Empirical Study On Effect Of Reactive Nitrogen Metabolites And Their Scavengers To NK Cell-mediated Killing Of K562 Cell

[Objective]: To explore the influence of RNM(reactive nitrogen metabolites) as supressor and the effect of Tiopronin and Glutamylcysteinylglycine as immunoadjuvant to reverse RNM and ROM's suppressing influence in NK cell-mediated killing of K562 cell line.[Methods]:1.exogenous ONOO- was administered in the NK cell and K562 cell culture system,,and the changes of RNM and ROM productions and NK cell activity were observed. 2. IL-2 and PHA were administered as reactivator to increase RNM and ROM productions, and the corresponding changes of KIR and NK cell activity were observed. Then different concentrations of Tiopronin,Glutamylcysteinylglycine and Histamine dihydrochloride were administered in the cultivated systems with monocytes plus NK cells and K562 cells (E/T=10/1,E/MO=10/2). At the same time, RNM and ROM productions,KIR,IL-6,TNF-βand IFN-γproductions were measured on schedule.[Results]:1. After exogenous ONOO- was administered in the NK cell and K562 cell culture system, K562 cell population and NK cell activity degraded significantly, while RNM and ROM productions increased. And with higher more K562 cells, RNM and ROM productions increased accordingly. 2.①With more monocytes,RNM and ROM productions slightly increased accordingly in the monocyte culture system. But after the activation of IL-2/PHA, RNM production increased from 81.8898±0.7874umom/l to 91.0308±0.8956umom/l,and ROM production increased from 193.1685±9.6505U/ml to 356.4275±5.9642U/ml (P<0.05)in the MO=10 group.②After IL-2 and PHA were administered in the cultivated systems mixing with NK cells and K562 cells as the E/T(NK/K562) ratio was 10/1,RNM production increased slightly,and ROM production increased from 58.6326±2.5141U/ml to 141.915±8.2593U/m(lP<0.05) at the same time, while KIR increased from 79.98% to 93.29%(P<0.05). After monocytes being appended as E/MO(NK/monocyte) ratio was 10/2,10/5 and 10/10 ,RNM and ROM productions increased correspondingly.ButIL-6,TNF-βand IFN-γ productions and KIR changed on the other round. Controlling by K562 cells, the negative correlation coefficient of RNM to KIR exceeds that of ROM to KIR (r= -0.8511,-0.7141 respectively).③When TIP,GSH and DHT were administered in the cultivated systems mixing with NK cells,K562 cells,monocytes,IL-2 and PHA, RNM productions dropped from 73.7390±3.7908 umom/l to 64.5516±2.1570umom/l,63.0222±2.3090 umom/l,73.1052±1.4528 umom/l respectively, and ROM productions dropped from 321.5488±10.5030 U/ml to 55.1665±7.0950 U/ml,54.6136±5.9515 U/ml,108.6608±6.2110U/ml respectively,while KIR ascensused from 65.78% to 84.47%,84.77%,79.56% respectively. With higher concentration of TIP,GSH or DHT, the RNM and ROM productions decreased accordingly,while IL-6,TNF-βand IFN-γproductions as well as KIR increased at the same time. There showed reverse correlation between RNM and ROM productions and KIR( r= -0.792 and -0.933 respectively). In scavenging RNM and ROM, and enhancing the anti-neoplasm activity of NK cells,TIP is as good as GHS(P>0.05),but bothes of them are better than DHT(P<0.05).[Conclusions]:1.Exogenous ONOO- is deleterious to NK cells and K562 cells.And it could convert to ROM or/and induce cell to generate ROM. 2.Monocytes are the major resources of RNM and ROM, while K562 cells and NK cells are the minor ones. 3.Bothes of RNM and ROM can disable NK cells in killing neoplasm cells (K562 cells). 4.Bothes of TIP and GHS can protect NK cells via scavenging RNM and ROM,and enhance the anti-neoplasm activity of NK cells. At the same time ,there shows a dose-effect relationship between the doses of TIP and GHS and their capability of scavenging RNM and ROM.While DHT cannot scavenge RNM. 5. In scavenging RNM and ROM and up-regulating KIR,TIP is as good as GSH,but bothes of them are better than DHT. With less side effect, TIP and GSH may take the place of DHT as immunoadjuvant during the adoptive immunotherapy in leukemia.