, Danièle

doi:10.1016/j.theriogenology.2019.03.004

Cited by 6 publications

(6 citation statements)

References 25 publications

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“…Owing to its long half-life in blood, a single dose of eCG, as opposed to multiple doses, is adequate to stimulate ovarian gene expression [13]. Furthermore, eCG and human CG (hCG) together stimulate ovulation in rats and mice [14,15].…”

Section: Introductionmentioning

confidence: 99%

“…Non-functional oocytes in N-eCG and R-eCG mutants were approximately 20 and 2%, respectively. Numerous studies have reported the effects of a combination of eCG and hCG on reproductive performance and estrous synchronization [13][14][15][16]. However, no studies have investigated the effects of N-eCG and R-eCG on gene regulation through RNA-based microarray analysis.…”

Section: Introductionmentioning

confidence: 99%

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Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

et al. 2020

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Background Equine chorionic gonadotropin (eCG) induces super-ovulation in laboratory animals. Notwithstanding its extensive usage, limited information is available regarding the differences between the in vivo effects of natural eCG (N-eCG) and recombinant eCG (R-eCG). This study aimed to investigate the gene expression profiles of mouse ovaries upon stimulation with N-eCG and R-eCG produced from CHO-suspension (CHO-S) cells. R-eCG gene was constructed and transfected into CHO-S cells and quantified. Subsequently, we determined the metabolic clearance rate (MCR) of N-eCG and R-eCG up to 24 h after intravenous administration through the mice tail vein and identified differentially expressed genes in both ovarian tissues, via quantitative real-time PCR (qRT-PCR) and immunohistochemistry (IHC). Results R-eCG was markedly expressed initially after transfection and maintained until recovery on day 9. Glycan chains were substantially modified in R-eCG protein produced from CHO-S cells and eliminated through PNGase F treatment. The MCR was higher for R-eCG than for N-eCG, and no significant difference was observed after 60 min. Notwithstanding their low concentrations, R-eCG and N-eCG were detected in the blood at 24 h post-injection. Microarray analysis of ovarian tissue revealed that 20 of 12,816 genes assessed therein were significantly up-regulated and 43 genes were down-regulated by > 2-fold in the group that received R-eCG (63 [0.49%] differentially regulated genes in total). The microarray results were concurrent with and hence validated by those of RT-PCR, qRT-PCR, and IHC analyses. Conclusions The present results indicate that R-eCG can be adequately produced through a cell-based expression system through post-translational modification of eCG and can induce ovulation in vivo. These results provide novel insights into the molecular mechanisms underlying the up- or down-regulation of specific ovarian genes and the production of R-eCG with enhanced biological activity in vivo.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

et al. 2020

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show abstract

“…Non-functional oocytes with the N-eCG and R-eCG mutants have been determined approximately 20% and 2%, respectively. Numerous studies have reported the effects of a combination of eCG and hCG on reproductive performance and estrous synchronization [13][14][15][16]. However, no studies have investigated the effects of N-eCG and R-eCG on gene regulation through RNA-based microarray analysis.…”

