Identification of donor hematopoietic progenitor cells after allogeneic liver transplantation

Abstract

The persistence of multilineage microchimerism after solid organ transplantation suggests long-lasting engraftment of donor hematopoietic progenitor cells (HPCs) originated from the transplanted organs. This study describes a method developed to identify donor HPCs in the recipient bone marrow and reports preliminary results obtained after allogeneic liver transplantation. METHODS Inbred male Lewis (LEW) and Brown Norway (BN) rats were used for experiments. Plastic dish nonadherent bone marrow cells (BMC) were prepared and cultured with 2% PWM-SCM for 6 days in a fibrin clot culture system to detect colony-forming units in culture (CFU-C). Clusters (10 to 50 cells per aggregate) and colonies (>50 cells per aggregate) in cultures were counted as CFU-C. LEW-derived CFU-C was identified by in situ immunohistochemical staining of cultured cells in the fibrin clot, using a standard three-step avidin-biotin complex method with monoclonal antibody L21-6, which recognizes the class II MHC antigens of LEW but not BN rats. RESULTS CFU-C generated from 5 × 10 4 naive LEW BMC was 54 ± 18, of which 39 ± 9% (21 ± 7) was composed of L21-6 strong positive cells, including macrophages and dendritic-like cells. CFU-C induced from 5 × 10 4 normal BN BMC was 43 ± 4, and none of CFU-C contained L21-6 + cells. When normal LEW and BN BMC were mixed at various ratios (1:10, 1:100, 1:1000), L21-6 + CFU-C derived from the mixture were 0.2 ± 0.4, 2.3 ± 1.4, and 21.3 ± 5.7, respectively, and correlated to the number of LEW BMC contained in the culture. Finally, the engraftment of donor HPCs in transplant recipients was examined in a LEW to BN liver transplantation model, in which rejection was controlled with a short course of tacrolimus treatment (1 mg/kg for 14 days). L21-6 + CFU-C in BN recipient BMC (5 × 10 5) obtained 14 and 30 days after LEW liver transplantation were 1.0 ± 0.6 and 0.3 ± 0.3, respectively. The frequency of donor HPCs in the bone marrow of these recipients is estimated as 0.1 % to 0.5%, according to L21-6 + CFU-C counts obtained in the previous mixed culture experiment. CONCLUSION This study provides the strong evidence that HPCs in the liver graft were able to engraft into the recipient bone marrow after transplantation. The role of donor HPCs in maintaining

Functional Immune Anatomy of the Liver—As an Allograft

“…BD, bile duct; HA, hepatic artery; HMS, hepatic microcirculatory unit; PV, portal vein. Constant exposure to intestinal microbial products fosters a “tolerogenic” microenvironment with relatively low co‐stimulatory and MHC class II expression on antigen presenting cells (APC), including the endothelium . Sinusoids are the majority microvasculature, lined by liver sinusoidal endothelial cells (LSEC) and Kupffer cells (KC) , which scavenge particulates/antigens, and regulate immune responses , liver regeneration , and fibrogenesis . Tremendous parenchymal regenerative abilities include collagenase‐mediated matrix remodeling , which can reverse fibrosis after elimination of immune injury. This is not observed with other solid organ allografts . A variety of immune leukocytes (classical T and B cells, natural killer [NK], NK T cells [NKT], and γδ‐T cells ) are normal hepatic inhabitants, including hematopoietic stem cells .…”

Heterogeneity and plasticity of hepatocyte lineage cells

“…126 Passenger liver hematopoietic cells 126,127 migrate to the bone marrow after liver transplantation and may establish persistent hematolymphopoietic chimerism. [128][129][130] In experimental models, the syngeneic parenchymal environment of the liver allograft appears to constitute a privileged site for persistent progenitor donor hematopoietic cells. 131 Some studies have also shown that hepatic oval cells and hematopoietic stem cells share CD34, Thy-1, and C-kit mRNA and protein.…”

The role of progenitor cells in repair of liver injury and in liver transplantation

“…The adult liver contains HSCs 93,94 and may serve as a major site of extramedullary hematopoiesis 93 . Passenger liver hematopoietic cells 95,96 migrate to the bone marrow after liver transplantation and may establish persistent hematolymphopoietic chimerism 97–99 . In experimental models, the syngeneic parenchymal environment of the liver allograft appears to constitute a privileged site for persistent progenitor donor hematopoietic cells 100 .…”