Conceptus interferon gamma is essential for establishment of pregnancy in the pig

2021 ◽  
Author(s):  
Destiny N Johns ◽  
Caroline G Lucas ◽  
Caroline A Pfeiffer ◽  
Paula R Chen ◽  
Ashley E Meyer ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Somatic Cell ◽  
Interferon Gamma ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Secretory Protein ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Proinflammatory Response ◽  
Ifng Production

Abstract Establishment and maintenance of pregnancy in the pig is a complex process that relies on conceptus regulation of the maternal proinflammatory response to endometrial attachment. Following elongation, pig conceptuses secrete interferon gamma (IFNG) during attachment to the endometrial luminal epithelium. The objective here was to determine if conceptus production of IFNG is important for early development and establishment of pregnancy. CRISPR/Cas9 gene editing and somatic cell nuclear transfer technologies were used to create an IFNG loss-of-function study in pigs. Wild-type (IFNG+/+) and null (IFNG−/−) fibroblast cells were used to create embryos through somatic cell nuclear transfer. IFNG expression was not detected in IFNG−/− conceptuses on either day 15 or day 17 of pregnancy. Ablation of conceptus IFNG production resulted in the reduction of stromal CD3+ and mast cells which localized to the site of conceptus attachment on day 15. The uteri of recipients with IFNG−/− conceptuses were inflamed, hyperemic and there was an abundance of erythrocytes in the uterine lumen associated with the degenerating conceptuses. The endometrial stromal extracellular matrix was altered in the IFNG−/− embryo pregnancies and there was an increased endometrial mRNA levels for collagen XVII (COL17A1), matrilin 1 (MATN1), secreted phosphoprotein 1 (SPP1) and cysteine-rich secretory protein 3 (CRISP3), which are involved with repair and remodeling of the extracellular matrix. These results indicate conceptus IFNG production is essential in modulating the endometrial proinflammatory response for conceptus attachment and survival in pigs.

38 Improved Cloning Efficiency and Developmental Potential in Bovine Somatic Cell Nuclear Transfer with the New Technology

2018 ◽  
Vol 30 (1) ◽  
pp. 158
Author(s):  
L. Xu ◽  
M.-D. Joo ◽  
A. Mesalam ◽  
S.-H. Song ◽  
S. Zhang ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Reverse Transcription ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Total Cell ◽  
Developmental Competence ◽  
Mrna Levels ◽  
Methyl Transferase ◽  
Developmental Potential

Bovine somatic cell nuclear transfer (SCNT) embryos can develop to the blastocyst stage at a rate similar to that of embryos produced by IVF; however, its efficiency remains low. In this study, we examined the effects of cytoplasm restoration of enucleated oocyte, by injecting ~30% of the cytoplasm of a donor oocyte to restore the enucleated oocyte cytoplasm volume to normal, on the developmental competence and quality of bovine cloned embryos during pre-implantation using the TUNEL assay, quantitative reverse transcription PCR (RT-qPCR) and immunocytochemistry. The experiment was conducted in 6 replicates. The differences in embryo development and expression levels of the various genes between experimental groups were analysed by one-way ANOVA. The level of statistical significance was set at P < 0.05. The percentages of embryos that underwent cleavage and formed a blastocyst were significantly higher (P < 0.05) in the cytoplasmic injected group than in the traditional SCNT group (61.5 ± 1.3% v. 39.7 ± 2.1% and 28.9 ± 0.8% v. 20.2 ± 1.3%, respectively). Furthermore, the beneficial effects of cytoplasmic injection on the cloned embryos were associated with a significantly increased (P < 0.05) total cell number in Day 8 blastocysts compared with the traditional SCNT group (176.2 ± 6.5 v. 119.3 ± 7.7; P < 0.05); however, there was no difference (P > 0.05) between the number of apoptotic cells per blastocyst in the cytoplasmic injected group and in the traditional SCNT group (3.5 ± 1.1 v. 4.1 ± 0.8). Moreover, cytoplasm restoration of enucleated oocyte significantly increased (P < 0.05) mitochondrial activity, as identified by MitoTracker Green (Thermo Fisher Scientific, Waltham, MA, USA). Reverse transcription-qPCR showed that the mRNA levels of DNA methyl-transferase 1 and DNA methyl-transferase 3a were significantly decreased (P < 0.05) in cytoplasmic injected group compared with the traditional SCNT group, but did not significantly differ (P > 0.05) between the cytoplasmic injected and IVF groups. Taken together, these data suggest that cytoplasm restoration of enucleated oocyte improves in vitro developmental competence and quality of bovine cloned embryos, as evidenced by increased total cell numbers, reprogramming efficiency, and mitochondria activity. This work was partly supported by grant from the Next-Generation BioGreen21 (No. PJ01107703), IPET (No. 315017-5 and 117029-3), Allergy free cat (Co. Felix Pets) and BK21plus.

