The gold standard improves: a better assay for HSCs

Comment on McKenzie et al, page 1259

The combination of intrafemoral injection and depletion of CD122⁺ cells provides a new and improved approach for xenograft analysis of human hematopoietic stem and progenitor cells.

Transplantation of human hematopoietic stem cells (HSCs) into immune-deficient mice has become the accepted “gold” standard for in vivo analysis of primitive cells. Studies from several investigators have demonstrated that productive engraftment and growth of human hematopoietic cells in mice are mediated by stem and progenitor cell populations.¹  Thus, successful xenografts can be used in a fashion analogous to syngeneic systems to detect and quantitate stem cell activity. Over the past 10 to 15 years, the approach has improved as strains of mice more permissive to supporting human tissue have become available. Most prominently, the nonobese/diabetic severe combined immune deficiency (NOD/LtSzscid/scid or NOD/SCID) mouse has emerged as the most commonly used host for assessing properties of human HSCs. While conventional use of this strain is a powerful tool, recent reports have suggested that at least some stem and progenitor populations are not readily detected in NOD/SCID mice. In this issue of Blood, McKenzie and colleagues describe a simple modification of the basic NOD/SCID transplant system that greatly improves sensitivity of the assay.

The use of immune-deficient mice as recipients of human stem cell grafts is a common strategy available to most laboratories.²  The NOD/SCID strain was initially derived from the SCID mouse by Shultz and colleagues at Jackson Laboratories.³  In comparison to SCID mice, which do not make mature T and B cells, NOD/SCID mice have an excellent capacity for engraftment with human cells due to additional defects in natural killer (NK) cell and macrophage function, and circulating complement.³  NOD/SCID-β₂-microglobulin-null (NOD/SCID-β₂m^–/–) mice have a more absolute immunodeficiency than the NOD/SCID strain, and have virtually no NK cell function.⁴  They are thus even more permissive than NOD/SCID mice for supporting xenografts. Unfortunately, due to a short lifespan and poor breeding characteristics, the NOD/SCID-β₂m^–/– strain has serious logistical limitations. Therefore, methods to reduce the residual NK cell activity in the NOD/SCID strain, which breeds robustly, would be very beneficial.

In the manuscript by McKenzie et al, studies demonstrate that in vivo administration of anti-CD122 antibody, which is targeted to murine NK cells, renders the NOD/SCID strain more like the NOD/SCID-β₂m^–/– strain by depleting residual NK activity. The authors also assessed 2 different routes of injection: intravenous versus intrafemoral. These investigators have previously demonstrated that injection of cells directly into the bone marrow cavity of the femur enables identification of a population of “short-term” or rapidly reconstituting progenitors, which are not observed using intravenous injection.⁵  The intrafemoral injection technique is thus useful for cell populations such as maturing progenitors that may be compromised in their ability to home to the bone marrow. The intravenous and intrafemoral injection techniques were compared in NOD/SCID–, NOD/SCID-β₂m^–/––, and anti-CD122 mAb is injected once to condition NOD/SCID mice. Mice that received the same populations of human cells, injected via the intrafemoral versus intravenous routes, were evaluated for the level of human cell engraftment at 2 and 6 weeks after transplantation. These experiments demonstrated that the most effective approach was intrafemoral injection of anti-CD122–treated NOD/SCID mice, which generated even higher engraftment than the NOD/SCID-β₂m^–/– strain. While the improved sensitivity was particularly striking at 2 weeks after transplantation, analysis at 6 weeks also showed substantially higher human cell engraftment in comparison to standard intravenous injection of NOD/SCID mice. These data thus describe a simple and convenient model to assess rapid engraftment of human stem cell populations that might lack normal bone marrow homing capacities, or that are available in numbers too limiting to use conventional intravenous injection routes. ▪