Barnea Itay, Luria Lior, Girsault Arik, Dabah Ofira, Dudaie Matan, Mirsky Simcha K, Merkel Drorit, Shaked Natan T
Department of Biomedical Engineering, Tel Aviv University, Tel Aviv 6997801, Israel.
MDS Center, Sheba Medical Center, Ramat Gan 5266202, Israel.
Bioengineering (Basel). 2024 Mar 6;11(3):256. doi: 10.3390/bioengineering11030256.
Myelodysplastic syndromes (MDSs) are a group of potentially deadly diseases that affect the morphology and function of neutrophils. Rapid diagnosis of MDS is crucial for the initiation of treatment that can vastly improve disease outcome. In this work, we present a new approach for detecting morphological differences between neutrophils isolated from blood samples of high-risk MDS patients and blood bank donors (BBDs). Using fluorescent flow cytometry, neutrophils were stained with 2',7'-dichlorofluorescin diacetate (DCF), which reacts with reactive oxygen species (ROS), and Hoechst, which binds to DNA. We observed that BBDs possessed two cell clusters (designated H and L), whereas MDS patients possessed a single cluster (L). Later, we used FACS to sort the H and the L cells and used interferometric phase microscopy (IPM) to image the cells without utilizing cell staining. IPM images showed that H cells are characterized by low optical path delay (OPD) in the nucleus relative to the cytoplasm, especially in cell vesicles containing ROS, whereas L cells are characterized by low OPD in the cytoplasm relative to the nucleus and no ROS-containing vesicles. Moreover, L cells present a higher average OPD and dry mass compared to H cells. When examining neutrophils from MDS patients and BBDs by IPM during flow, we identified ~20% of cells as H cells in BBDs in contrast to ~4% in MDS patients. These results indicate that IPM can be utilized for the diagnosis of complex hematological pathologies such as MDS.
骨髓增生异常综合征（MDS）是一组可能致命的疾病，会影响中性粒细胞的形态和功能。MDS的快速诊断对于启动能够大幅改善疾病预后的治疗至关重要。在这项工作中，我们提出了一种新方法，用于检测从高危MDS患者和血库供体（BBD）的血液样本中分离出的中性粒细胞之间的形态差异。使用荧光流式细胞术，中性粒细胞用与活性氧（ROS）反应的二乙酸2',7'-二氯荧光素（DCF）和与DNA结合的Hoechst进行染色。我们观察到BBD有两个细胞簇（分别命名为H和L），而MDS患者只有一个簇（L）。后来，我们使用荧光激活细胞分选术（FACS）对H细胞和L细胞进行分选，并使用干涉相显微镜（IPM）对细胞进行成像，无需进行细胞染色。IPM图像显示，H细胞的特征是细胞核相对于细胞质的光程延迟（OPD）较低，特别是在含有ROS的细胞小泡中，而L细胞的特征是细胞质相对于细胞核的OPD较低且没有含ROS的小泡。此外，与H细胞相比，L细胞的平均OPD和干质量更高。在流动过程中通过IPM检查MDS患者和BBD的中性粒细胞时，我们发现BBD中约20%的细胞为H细胞，而MDS患者中这一比例约为4%。这些结果表明，IPM可用于诊断诸如MDS等复杂的血液学疾病。
