Derivation of induced pluripotent stem cells line (RCPCMi007-A-1) with inactivation of the beta-2-microglobulin gene by CRISPR/Cas9 genome editing.

The mismatch of HLA haplotypes between donor and recipient adversely affects the outcome of tissue transplantation. TheB2Mgene knockout (B2M-KO) disrupts the HLA I heterodimer formation; therefore,B2M-KO cells have reduced immunogenicity to allogeneic CD8+ T cells. Thus, theB2M-KO IPSCs and their derivatives can potentially solve a problem of the immunological compatibility in allogeneic transplantations. Using CRISPR/Cas9-mediated genome editing, we generated a human B2M-KO iPSC line (RCPCMi007-A-1). The RCPCMi007-A-1 iPSCs express pluripotency markers, have typical stem cell morphology, maintain normal karyotype, and the ability to differentiate into three germ layers.

[At Home among Strangers: Is It Possible to Create Hypoimmunogenic Pluripotent Stem Cell Lines?]

Human pluripotent stem cells, which include embryonic stem cells and induced pluripotent cells (iPSCs), are capable of unlimited division and differentiation into all cells of the body. These cells are considered as a potential source of various types of cells for transplantations. The use of autologous iPSCs is not potentially associated with immune rejection and does not require immunosuppression required for allogeneic grafts. However, the high cost of this technology and the duration of obtaining iPSCs and differentiated cells may limit the use of autologous iPSCs in clinical practice. In addition, full equivalence and immunological compatibility of autologous iPSCs and their derivatives have been repeatedly questioned. One approach to solving the problem of the immunological compatibility of allogeneic derivatives of iPSCs can be the establishment of cell lines with reduced immunogenicity. Differentiated derivatives of such iPSCs may be suitable for transplantation to any patient. This review discusses the strategies for evading immune surveillance in normal and tumor processes that can be used to establish stem cell lines with reduced immunogenicity.

"Necessity Is the Mother of Invention" or Inexpensive, Reliable, and Reproducible Protocol for Generating Organoids.

Organoids are three-dimensional (3D) cell cultures that replicate some of the key features of morphology, spatial architecture, and functions of a particular organ. Organoids can be generated from both adult and pluripotent stem cells (PSCs), and complex organoids can also be obtained by combining different types of cells, including differentiated cells. The ability of pluripotent cells to self-organize into organotypic structures containing several cell subtypes specific for a particular organ was used for creating organoids of the brain, eye, kidney, intestine, and other organs. Despite the advantages of using PSCs for obtaining organoids, an essential shortcoming that prevents their widespread use has been a low yield when they are obtained from a PSC monolayer culture and a large variation in size. This leads to great heterogeneity on further differentiation. In this article, we describe our own protocol for generating standardized organoids, with emphasis on a method for generating brain organoids, which allows scaling-up experiments and makes their cultivation less expensive and easier.