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Purpose To evaluate the effect of the nonsteroidal antiinflammatory drugs tolfenamic acid and flunixin meglumine in pregnancy rate and embryo survival of recipient mice subjected to embryo transfer. Methods A total of 142 recipient females were transferred with 2,931 embryos and treated with a single injection of tolfenamic acid (1 mg/kg; n=54 females with 1,129 embryos), flunixin meglumine (2.5 mg/kg; n=46 females with 942 embryos), or bi-distilled water (10 mL/kg) as control group (n=42 females with 860 embryos). Pregnancy was checked 2 weeks after embryo transfer, delivery was registered on the due date, and litter size was recorded on Day 7 after birth. Results Pregnancy rate of tolfenamic acid treated females was significantly higher than flunixin group (P<0.05) and showed a tendency to be higher when compared to the control group (P=0.06). The number of pups born from transferred embryos in pregnant females was significantly higher for both treatment groups compared to controls (P<0.05). Number of pups from total transferred embryos was higher for both treatment groups (P<0.05) when compared to controls. Conclusion The use of tolfenamic acid at the time of embryo transfer improves both pregnancy rate and number of live pups in recipient mice, with optimal effects observed with flunixin meglumine. We suggest that the use of tolfenamic acid has beneficial effects on the maintenance of pregnancy and embryo survival in recipient mice, which should be taken into account for further studies in other mammalian females.

Transgenic mice have important implications in biomedicine, and are widely employed to understand gene functions and their regulation. The improvement of transgenic efficiency is relevant because of the low rate of success for this technology. CXCL2 is a chemokine secreted by macrophages and epithelial cells under proinflammatory stimulus of the innate immune response such as bacterial endotoxins. The main effect of CXCL2 is the recruitment of neutrophils to the site of production to fight infections. The objective of this study was to evaluate the effect of 2 DNA concentrations in the efficiency of the transgenesis process. To this aim we used a luciferase reporter under the control of CXCL2 promoter for the generation of a transgenic line to report activation of innate immune response. A total of 1727 1-cell embryos were divided into 2 experimental groups to be microinjected with 0.5 or 1.0 ng μL-1 of DNA in 25 sessions. Three-week-old B6SJL F1 females (n = 131) were superovulated with 5 IU of eCG i.p. (Novormon, Syntex, Buenos Aires, Argentina) and 5 IU of hCG i.p. (Ovusyn, Syntex) 46 h later, and mated with B6SJL F1 stud males. At the moment of hCG treatment, foster females were mated with vasectomized males to induce pseudogestation. Donor females were euthanized by cervical dislocation 20 h after hCG treatment, and embryos were recovered from the ampulla, denuded in 300 μg mL-1 hyaluronidase (Sigma, St. Louis, MO, USA) and incubated at 37°C with 5% CO2, in drops of M16 media (Sigma) under mineral oil, until microinjection. DNA construction consisted of the luciferase reporter gene under the control of the murine CXCL2 gene promoter. Embryos were microinjected into 1 pronucleus under an inverted microscope (Nikon, NY, USA) using glass microtools and mechanic micromanipulators (Eppendorf, Hamburg, Germany). Intact/injected embryos were assessed 30 min after microinjection. Fifteen to 20 embryos per foster female were transferred in both oviducts. Birth rate, survival of pups at Day 7 after birth, number of transgenic pups assessed by standard PCR, and overall transgenic efficiency was registered for each group. Data were analyzed by Yates-corrected chi-square test. No statistical differences were founded except for a higher number of pups alive/embryo transferred in the lower DNA concentration, suggesting the advantage of using 0.5 ng μL-1 v. 1.0 ng μL-1. Table 1.Effect of DNA concentration in the generation of CXCL2-luc transgenic mice

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