Dataset Information

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

ABSTRACT: 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.

SUBMITTER: Min KS

PROVIDER: S-EPMC7661263 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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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).

Min Kwan-Sik KS Park Jong-Ju JJ Lee So-Yun SY Byambaragchaa Munkhzaya M Kang Myung-Hwa MH

BMC biotechnology 20201111 1

<h4>Background</h4>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 an ...[more]

PMID: 33176770

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Project description:We hypothesized that stimulatory and superovulatory treatments, using equine chorionic gonadotropin (eCG), modulate the expression of genes related to insulin, cellular modelling and angiogenesis signaling pathways in the bovine corpus luteum (CL). Therefore, we investigated: 1-the effect of these treatments on circulating insulin and somatomedin C concentrations and on gene and protein expression of INSR, IGF1 and IGFR1, as well as other insulin signaling molecules; 2-the effects of eCG on gene and protein expression of INSR, IGF1, GLUT4 and NFKB1A in bovine luteal cells; and 3-the effect of stimulatory and superovulatory treatments on gene and protein expression of ANG, ANGPT1, NOS2, ADM, PRSS2, MMP9 and PLAU. Serum insulin did not differ among groups (P = 0.96). However, serum somatomedin C levels were higher in both stimulated and superovulated groups compared to the control (P = 0.01). In stimulated cows, lower expression of INSR mRNA and higher expression of NFKB1A mRNA and IGF1 protein were observed. In superovulated cows, lower INSR mRNA expression, but higher INSR protein expression and higher IGF1, IGFR1 and NFKB1A gene and protein expression were observed. Expression of angiogenesis and cellular modelling pathway-related factors were as follows: ANGPT1 and PLAU protein expression were higher and MMP9 gene and protein expression were lower in stimulated animals. In superovulated cows, ANGPT1 mRNA expression was higher and ANG mRNA expression was lower. PRSS2 gene and protein expression were lower in both stimulated and superovulated animals related to the control. In vitro, eCG stimulated luteal cells P4 production as well as INSR and GLUT4 protein expression. In summary, our results suggest that superovulatory treatment induced ovarian proliferative changes accompanied by increased expression of genes providing the CL more energy substrate, whereas stimulatory treatment increased lipogenic activity, angiogenesis and plasticity of the extracellular matrix (ECM).

Project description:Immunopathological outcomes in Systemic Lupus Erythematosus (SLE; or lupus) are believed to be autoantibody-mediated. Conditions which promote a Th2 skew (such as pregnancy) should encourage antibody production, worsening antibody-mediated diseases while ameliorating Th1/Th17-mediated diseases. Although an increased propensity toward autoreactivity can be observed in pregnant lupus patients and in pregnant lupus-prone mice, whether a unique human pregnancy-specific factor can contribute to such effects is unknown. This study assessed whether human chorionic gonadotropin (hCG, a pregnancy-specific hormone of diverse function) at physiological concentrations could mediate stimulatory influences on immune parameters in non-pregnant, lupus-prone mice, in light of the hormone's ameliorating effects on Th1-mediated autoimmunity in murine models. Results demonstrate that administration of hCG heightened global autoreactivity in such mice; antibodies to dsDNA, RNP68, Protein S, Protein C, β2-glycoprotein 1, and several phospholipids were enhanced, and hormone administration had adverse effects on animal survival. Specifically in splenic cell cultures containing cells derived from lupus-prone mice, hCG demonstrated synergistic effects with TLR ligands (up-modulation of costimulatory markers on B cells) as well as with TCR stimuli (enhanced proliferative responses, enhanced levels of cytokines, and the phosphorylation of p38). In both instances, enhanced synthesis of lupus-associated cytokines was observed, in addition to the heightened generation of autoantibodies reactive toward apoptotic blebs. These results suggest that selective transducive, proliferative, and differentiative effects of hCG on adaptive immune cells may drive autoreactive responses in a lupus environment, and may also potentially provide insights into the association between the presence of higher hCG levels (or the administration of hCG) with the presence (or appearance) of humoral autoimmunity.

Dataset Information

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

Publications

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

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