Objective: To investigate the mechanisms underlying bone marrow damage by iron overload in pancytopenic patients with positive BMMNC-Coombs test (IRP).
Methods: Twenty-one iron overloading, 26 non-iron overloading IRP patients and 10 normal controls were enrolled in this study. The expressions of ROS, Bcl-2, Caspase-3 and apoptosis of BMMNC were analyzed by flow cytometry (FCM). Antioxidants were added to iron overloading IRP BMMNC, and then the changes of indices above were detected by FCM. The number and apoptosis of T lymphocytes of IRP patients were also detected.
Results: ROS and apoptosis of BMMNC, myelocytes, erythrocytes and stem cells of iron overloading IRP patients were significantly higher than that of non-iron overloading IRP ones and normal controls (P < 0.05). The expressions of Bcl-2 on BMMNC, erythrocytes and stem cells of iron overloading IRP patients were significantly lower than those of non-iron overloading IRP ones (P < 0.05). The levels of Caspase-3 on myelocytes, erythrocytes and stem cells of iron overloading IRP patients were significantly higher than those of non-iron overloading IRP ones and normal controls (P < 0.05). After treatment with antioxidants, the expressions of ROS, Caspase-3 and apoptosis of iron overloading IRP BMMNC significantly decreased, but opposite for Bcl-2. The percentages of CD4(+) lymphocytes [ ( 40.86 ± 8.74）%] and CD4(+)/CD8(+) (1.44 ± 0.36) in PB of iron overloading IRP patients were significantly higher than that of non-iron overloading IRP ones [(35.96 ± 7.03)% and 1.14 ± 0.37] and normal controls [(28.00 ± 6.73)% and 0.79 ± 0.21], respectively (P < 0.05), as opposite for CD8(+) lymphocytes (P < 0.05). The apoptosis of CD8(+) lymphocytes [(27.35 ± 10.76)%] and the ratio of CD8(+) apoptosis/CD4(+) apoptosis (2.51 ± 0.81) in BM of iron overloading IRP patients were significantly higher than those of non-iron overloading IRP ones [(15.47 ± 8.99)%] and normal controls (1.39 ± 0.47), respectively (P < 0.05). The apoptosis of erythrocytes and stem cells coated with auto-antibodies in BM of iron overloading IRP patients were significantly higher than those of non-iron overloading IRP and normal controls.
Conclusion: Mechanisms underlying bone marrow damage by iron overload might be through the follows: ①The increased ROS induced by excessive iron deposition affected the expressions of Caspase-3 and Bcl-2, which caused more BMMNC apoptosis; ②The abnormal number and ratio of T lymphocytes caused by iron overload aggravated the abnormality of immunity of IRP; ③Iron overload may increase the damage to erythrocytes and stem cells coated with auto-antibodies.
目的 探究铁过载对骨髓单个核细胞（BMMNC）Coombs试验阳性全血细胞减少症（又称免疫相关性全血细胞减少症，IRP）患者骨髓造血损伤的机制。方法 采用流式细胞术检测21例铁过载IRP患者、26例非铁过载IRP患者及10名正常对照者骨髓造血细胞活性氧自由基（ROS）、Bcl-2、Caspase-3水平及凋亡情况；并检测铁过载组BMMNC经N-乙酰半胱氨酸处理后上述指标的变化；同时检测IRP患者T细胞数量、比例及凋亡率。结果 铁过载组BMMNC、粒系、红系和干细胞ROS水平及凋亡率均明显高于非铁过载组及正常对照组（P<0.05）；铁过载组BMMNC、红系和干细胞Bcl-2水平均明显低于非铁过载组（P<0.05）；铁过载组粒系、红系和干细胞Caspase-3水平均明显高于非铁过载组及正常对照组（P<0.05）。铁过载组经N-乙酰半胱氨酸处理后，BMMNC ROS、Caspase-3水平及凋亡率均较前明显下降，Bcl-2水平较前明显升高。铁过载组外周血CD4+细胞、CD4+/CD8+细胞比值分别为（40.86±8.74）%及1.44±0.36，均明显高于非铁过载组[（35.96±7.03）%及1.14±0.37]和正常对照组[（28.00±6.73）%及0.79±0.21]（P<0.05），铁过载组CD8+细胞百分率明显低于非铁过载组和正常对照组（P<0.05）。铁过载组骨髓CD8+细胞凋亡率、CD8+细胞凋亡率/CD4+细胞凋亡率比值分别为（27.35±10.76）%及2.52±0.81，均明显高于非铁过载组[（15.47±8.99）%及1.39±0.47]（P<0.05）。在骨髓有核红细胞膜抗体或干细胞膜抗体阳性IRP患者中，铁过载组骨髓红系或干细胞凋亡率均高于非铁过载组（P<0.05）。结论 铁过载可能通过以下机制影响IRP患者骨髓造血功能：①过量铁沉积可诱导骨髓造血细胞ROS生成增加，影响Bcl-2、Caspase-3凋亡信号的表达，引起IRP患者骨髓造血细胞凋亡增加；②铁过载可通过影响T细胞数量及比例加重IRP患者免疫异常；③铁过载可增加IRP患者被覆自身抗体的骨髓有核红细胞和干细胞的破坏。