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Category: bioengineering – Page 141

Circa 2017

Efficient intracellular delivery of biologically active macromolecules has been a challenging but important process for manipulating live cells for research and therapeutic purposes. There have been limited transfection techniques that can deliver multiple types of active molecules simultaneously into single-cells as well as different types of molecules into physically connected individual neighboring cells separately with high precision and low cytotoxicity. Here, a high frequency ultrasound-based remote intracellular delivery technique capable of delivery of multiple DNA plasmids, messenger RNAs, and recombinant proteins is developed to allow high spatiotemporal visualization and analysis of gene and protein expressions as well as single-cell gene editing using clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein-9 nuclease (Cas9), a method called acoustic-transfection. Acoustic-transfection has advantages over typical sonoporation because acoustic-transfection utilizing ultra-high frequency ultrasound over 150 MHz can directly deliver gene and proteins into cytoplasm without microbubbles, which enables controlled and local intracellular delivery to acoustic-transfection technique. Acoustic-transfection was further demonstrated to deliver CRISPR-Cas9 systems to successfully modify and reprogram the genome of single live cells, providing the evidence of the acoustic-transfection technique for precise genome editing using CRISPR-Cas9.

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A research group at ETH Zurich, Switzerland, has made it possible to edit hundreds of genes at once with CRISPR gene editing.

CRISPR gene editing has revolutionized the biotech industry by providing an easy and quick way to genetically modify organisms. So far, however, CRISPR techniques have only managed to edit a maximum of seven genes at once. This limits the potential of the technique in creating cell therapies, since whole networks of genes need to be reprogrammed to control each cell’s fate.

The Swiss research group devised a way to overcome this limitation with a CRISPR technique able to edit 25 genes in one go. This number could also be increased to up to hundreds of genes at a time. This method therefore makes it possible to edit gene networks, and reprogram stem cells to become cell therapies such as skin cells or insulin-producing pancreatic cells.

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Animal models provide benefits for biomedical research, but translating such findings to human physiology can be difficult. The human heart’s energy needs and functions are difficult to reproduce in other animals, such as mice and rats. One new system looks to circumvent these issues and provide a functional view of how different treatments can help ailing cells in the heart following oxygen and nutrient deprivations.

Researchers have unveiled a new silicon chip that holds human lab-grown for assessing the effectiveness of new drugs. The system includes heart cells, called cardiomyocytes, patterned on the chip with electrodes that can both stimulate and measure within the cells. The researchers discuss their work in this week’s APL Bioengineering.

These capabilities provide a way for determining how the restriction of blood supply, a dangerous state known as ischemia, changes a heart’s conduction velocity, beat frequency and important electrical intervals associated with heart function.

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The ability to modify multiple genetic elements simultaneously would help to elucidate and control the gene interactions and networks underlying complex cellular functions. However, current genome engineering technologies are limited in both the number and the type of perturbations that can be performed simultaneously. Here, we demonstrate that both Cas12a and a clustered regularly interspaced short palindromic repeat (CRISPR) array can be encoded in a single transcript by adding a stabilizer tertiary RNA structure. By leveraging this system, we illustrate constitutive, conditional, inducible, orthogonal and multiplexed genome engineering of endogenous targets using up to 25 individual CRISPR RNAs delivered on a single plasmid. Our method provides a powerful platform to investigate and orchestrate the sophisticated genetic programs underlying complex cell behaviors.

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