ORP4L expression induces T-cell malignant transformation and T-cell leukemogenesis in mice. (A) The experimental design. Normal murine CD4⁺ T cells were infected with lentivirus with or without ORP4L expression and cultured for 16 weeks in vitro; then, the cells were used for malignant transformation assay or B-NDG mice transplantation. (B) Cell death analysis of T cells infected with lentivirus carrying ORP4L or not. After culture for 16 weeks in vitro, the cell death rates were detected at 0, 3, 6, and 9 days after removing the IL-2 from the culture medium. Mean ± standard deviation (SD; n = 3 experiments; Student t test). (C) Kaplan-Meier comparative survival analysis of B-NDG mouse recipients of T-cell transplants with or without ORP4L expression (n = 8 mice each group, log-rank test). (D) The percentage of CD3⁺CD4⁺, CD4⁺CD8⁺, CD44⁺CD25⁺, and CD3⁺c-Kit⁺ cells in the peripheral blood of B-NDG mice treated as in panel C (n = 8 mice each group; Student t test). (E) Representative images of peripheral blood smear from B-NDG mice treated as in panel C. The number of abnormal lymphocytes is indicated below the smears for both cohorts. Bar represents 10 μm. (F-G) Splenomegaly (F) and hepatomegaly (G) in B-NDG mice treated as in panel C. Representative organs are shown on the left, with organ weights on the right. Mean ± SD (n = 8 mice each group; Student t test). (H) Representative histologic hematoxylin and eosin–stained sections showing T-cell infiltration in spleen, liver, and lung of B-NDG mice treated as in panel C. Immunohistochemical human CD3-specific antibody staining (insets) of the tissues are shown. Bar represents 100 μm. (I) Western blot analysis of ORP4L expression in T cells. ORP4L expression in T cells infected with lentivirus with or without ORP4L before transplantation (top) or ORP4L expression in T cells isolated from 2 B-NDG mice after transplantation when the mice are sick (bottom). **P < .01; ***P < .001.