mentioning

confidence: 99%

Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

Min

Park

Lee

et al. 2020

Preprint

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Background: Equine chorionic gonadotropin (eCG) induces super-ovulation in laboratory animals. Notwithstanding its extensive usage, limited information is available regarding the differences between the in vivo effects of natural eCG (N-eCG) and recombinant eCG (R-eCG). This study aimed to investigate the gene expression profiles of mouse ovaries upon stimulation with N-eCG and R-eCG. R-eCG gene was cloned and transfected into CHO-suspension (CHO-S) cells and quantified. Thereafter, we determined the metabolic clearance rate (MCR) of N-eCG and R-eCG up to 24 h after intravenous administration through the tail vein and identified differentially expressed genes in both ovarian tissues, via quantitative real-time PCR (qRT-PCR) and immunohistochemistry (IHC). Results: R-eCG was markedly expressed initially after transfection and maintained until recovery on day 9. Oligosaccharide glycans were eliminated approximately 10 kDa through PNGase F treatment in R-eCG protein. The MCR was slightly lower for R-eCG than for N-eCG, and no significant difference was observed after 60 min. Notwithstanding their low concentration, R-eCG and N-eCG were detected in the blood at 24h post-injection. Microarray analysis of ovarian tissue revealed that 20 of 12,816 genes assessed therein were significantly up-regulated and 43 genes were down-regulated by >2-fold in the group receiving R-eCG (63 [0.49%] differentially regulated genes in total). The microarray results were concurrent with and hence validated by those of RT-PCR, qRT-PCR, and IHC analyses. Conclusions: The present results indicate that R-eCG can be adequately produced through a cell-based expression system through post-translational modification of eCG and can induce ovulation in vivo . These results provide novel insights into the molecular mechanisms underlying the up- or down-regulation of specific ovarian genes and the production of R-eCG with enhanced biological activity in vivo.

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“…Approximately 20% of non-functional oocytes with native eCG and only 2% with the rec-eCGs have been determined thus far. Numerous studies have reported the effects of a combination of eCG and hCG on reproductive performance and estrous synchronization [13][14][15][16]. However, no studies have investigated the effects of native eCG and rec-eCG on gene regulation through RNA-based microarray analysis.…”

mentioning

confidence: 99%

Comparative gene expression profiling of mouse ovaries upon stimulation with native equine chorionic gonadotropin (eCG) and tethered recombinant-eCG

Min

Park

Lee

et al. 2020

Preprint

View full text Add to dashboard Cite

Background: Equine chorionic gonadotropin (eCG) induces super-ovulation in laboratory animals. Notwithstanding its extensive usage, limited information is available regarding the differences between the in vivo effects of native eCG and recombinant eCG (rec-eCG). This study aimed to investigate the gene expression profiles of mouse ovaries upon stimulation with native eCG and rec-eCG produced from CHO-suspension (CHO-S) cells. eCG and rec-eCG were cloned and transfected into CHO-S cells and quantified. Thereafter, we determined the metabolic clearance rate (MCR) of native eCG and rec-eCG up to 24 h after intravenous administration through the tail vein and identified differentially expressed genes in both ovarian tissues, via quantitative real-time PCR (qRT-PCR) and immunohistochemistry (IHC).Results: Rec-eCG was markedly up-regulated initially after transfection and maintained until recovery on day 9. Oligosaccharide chains were substantially modified in rec-eCG protein produced from CHO-S cells and eliminated through PNGase F treatment. The MCR was slightly lower for rec-eCG than for eCG, and no significant difference was observed after 60 min. Notwithstanding their low concentration, rec-eCG and native eCG were detected in the blood at 24h post-injection. Microarray analysis of ovarian tissue revealed that 20 of 12,816 genes assessed therein were significantly up-regulated and 43 genes were down-regulated by >2-fold in the group receiving rec-eCG (63 [0.49%] differentially regulated genes in total). The microarray results were concurrent with and hence validated by those of RT-PCR, qRT-PCR, and IHC analyses.Conclusions: The present results indicate that rec-eCG can be adequately produced through a cell-based expression system through post-translational modification of eCG and can induce ovulation in vivo. These results provide novel insights into the molecular mechanisms underlying the up- or down-regulation of specific ovarian genes and the production of rec-eCG with enhanced biological activity in vivo.

show abstract

Choice of protocol for the in vivo bioassay of equine Chorionic Gonadotropin (eCG / PMSG) in immature female rats

Cited by 6 publications

References 25 publications

Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

Comparative gene expression profiling of mouse ovaries upon stimulation with natural equine chorionic gonadotropin (N-eCG) and tethered recombinant-eCG (R-eCG)

Comparative gene expression profiling of mouse ovaries upon stimulation with native equine chorionic gonadotropin (eCG) and tethered recombinant-eCG

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