White blood cell (WBC) segmentation, which is important for cytometry, is a challenging issue because of the morphological diversity of WBCs and the complex and uncertain background of blood smear images. This paper proposes a novel method for the nucleus and cytoplasm segmentation of WBCs for cytometry. A color adjustment step was also introduced before segmentation. Color space decomposition and k-means clustering were combined for segmentation. A database including 300 microscopic blood smear images were used to evaluate the performance of our method. The proposed segmentation method achieves 95.7% and 91.3% overall accuracy for nucleus segmentation and cytoplasm segmentation, respectively. Experimental results demonstrate that the proposed method can segment WBCs effectively with high accuracy.

PD1 blockade enhances cytotoxicity of in vitro expanded natural killer cells towards myeloma cells

This paper presents a novel method for segmentation of white blood cells (WBCs) in peripheral blood and bone marrow images under different lights through mean shift clustering, color space conversion and nucleus mark watershed operation (NMWO). The proposed method focuses on obtaining seed points. First, color space transformation and image enhancement techniques are used to obtain nucleus groups as inside seeds. Second, mean shift clustering, selection of the C channel component in the CMYK model, and illumination intensity adjustment are employed to acquire WBCs as outside seeds. Third, the seeds and NMWO are employed to precisely determine WBCs and solve the cell adhesion problem. Morphological operations are further used to improve segmentation accuracy. Experimental results demonstrate that the algorithm exhibits higher segmentation accuracy and robustness compared with traditional methods.

Disulfiram (DSF) is an FDA approved anti-alcoholism drug in use for more than 60 years. Recently, antitumor activity of the DSF/copper (DSF/Cu) complex has been identified. Its anti-multiple myeloma activity, however, has barely been investigated. In the present study, our results demonstrated that the DSF/Cu complex induced apoptosis of MM cells and MM primary cells. The results indicated that DSF/Cu significantly induced cell cycle arrest at the G2/M phase in MM.1S and RPMI8226 cells. Moreover, JC-1 and Western blot results showed that DSF/Cu disrupted mitochondrial membrane integrity and cleaved caspase-8 in MM cells, respectively, suggesting that it induced activation of extrinsic and intrinsic apoptosis pathways. Interestingly, DSF/Cu induced caspase-3 activation was partly blocked by Z-VAD-FMK (zVAD), a pan-caspase inhibitor, indicating at caspase-dependent and -independent paths involved in DSF/Cu induced myeloma cell apoptosis machinery. Additionally, activation of the c-Jun N-terminal kinase (JNK) signaling pathway was observed in DSF/Cu treated MM cells. More importantly, our results demonstrated that DSF/Cu significantly reduced tumor volumes and prolonged overall survival of MM bearing mice when compared with the controls. Taken together, our novel findings showed that DSF/Cu has potent anti-myeloma activity in vitro and in vivo highlighting valuable clinical potential of DSF/Cu in MM treatment.

Multiple myeloma (MM) is still an incurable plasma cell tumor. Natural killer (NK) cells are characterized by efficient anti-tumor activity, and their activity is one basis of cancer immunotherapeutic strategies. Tim-3, one of the immune checkpoint molecules, negatively regulates NK cell activity. To evaluate roles of the Tim-3 pathway blocking in the regulation of NK cell mediated- anti-MM activity in vitro and in vivo, anti-Tim-3 and/or anti-its ligand (HMGB1, CEACAM1 or Galetin-9) antibodies were applied respectively to block the Tim-3 pathway in the present study. Our results showed that Tim-3 was highly expressed on NK cells, in particular on in vitro expanded NK (exNK) cells. NK cells with Tim-3 blockade displayed a significantly higher degranulation and cytolytic activity against both human MM cell lines and primary MM cells, compared to the isotype control antibody-treated NK cells. The increased NK cell cytolytic activity by Tim-3 blocking was associated with up-regulation of cytotoxicity-related molecules, including perforin, granzyme B, TNF-α and IFN-γ. Ligand (HMGB1, CEACAM1 or Galetin-9) expression on MM cells was at different levels, and accordingly, the improvement in NK cell-mediated killing activity by different ligand blocking were also varying. Tim-3 blocking showed much more efficient enhancement of NK cell cytolytic activity than its ligand blockings. More importantly, exNK cells with Tim-3 blockade significantly inhibited MM tumor growth and prolonged the survival of MM-bearing NOD/SCID mice. Our results also showed that NK cells from peripheral blood and bone marrow of MM patients expressed much higher levels of Tim-3 than their counterparts from controls. Taken together, Tim-3 may be an important target molecule used for developing an antibody and/or NK cell based immunotherapeutic strategies for MM.

Hemophagocytic lymphohistiocytosis in adults is largely underdiagnosed. To improve the rate and accuracy of diagnosis in adults, the clinical and laboratory characteristics of hemophagocytic lymphohistiocytosis were analyzed in and compared between adults and children in a Chinese cohort. Data from 50 hemophagocytic lymphohistiocytosis patients, including 34 adults and 16 children who fulfilled the 2004 hemophagocytic lymphohistiocytosis diagnostic criteria, were collected and analyzed. 1. Etiological factors: The proportion of Epstein-Barr virus infection was lower in adults compared with children, whereas fungal infection and natural killer/T cell lymphoma were more frequent in adults (P19 mmol/L and lactate dehydrogenase >500 U/L compared with adults (P<0.05). 3. The time interval from the onset of symptoms to clinical diagnosis was significantly shorter in pediatric patients than in adults (P<0.05). Certain clinical features were different between the two groups. The less characteristic clinical presentation of hemophagocytic lymphohistiocytosis in adults may make the disease more difficult to diagnose. Our findings suggest that hemophagocytic lymphohistiocytosis should be considered when an adult patient presents with the above-mentioned symptoms.