Abstract
Background:
Several attempts have been made to expand human NK cells from peripheral blood mononuclear cells (PBMCs). This study examined the selective expansion of NK cells using interleukin 2 (IL-2) plus the K562 cell line, the expression of the NK cell receptors, and the cytotoxic activity.
Methods:
The PBMCs from seven healthy volunteers were cultured in a medium containing the IL-2 plus the K562 cell line for 14 days. The expression of the activating and inhibitory receptors on the resting NK cells and the 72 hr-expanded NK cells were analyzed. A flow cytometric cytotoxic assay was used to determined the killing activity of the non-expanded NK cells and the 7 day-expanded NK cells against the K562 target cells.
Results:
The NK cells from PBMCs expanded 4.5-fold after 7 days, and contained 56.5% CD3-CD56+ cells. The IL-2 or IL-2 plus K562 increased the expression levels of CD158b (MFI, mean florescence intensity), CD158e1/e2 (MFI), and NKp44 (MFI), while it decreased the expression levels of NKp30 (%), CD16 (MFI), and 2B4 (MFI). The non-expanded NK cells lysed 9.0% and 27.6% of the K562 target cells in the 1:1 and 5:1 effector and target ratio, respectively, and the 7-day expanded NK cells lysed 36.9% and 57.2% of the K562 target cells, respectively.
Conclusion:
The selective expansion of CD3-CD56+ NK cells occurred only during 7 days of culture. IL-2 or IL-2 plus the K562 cells altered the expression of various activating and inhibitory receptors of NK cells, and the cytotoxicity of the expanded NK cells was higher than in the non-expanded cells.
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Fig. 1
The change of CD158b expression on gated CD3- CD56+ NK cells in response to IL-2 alone and IL-2 plus K562 cell line. Representative data using PE-labeled CD 158b was shown. The increased expression was observed for CD 158b receptors, indicating that IL-2 alone (B) and IL2+K562 (C) induce up-regulation of CD158b compared to uncultured cells (A).
![kjh-41-8f1.tif](/upload/SynapseXML/0072kjh/thumb/kjh-41-8f1.gif)
Fig. 2
Flow cytometric measurement of NK cytotoxicity using staining with FITC-congugated anti-CD45 and side scatter profiles to identify different cell populations and uptake of propidium iodide (PI) to detect cell death. Target cells and effectors (1:5) incubated for 4 hrs, are identified by region 1 (R1) and region 2 (R2) repectively. Percentages of dead cells in different regions are calculated from histograms showing PI uptake as measured by the intensity of fluorescence.
![kjh-41-8f2.tif](/upload/SynapseXML/0072kjh/thumb/kjh-41-8f2.gif)
Table 1.
Median cell count, percentage of CD3-CD56+ cells, and fold expansion rate from peripheral blood mononuclear cells of seven donors. The cells were cultured in IL-2 (500U/mL) and irradiated K562 cell line as a feeder cell
Table 2.
The change of activating and inhibitory natural killer cell receptors when lymphocytes are treated with IL-2 alone or IL-2 with irradiated K562 cell line as feeder cell