Manipulation of the Wnt/b-catenin pathway during pig´s oocytes IVM aiming to produce clones by SCNT

• Grant number: 23/12977-0
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• Start date: May 01, 2024
• End date: September 30, 2027
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Animal Reproduction
• Principal Investigator: Flávio Vieira Meirelles
• Grantee: Thais Naomi Goncalves Nesiyama
• Host Institution: Faculdade de Zootecnia e Engenharia de Alimentos (FZEA). Universidade de São Paulo (USP). Pirassununga , SP, Brazil
• Associated research grant: 13/08135-2 - CTC - Center for Cell-Based Therapy, AP.CEPID
• Associated scholarship(s): 25/03340-4 - UNDESTANDING THE RSPO2 ROLE IN SWINE OOCYTE MATURATION AND EMBRYONIC DEVELOPMENT, BE.EP.DD

Abstract

Pigs have been used as an experimental model in biomedical research since Ancient Greece, mainly due to their physiological, anatomical, and genetic similarity with humans. Studies with transgenic porcine models are carried out to promote advances in scientific research, through the use of two tools, such as CrispR-Cas9 and in vitro production (IVP) of clone embryos through somatic cell nuclear transfer (SCNT). The oocyte in vitro maturation (IVM) step is fundamental for IVP and requires a series of complex nuclear, cytoplasmic, and molecular events. In this way, studies during the IVM stage can contribute to the IVP of porcine clone embryos. Studies with manipulation of the Wnt/²-catenin pathway during IVM, using R-spondin 2 (RSPO2) or Dickkopf-related protein 1 (DKK1), have shown to improve oocyte maturation and embryo development. Thus, the objective of this study will be to evaluate the effects of the association of treatments of swine oocytes with RSPO2 and DKK1 during IVM, aiming at the production of parthenotes and clone embryos (Mini-pigs and Onco-pigs) through SCNT. The manipulation of the Wnt/²-catenin pathway will be performed by supplementation with RSPO2 and DKK1 to the IVM media, where 10 groups will be established: Control (CTL/CTL), Negative Control (CTL without PFF/ CTL without PFF), A (DKK1/DKK1), B (RSPO2/RSPO2), C (DKK1/RSPO2) and D (RSPO2/DKK1), E (CTL/DKK1), F (DKK1/CTL), G (CTL/RSPO2), and H (RSPO2/ CTL). Matured oocytes will be activated by parthenogenesis and the rates of maturation will be evaluated, as well as the activation and development of blastocysts. The treatment with the best rates will be chosen for cloning, the embryos produced will be transferred to synchronized adult females, and any pregnancies will be followed up until delivery, and their offspring evaluated. Binary data results will be analyzed using the Chi-square test and the others using ANOVA. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

NESIYAMA, THAIS NAOMI GONSALVES; SANGALLI, JULIANO RODRIGUES; DE BEM, TIAGO HENRIQUE CAMARA; RECCHIA, KAIANA; MARTINS, SIMONE MARIA MASSAMI KITAMURA; DE ANDRADE, ANDRE FURUGEN CESAR; FERST, JULIANA GERMANO; ALMEIDA, GUSTAVO HENRIQUE DONA RODRIGUES; MARQUES, MARIANA GROKE; DORIA, RENATA GEBARA SAMPAIO; et al. Swine clones: potential application for animal production and animal models. ANIMAL REPRODUCTION, v. 22, n. 1, p. 13-pg., 2025-01-01. (21/09886-8, 23/12977-0, 13/08135-2)