**Preparation of Conditioned Medium (CM) of peripheral blood mononuclear cells cultured with phycocyanin**

130 Blood Cell – An Overview of Studies in Hematology

**cell lines** 

(Koren et al, 1979).

mature cells.

possibly contribute to the radiation protection effects.

oxygen species in mice and against reduced levels of the leukocytes in the blood and nucleated cells in the bone marrow in dog (Zhang et al., 2001; Verma et al., 2006). Doctors in Belarus reported that ingestion of 5 g of *Spirulina* a day resulted in the reduction of Cesium-137 in urine by 50%, in children subjected to low level of radiation over a long period of time (Loseva and Dardynskaya, 1993). Rahadiya and Patel in India (Rabadiya and Patel, 2010) also reported radiation-effect-reducing activity of *Spirulina* in their review. Anti-oxidant and anti-inflammatory effects as well as proliferation and differentiation activity of *Spirulina*

**5. Effect of phycocyanin on differentiation of human myeloid leukemia** 

A study of aerosolized GM-CSF demonstrated tolerance and possible efficacy in patients with malignant metastases to the lungs, possibly through upregulation of antigen-specific cytotoxic T-cells (Rao et al., 2003). It is known that various food compounds and the metabolites involving phycocyanin can influence the processes in cellular differentiation, apoptosis, and proliferative potential, and there is considerable evidence that vitamins and micronutrients are able to regulate gene expression of cancer cells, resulting in influence on the carcinogenic process (Sacha et al., 2005). All-trans-retinoic acid and vitamin D3 are known as one of the physiologic agents which can modulate the proliferation and differentiation of hematopoietic cells (Collins, 2002). The vitamin plus interferon-γ (IFNγ) treatment and enrichment with polyunsaturated fatty acids such as arachidonic acid, eicosapentaenoic acid or docosahexaenoic acid also significantly enhanced immunoregulatory effects, or enhanced the expression of monocytic surface antigens CD11b and CD14 on human premonocytic U937 cells (Obermeier et al., 1995). In this section, we investigated the effects of phycocyanin on differentiation and morphological and cytochemical changes of human myeloid leukemia cell

lines, U937 and HL-60 cells, generally used for the studies of cell differentiation.

A human hematopoietic cell line, U-937, was derived from a patient with generalized histiocytic lymphoma. The histiocytic origin of the cell line was shown by its capacity of lysozyme production and the strong esterase activity (naphtol AS-D acetate esterase inhibited by NaF) of the cells (Sundstrom and Nillson, 1976). The cell line was morphologically identical to that of the tumor cells in the pleural effusion, and is known to be functionally differentiated to phagocytic macrophage by cytokines from lymphocytes

A continuous human myeloid cell line, HL-60, was derived from the peripheral blood leukocytes of a patient with acute promyelocytic leukemia and established. The predominant cell type is a neutrophilic promyelocyte with prominent nuclear/cytoplasmic asynchrony (Gallangher et al., 1979). HL-60 cells lack specific markers for lymphoid cells, but express surface receptors for Fc fragment and complement (C3), which have been associated with differentiated granulocytes. They exhibit phagocytic activity and responsiveness to a chemotactic stimulus commensurate with the proportion of Human peripheral blood mononuclear cells (PBMCs) from 7 healthy volunteers were separated by density centrifugation (800 g, 30 mins) using a Lymphoprep™ (Density 1.077 g/mL, NYCOMED) under approving by the Institutional Review Board of our University. Cells from each subject were suspended in RPMI-FBS medium and adjusted to 1 x 106 cells/mL and were cultured with and without 2 mg/mL of phycocyanin (Phyc) using 24-well cultured plates. The conditioned mediums (CMs), both with and without Phyc (Phyco-CM and Cont-CM, respectively), were harvested on the 7th day and filtered through an 0.45 μmφ filter to remove cell debris.

Cell growth of U937 as well as HL-60 cells supplemented with Phyco-CM resulted in significant inhibition during the 7-day culture in comparison with those of control without supplementation, while supplementation with 2 mg/mL Phyc itself had no effect on growth in these cells. Both U937 and HL-60 cells stimulated with Phyc and Phyco-CM were more than 80% viable, as were those cells stimulated with phorbol-12-myristate-13-acetate (PMA) 0.02 μg/mL as a positive control of differentiation.
