Fernanda Maria Policarpo Tonelli

Gene delivery to Nile tilapia spermatogonial stem cells using carboxi-functionalized multiwall carbon nanotubes

Carboxi-functionalized multiwall carbon nanotubes (fMWCNTs), when complexed with DNA, can promote gene delivery to Nile tilapia spermatogonial stem cells with higher transfection efficiency than cationic lipids or electroporation, causing also less cell death.

Gene delivery to Nile tilapia cells for transgenesis and the role of PI3K-c2α in angiogenesis

Microinjection is commonly performed to achieve fish transgenesis; however, due to difficulties associated with this technique, new strategies are being developed. Here we evaluate the potential of lentiviral particles to genetically modify Nile tilapia cells to achieve transgenesis using three different approaches: spermatogonial stem cell (SSC) genetic modification and transplantation (SC), in vivo transduction of gametes (GT), and fertilised egg transduction (ET). The SC protocol using larvae generates animals with sustained production of modified sperm (80% of animals with 77% maximum sperm fluorescence [MSF]), but is a time-consuming protocol (sexual maturity in Nile tilapia is achieved at 6 months of age). GT is a faster technique, but the modified gamete production is temporary (70% of animals with 52% MSF). ET is an easier way to obtain mosaic transgenic animals compared to microinjection of eggs, but non-site-directed integration in the fish genome can be a problem. In this study, PI3Kc2α gene disruption impaired development during the embryo stage and caused premature death. The manipulator should choose a technique based on the time available for transgenic obtainment and if this generation is required to be continuous or not.

Efficient and safe gene transfection in fish spermatogonial stem cells using nanomaterials

Multiwalled carbon nanotubes (MWCNTs), nanographene oxide (NGO), and gold nanorods (NRs) can be functionalized and complexed to DNA to promote efficient gene delivery to Nile tilapia spermatogonial stem cells inducing less cell death than electroporation and the commercial reagents tested. Therefore, nanomaterials can contribute to achieve fish transgenesis.

Stem Cells and Tissue Engineering

Tissue engineering is a research field that aims to offer efficient strategies for tissue repair in an effective way, and it has been developing rapidly in the last years. In this context, embryonic stem cells (ESCs) and adult stem cells (ASCs) are the most promising tools to repair tissue damage and loss of function. New protocols using stem cells and different approaches for scaffolds, production are now available for scientific experimentation. In this chapter, we review the use of stem cells in regenerative medicine and also discuss some important strategies to verify stem cell’s differentiation in functional cells for efficient tissue repair. Additionally, we present a protocol to induce pluripotent stem cells to differentiate into bone cells, in order to generate an in vivo material for bone repair.