Tehran University Medical Journal

---

Natural Killer (NK) cells are the main lymphocyte population expressing P75 B chain of the IL-2 receptor (IL-2R). Consequently, incubation of peripheral blood lymphocytes with IL-2 induce selective activation of NK cells and results in NK activity and generation of Lymphokine activated killer (LAK) cells activity and proliferation. One of the early events during IL-2 activation of peripheral blood lymphocyte in both rodents and humans is adherence of some NK cells to plastic surface. The cells adherence to plastic after 24 hr of culture with IL-2 are almost exclusively CD56+, have the morphology large granular cells to yield a highly entiched population of activated NK cells that have been used for systemic adoptive immunotherapy. To test these hypothesis, we used highly purified population of human peripheral NK cells through the biological and nonimmunclogical phenotyping technique. Blood mononuclear cells were separated by centrifugation of ficol-hypaque gradient from normal blood donor (20-30 years age). We depleted after purification of nonadherent cells with nylonwool. We collected with rosette technique to remove cells with high affinity SRBC receptors. These cells separate in two parts A-NK and NA-NK by mononuclear celss activated supernatant media. The main objective results of this study show that the subpopulation of human NK cell which develope early adherent to plastic surface in the presence of supernatant mononuclear celss activation media was functionally more cytotoxic and killed K562 targets in single cell sytotoxicity manner and LDH activity assay than nonadherent NK cells and resting NK cells

---

Introduction: PMA is known to induce the differentiation of monocytes to macrophages. This agent also increases the killing effect of the monocytes/macrophages through oxidative burst and can be used as a stimulant for oxidative burst assay. The present experimental study was intended to investigate the in vitro effects of PMA on the differentiation, morphological changes, cell adherence and the viability of monocyted-derived macrophages (MDMs). Besides, MDM capacity for free radical production was assessed, indicating the oxidative burst events.

Materials and Methods: This experimental study has been design in Department of Immunology of S.B.M.U in Tehran Iran (year 2000). Peripheral mononuclear cells from adult Balb/c mice were isolated and cultured in complete tissue culture medium and divided in two group: control, (without PMA) and test (were added Pma=450 ng/ml). MDMs wee counted on the hours 1, 2, 3, 4, 6 and 18 and their characteristics were confirmed by morphological analysis (histological features) in both groups. Viability of MDMs was assessed using trypan blue. In the peak time of MDMs activation the oxidative burst was determined by NBT reduction.

Results: The obtained results suggested that PMA had significant effect on the differentiation of monocytes to macrophages. The morphologic maturation tended to occur in earlier stages in the PMA treated cells comparing to the control MDMs. Also, the number of adherent cells was considerably more in PMA stimulated monocytes. The peak time of cell adherence in the presence of PMA was no the second hour. As the incubation period increased, the significant difference between the numbers of adherent cells in two culture systems decreased. However, viability decreased significantly in the PMA treated MDMs, i.e. PMA treatment induced rapid apoptosis in the MDMs after their activation. PMA stimulated MDMs markedly (60%). Also we mentioned that the primary un-stimulated MDMs only revealed (55%) of NBT reduction after treatment with PMA at NBT reduction stage.

Conclusion: Phagocytic function and oxidative burst assay in monocyte-macrophage lineage can be a diagnostic tool for identification and management of some Immunological abnormality and defect and can be establish distinct from other phagocytic system assessment.