Effects Of Low-dose Radiation On Immunological Competence Of NK Cells From The Peripheral Blood In Patients With Tumor And Mechanism

Background and purpose：In the normal cells， low-dose radiation (LDR) has both hormesis and adaptiveresponse. At cellular level, the hormesis mainly embodies in accelerating theproliferation and enhancing immune activity of normal cells. And low-dose radiationcan reduce the damage which is caused by the follow-up of high-dose radiation, this isadaptive response. However, in the tumor cells, LDR can inhibit the growth of tumorcells and enhance the killing effect of high-dose radiation on the tumor cells. LDRused in cancer therapy, not only could protect the body normal tissue, but alsoincrease the doses of cancer radiotherapy and chemotherapy, thereby enhancing theeffect of cancer treatment, which will explore a new mode for the comprehensivetherapy of cancer. The cell immunotherapy of cancer is a therapy in which theautologous or xenogenous anti-tumor effector cells from the patients with cancer areinduced, activated and amplified by cytokines in vitro, and then reinfusion back intothe patient, directly or indirectly killing malignant tumors cells, and in order toimprove the ability of anti-tumors. However, whether the activity of immune cells isenhanced and the efficacy of adoptive cellular immunotherapy is improved whenLDR acting on the immune cells, and the molecular mechanism in whichLDR-induced immune-enhancing effect, have not been fully elucidated yet. The studyon the effects of LDR on immunological competence of immune cells and itsmechanism, will provide a theroretical basis for the treatment of LDR in cancer.In the present study, the effect of LDR on immune activity of NK cells fromperipheral blood of patients with malignant tumors was reaserched, the changes inkilling activity, cytokine and protein expression were detected, and its mechanismfrom the perspective of p38MAPK signal transduction pathway was studied. Method:First of all, we isolated and cultured NK cells from peripheral blood of24patients with malignant tumors, counted the number of these cells daily and observedthe morphological changes before and after culture by inverted microscope, anddetected the changes of surface features CD3－CD56＋NK cell percentage using flowcytometry, to realize the expansion capacity of NK cells in vitro. Second, lactatedehydrogenase cytotoxicity assay was used to determinate changes of killing activityof NK cells after different doses of radiation (25,75,200and500mGy). We detectedthe secretion levels of cytokine TNF-α及IFN-γ in the culture system supernatant byELISA test kit. After75mGy radiation, the expression levels of anti-associatedprotein FasL, perforin, and p38MAPK pathway were determinated using Westernblot.Results:1. After induction and expansion for14days in vitro, NK cells expanded110folds, suggesting that NK cells from the peripheral blood in patients with malignanttumors can be induced and amplified.2. LDR-induced NK cells killing activity was significantly enhanced, especiallyafter75mGy radiation and cultured for24h, the strongest killing activity achieved79.33%.3. After LDR, cytokines TNF-α and IFN-γ secretion levels in NK cells culturesystem supernatant were elevated, The highest secretion levels were found after75mGy radiation and cultured for24h. The highest secretion level of TNF-α was(30.57±0.79) pg/ml, and IFN-γ reached (15.61±0.53) pg/ml.4. After NK cells received75mGy radiation and culcured for24h, expressionlevels of anti-associated protein FasL and perforin were significantly increased.5. Compared with the non-irradiated group, after LDR, the secretion levels ofNK cells destruction-related cytokines TNF-α and IFN-γ, and protein expressions ofp38MAPK, FasL and perforin were significantly increased (p<0.05), and showedpositively correlated with the cytotoxic activity of NK cells. After p38MAPK pathway activity was inhibited, the cytotoxic activity of NK cells, cytokines TNF-α,IFN-γ levels, and protein expressions of p38MAPK pathway, FasL and perforin weremarkedly reduced.Conclusions:1. Low-dose radiation induces NK cells from the peripheral blood in patientswith malignant tumors to enhance their immune activity.2. The mechanism of LDR induced immune-enhancing effects may be closelyrelated to activation of p38MAPK signaling pathway.The chief innovation of the present study is that after NK cells are induced byLDR in vitro, their stronger immune activity is found. The mechanism ofLDR-induced immune-enhancing effect of NK cells may be related to the expressionof p38MAPK signaling pathway.