261 PHENOTYPIC CHARACTERISTICS AND TISSUE-SPECIFIC IGF2R/IGF2 EXPRESSION PARTITION BOVINE FETAL OVERGROWTH ASSOCIATED WITH IN VITRO FERTILIZATION AND SOMATIC CELL NUCLEAR TRANSFER CLONING

2007 ◽  
Vol 19 (1) ◽  
pp. 247 ◽  
Author(s):  
D. Bebbere ◽  
S. E. Ulbrich ◽  
V. Zakhartchenko ◽  
M. Weppert ◽  
H.-D. Reichenbach ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Liver Weight ◽  
Mrna Levels ◽  
Crown Rump Length ◽  
Transcript Levels ◽  
Vitro Fertilization

Large offspring syndrome (LOS) in ruminants refers to various poorly defined organ pathologies that are associated with fetal overgrowth and are encountered after a range of embryo manipulations (Rhind et al. 2003 Nat. Rev. Genet. 4, 855–864). We have explored the effects of somatic cell nuclear transfer (NT) and in vitro fertilization (IVF) on phenotype and relative expression levels of 2 imprinted genes important for fetal growth, insulin-like growth factor 2 (IGF2) and its receptor (IGF2R). Viable bovine fetuses were recovered near the end of the first trimester of pregnancy, and skeletal muscle, liver, and lung were sampled for real-time RT-PCR analyses. We compared NT-fetuses (n = 23), IVF-fetuses (n = 24), and fetuses generated by artificial insemination (controls, n = 24) in order to separate abnormalities specific to cloning from effects of in vitro gamete and embryo manipulation. Nuclear transfer and IVF-fetuses, both derived from embryos cultured with 10% estrous cow serum, demonstrated significant fetal overgrowth. The increase in body weight relative to controls was similar for both groups (+22%, P &lt; 0.001, and +19%, P &lt; 0.001, respectively), but further analyses clearly separated the NT phenotype from the IVF phenotype. The NT-fetuses were characterized by a shorter crown-rump length but larger thorax circumference, which consequently produced a significantly reduced fetus length-to-thorax circumference ratio in comparison with IVF-fetuses and controls (-9% each, P &lt; 0.0001). Absolute liver weight was significantly increased in NT- and IVF-fetuses (+62%, P &lt; 0.0001, and +20%, P &lt; 0.0001, respectively), but relative liver weight was increased only in NT-fetuses (+30%, P &lt; 0.0001). Heart (P&lt; 0.0001) and kidney (P = 0.0003) mass also showed disproportionate organomegaly in NT-fetuses only, but relative lung weight (NT, P = 0.263; IVF, P = 0.317) was not affected by either embryo technique. Transcript abundance for IGF2 and IGF2R genes were strongly correlated in muscle (r = 0.835, P &lt; 0.0001), liver (r = 0.922, P &lt; 0.0001), and lung (r = 0.772, P &lt; 0.0001). The IGF2 and IGF2R transcript levels in muscle tissue from NT-fetuses were markedly reduced in comparison with both IVF-fetuses (-31%, P &lt; 0.0001, and -41%, P &lt; 0.0001, respectively) and controls (-31%, P &lt; 0.0001, and -41%, P &lt; 0.0001, respectively). In liver tissue, however, transcript levels for NT-fetuses were similar to those of controls, and IVF-fetuses showed markedly elevated, albeit non-significant, IGF2 (+86%, P = 0.0591) and IGF2R (+54%, P = 0.1305) mRNA levels relative to controls. Our data demonstrate that seemingly similar syndromes caused by NT or IVF procedures can be clearly partitioned with respect to phenotype and IGF2/IGF2R expression.

scholarly journals Author Correction: Application of interspecific Somatic Cell Nuclear Transfer (iSCNT) in sturgeons and an unexpectedly produced gynogenetic sterlet with homozygous quadruple haploid

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Effrosyni Fatira ◽  
Miloš Havelka ◽  
Catherine Labbé ◽  
Alexandra Depincé ◽  
Viktoriia Iegorova ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer

scholarly journals Stem cell therapies and benefaction of somatic cell nuclear transfer cloning in COVID-19 era

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Birbal Singh ◽  
Gorakh Mal ◽  
Vinod Verma ◽  
Ruchi Tiwari ◽  
Muhammad Imran Khan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Regenerative Medicine ◽  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Drug Testing ◽  
Human Interaction ◽  
Patient Specific ◽  
Wide Range

Abstract Background The global health emergency of COVID-19 has necessitated the development of multiple therapeutic modalities including vaccinations, antivirals, anti-inflammatory, and cytoimmunotherapies, etc. COVID-19 patients suffer from damage to various organs and vascular structures, so they present multiple health crises. Mesenchymal stem cells (MSCs) are of interest to treat acute respiratory distress syndrome (ARDS) caused by SARS-CoV-2 infection. Main body Stem cell-based therapies have been verified for prospective benefits in copious preclinical and clinical studies. MSCs confer potential benefits to develop various cell types and organoids for studying virus-human interaction, drug testing, regenerative medicine, and immunomodulatory effects in COVID-19 patients. Apart from paving the ways to augment stem cell research and therapies, somatic cell nuclear transfer (SCNT) holds unique ability for a wide range of health applications such as patient-specific or isogenic cells for regenerative medicine and breeding transgenic animals for biomedical applications. Being a potent cell genome-reprogramming tool, the SCNT has increased prominence of recombinant therapeutics and cellular medicine in the current era of COVID-19. As SCNT is used to generate patient-specific stem cells, it avoids dependence on embryos to obtain stem cells. Conclusions The nuclear transfer cloning, being an ideal tool to generate cloned embryos, and the embryonic stem cells will boost drug testing and cellular medicine in COVID-19.

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