The role of proteinase inhibitor (PI)-9 in hematopoietic cells remains unclear. To clarify the role of PI-9 in these cells, we compared the expressions of PI-9 mRNA and antigen with those of granzyme B (GrB). While the strongest expression of PI-9 mRNA was observed in a NK cell line YT-N10, it was also expressed in a B-acute lymphoblastic leukemia cell line U-Tree02, an Epstein-Barr Virus (EBV)-transformed B cell clone, a CD8+ T lymphocyte clone and a megakaryocytic cell line CMK, but not in a T cell line Jurkat. Phorbol 12-myristate 13 acetate (PMA) enhanced PI-9 mRNA expression in the CD8+ T lymphocyte clone and YT-N10 cells prior to GrB mRNA expression. IL-2 and IL-12 also had similar effects. PMA increased PI-9 mRNA expression in the EBV-transformed B cell clone and CMK cells, but IL-6 showed no effect. No changes were noted in PI-9 and GrB antigens after the addition of these agonists. Patients with graft-versus-host disease (GVHD) may have activated CTLs and NK cells. We therefore examined the expression of PI-9 and GrB mRNAs in eight patients after allogeneic hematopoietic stem cell transplantation with GVHD (n = 4) or without chronic GVHD (n = 4). Expression of GrB mRNA was significantly increased in three patients with GVHD and one patient without GVHD. Surprisingly, PI-9 mRNA expression was decreased in the eight patients. These results indicate that earlier synthesis of PI-9 may be essential for the prevention of autolysis of immunocompetent cells, and that the expression of PI-9 and GrB mRNAs may be controlled through different pathways.