Kinetics of Langerhans cell chimerism in the skin of dogs following 2 Gy TBI allogeneic hematopoietic stem cell transplantation

Abstract

Background: Langerhans cells (LC) are bone marrow-derived cells in the skin. The LC donor/recipient chimerism is assumed to influence the incidence and severity of graft-versus-host disease (GVHD) after hematopoietic stem cell transplantation (HSCT). In nonmyeloablative (NM) HSCT the appearance of acute GVHD is delayed when compared with myeloablative conditioning. Therefore, we examined the development of LC chimerism in a NM canine HSCT model.

Methods: 2 Gy conditioned dogs received bone marrow from dog leukocyte antigen identical littermates. Skin biopsies were obtained pre- and post-transplant. LC isolation was performed by immunomagnetic separation and chimerism analysis by PCR analyzing variable-number-of-tandem-repeat markers with subsequent capillary electrophoresis.

Results: All dogs engrafted. Compared to peripheral blood chimerism the development of LC chimerism was delayed (earliest at day +56). None of the dogs achieved complete donor LC chimerism, although two dogs manifested a 100 % donor chimerism in peripheral blood at days +91 and +77. Of interest, one dog remained LC chimeric despite loss of donor chimerism in the peripheral blood cells.

Conclusion: Our study indicates that LC donor chimerism correlates with chimerism development in the peripheral blood but occurs delayed following NM-HSCT.

Keywords: Chimerism; Dogs; Langerhans cells; Nonmyeloablative; Stem cell transplantation.

Fig. 1

Representative flow cytometric analysis of CD1a expressing epidermal canine Langerhans cells (LC). a Forward scatter (FSC) and side scatter (SSC) characteristics of canine LC. b FACS dot plot showing a purity of 95 % CD1a expressing LC after isolation with MiniMACS technology

Fig. 2

Electron microscopic image of a Langerhans cell. The figure insert shows a characteristic Birbeck granule (black arrow)

Fig. 3

Chimerism kinetics of Langerhans cells (LC) in comparison to the peripheral blood chimerism. Development of LC donor chimerism (bars) compared with donor chimerism of PBMC (solid line) and granulocytes (dotted line) after 2 Gy nonmyeloablative HSCT in two dogs. a Dog No. 3 with full donor chimerism in peripheral blood. Continuously increasing LC donor chimerism starting at day +56 after HSCT at a time when the dog experienced strong engraftment in the peripheral blood. Donor chimerism of LC developed delayed compared to donor chimerism in the peripheral blood and did not achieve the peripheral blood levels during the observation period b Dog No 6 with initial engraftment and subsequent late graft rejection. Despite high initial donor chimerism levels in the peripheral blood of 82 % (granulocytes d +28) and 62 % (PBMC d +21) first LC donor chimerism was not detected before day +112 probably as a consequence of decreasing peripheral blood chimerism levels starting 4 weeks after HSCT. Interestingly, although donor chimerism values of the peripheral blood continuously declined and the graft was eventually rejected at day +391 a continuously increasing LC donor chimerism was observed also beyond the date of graft rejection