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Since the gene-editing potential of CRISPR systems was realized in 2013, they have fundamentally transformed our ability to manipulate genomes and been utilized in laboratories across the world for a wide variety of applications. When this gene-editing power is combined with the proliferative potential of stem cells, scientists have leveled up their understanding of cell biology, human genetics and developmental biology, and the future potential of regenerative medicine.

As a consequence of CRISPR/Cas9 systems, next-generation genome sequencing, and stem cell technologies have matured, the possibilities for their combined use have also enlightened. Here are the latest advances in this rapidly evolving field to showcase the current progress in the intersection of gene-editing (such as gene-knockout, gene-knockin, and point mutation, etc.) stem cell research.

The ability to modify selectively specific genes provides a powerful tool for characterizing gene functions, performing gene therapy, correcting specific genetic mutations, eradicating diseases, engineering cells and organisms to achieve more innovative functions and obtain transgenic animals as models for disease studies. Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology has revolutionized genome engineering. The application of CRISPR/Cas9 systems for stem cell research allows researchers to develop new disease models to explore new therapeutic tools and address degenerative diseases. Moreover, there are challenges and future perspectives regarding the use of CRISPR/Cas9 as a helpful technology in the invention of new medical therapies.

1. Using the CRISPR/Cas9 to create stem cells that antagonize IL-1- / TNF-α-mediated inflammation

To create stem cells that antagonize IL-1- and TNF-α-mediated inflammation in an autoregulated manner, researchers applied the genome editing system CRISPR/Cas9. Transgenes encoding a firefly luciferase transcriptional reporter or a cytokine antagonist, either murine IL-1Ra or a chimeric human sTNFR1-murine immunoglobulin G (Bloquel et al., 2004), were targeted to the Ccl2 start codon in murine iPSCs cells (Diekman et al., 2012) using the CRISPR/Cas9 platform.

The results show that genome engineering can be applied successfully to rewire endogenous cell circuits to allow for prescribed input/output relationships between inflammatory mediators and their antagonists. This provides a foundation for cell-based drug delivery or cell-based vaccines via a rapidly responsive, autoregulated system. The customization of intrinsic cellular signaling pathways in stem cells also provides innovative ideas for safer and more effective therapeutic approaches for several kinds of diseases.

2. Using the CRISPR/Cas9 system in genetic modification of pluripotent or multipotent stem cells

Combined with the pluripotency of stem cells, the technology represents a powerful tool to generate various cell types for disease modeling, drug screening, toxicology, and targeted therapies. Generally, the CRISPR/Cas9 system has been applied in genetic modification of pluripotent or multipotent stem cells, after which the cells are differentiated into specific cell types and used for functional analysis or even clinical transplantation. Recent advancement in CRISPR/Cas9 technology has widened the scope of stem cell research and its therapeutic application. This review provides an overview of the current application and the prospect of CRISPR/Cas9 technology, particularly in stem cell research and therapy.

3. combining iPSC and CRISPR/Cas9 technologies for the investigation of the molecular and cellular mechanisms of inherited diseases

CRISPRi permits gene repression at the transcription level, as opposed to RNAi which controls genes at the mRNA level. This allows researchers to repress certain genes within stem cells and decipher their function. Kampmann explains: "For CRISPRi, we target a transcriptional repressor domain (the KRAB domain) to the transcription start site of genes to repress their expression. This knockdown approach is highly effective and lacks the notorious off-target effects of RNAi-based gene knockdown.”Using CRISPR/Cas9 systems(such as gene-knockout, gene-knockin, and point mutation, etc.) to mediate iPSC stem cells is helpful to investigate mechanisms of inherited diseases including immunological, metabolic, hematological, neurodegenerative, and cardiac diseases.

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