Objective To explore the re-constitution rule of T lymphocyte subsets in bone marrow of patients with hematological malignancies after allogenic hematopoietic stem cell transplantation (allo-HSCT), and its differences with those in the peripheral blood. Methods This study was a prospective study. We collected the bone marrow and peripheral blood samples from 41 patients with hematological malignancies receiving allo-HSCT treatment in Department of Hematology of our hospital from Sep. 2015 to Jan. 2017. During the same period, bone marrow and peripheral blood samples of 7 healthy donors were collected as control samples. Flow cytometry was used to evaluate the distribution of T lymphocyte subsets, including CD4⁺ T cells, CD8⁺ T cells, T-helper cell (Th)1 and Th2 before transplantation and 15, 30, 60, 90 and 180 d after transplantation. Luminex technique was used to evaluate Th1-related cytokines (interleukin 2 receptor[IL-2R] and interleukin 18[IL-18]). Results The proportions of CD4⁺ T cells in bone marrow of the patients with hematological malignancies were significantly lower versus the healthy controls 15 and 30 d after transplantation (both P<0.05), and no recovery was found 180 d after transplantation. The proportions of CD8⁺ T cells in bone marrow of the patients with hematological malignancies were significantly lower versus the healthy controls 15 and 30 d after transplantation (both P<0.01), and it recovered to normal level 60 d after transplantation. The total proportions of CD4⁺ and CD8⁺ T lymphocytes in bone marrow were both significantly lower than those in the peripheral blood of the patients with hematological malignancies (P=0.001, 0.002). The proportions of Th1 in bone marrow of the patients with hematological malignancies were significantly higher than those of the healthy controls 15, 30, 60, 90 and 180 d after transplantation (all P<0.05), and the total level was significantly higher than that in peripheral blood (P=0.006). The proportions of Th2 in bone marrow did not change significantly within 90 d after transplantation, but it was significantly higher 180 d after transplantation than that of the healthy controls (P=0.034), and the total level was similar to the total level of peripheral blood (P>0.05). The ratio of CD4⁺/CD8⁺ T lymphocytes in bone marrow was gradually decreased after transplantation, and significantly lower versus the healthy controls 180 d after transplantation (P=0.040). The ratios of Th1/Th2 in bone marrow were significantly higher than those of the healthy controls 15, 30, 60 and 90 d after transplantation (all P<0.01), while it was similar to the healthy control level 180 d after transplantation (P>0.05). The levels of IL-2R in bone marrow and peripheral blood of the patients with hematological malignancies were significantly higher than those of healthy controls 15, 30, 60 and 90 d after allo-HSCT (all P<0.05). The levels of IL-18 were significantly higher than those of healthy controls 15, 30 and 60 d after allo-HSCT (all P<0.05), and that in peripheral blood 90 d after transplantation was also significantly different from that of healthy controls (P=0.021). There were no significant differences in IL-2R or IL-18 between bone marrow and peripheral blood (both P>0.05). Conclusion After allo-HSCT, the re-constitution rules of different T lymphocyte subsets in bone marrow of patients with hematological malignancies are different, and are different from those in peripheral blood.