Reconstitution of B-1a Cells in Murine Allogeneic Transplantation of Umbilical Cord Blood Cells: A New Advantage of the Umbilical Cord Blood-Derived Hematopoietic System.
Authors: Sato Hideaki , Ito Kyoko , Shinohara Nobukata , Ito Koichi 
Published: October 2019

Abstract
Abstract 2427
Poster Board II-404

Background:
Umbilical cord blood cells (UCBCs) have been used for the treatment of various hematological diseases. In clinical situations, perfect major histocompatibility complex (MHC) matching cannot be expected in UCBC transplantation because UCBC transplantation essentially involves non-related donor-recipient combinations. Although even MHC-mismatched UCBCs can be used because of the low content of mature T cells, the extent of recovery of immune function in lymphocytes derived from MHC-mismatched UCBC transplantation has remained unclear. To resolve this issue, we examined the functional maturation of lymphocytes derived from a fully MHC-mismatched UCBC transplantation in a murine model system. For comparison, bone marrow cell (BMC) transplantation was also performed. In this study, we first found that B-1a cells, which produce natural IgM antibodies against pathogens such as Streptococcus pneumoniae and the influenza virus, can be generated from UCBCs but not from BMCs.

Methods:
RAG2(-/-)BALB/c [H-2d] mice exposed to 8-Gray irradiation were transplanted with 3 different doses (1×105, 5×105, or 1×106 cells) of UCBCs or BMCs, which had been depleted of mature T cells, obtained from green fluorescent protein-transgenic C57BL/6 (B6) [H-2b] mice. More than 16 weeks after transplantation, we observed successful reconstitution of immune cells in the recipients, as shown by flow cytometric analysis. In addition, the function of T and B cells developed from UCBCs and BMCs was evaluated by examining the rejection of skin grafts from third-party donors and the production of antibodies against intraperitoneally injected 2,4,6-trinitrophenyl-keyhole limpet hemocyanin (TNP-KLH), respectively. Furthermore, the ability of UCBCs and BMCs to repopulate natural IgM antibody-producing B-1a cells was examined using flow cytometry and enzyme-linked immunosorbent assay.

Results:
At 16 weeks after transplantation, a similar survival rate (40-100%), which was dose-dependent, was observed between recipient mice with UCBC and BMC transplants. Phenotypically mature fluorescent cells of donor origin, including T cells, B cells, monocytes, and granulocytes, were detected in the recipients' peripheral blood, even after MHC-mismatched UCBC and BMC transplantation. Functional analysis showed that mice with allogeneic UCBC and BMC transplants accepted skin grafts from both BALB/c and C57BL/6 mice. However, these chimeric mice completely rejected skin grafts from third-party C3H/HeJ [H-2k] mice, indicating that both CD8+ killer and CD4+ helper T cells were functionally mature in the recipients. In addition, TNP-KLH-immunized mice with UCBC and BMC transplants produced both TNP-specific IgM and IgG antibodies. These findings indicate the ability of recipient mice to develop antibody responses with Ig class switching to T-dependent antigens, thereby confirming that both B cells and CD4+ helper T cells derived from allogeneic UCBC were immunologically competent. In contrast, surprisingly, flow cytometric analysis revealed the presence of peritoneal B-1a cells only in UCBC recipients but not in BMC recipients; the presence of these cells appeared to correlate with the bacterial polysaccharide-reactive natural IgM level in UCBC recipients.

Conclusion:
Although UCBCs and BMCs essentially have similar ability for reconstituting the hematopoietic system, this study showed the promising potential of UCBCs in reconstituting B-1a cells even in an allogeneic environment, thereby replenishing polysaccharide-reactive IgM as a front-line defensive factor against invading pathogens. On the basis of our findings, we strongly recommend the use of UCBCs in hematopoietic stem cell transplantation.

Disclosures:
No relevant conflicts of interest to declare.