Effect Of Lentinan On Killing Of Lung Cancer A549 Cells By Differentiation Of ??T Cells In Vitro

Objective After ??T cells were isolated from healthy adult peripheral blood,they were induced to differentiate in vitro by various cytokines,and then formed into cell systems,and then different concentrations of Lentinus edodes polysaccharide(Lentinan,LNT)were added to determine the effect of lentinan on the anti-tumor activity of ??T cells,to provide a theoretical basis for clinical application of lentinan to improve the efficacy of adoptive immunotherapy of ??T cells.Methods Mononuclear cells were obtained from peripheral blood of healthy adults by density gradient centrifugation under sterile conditions,induced and increase ??T cells by cytokines such as recombinant human IL-2(1000 IU/ml)and zoledronic acid(5 umol/l).??T cells were counted on days 0,4,7,and 10 of in vitro culture by direct cell counting.The ??T cells on the 10 th day of in vitro culture were divided into 12 groups,and each group was set up with 3 duplicate wells,of which 6 groups were experimental groups,and 1640 culture solution dissolved lentinan was injected to make the final concentration of 0ug/ml,5ug/ml,10ug/ml,25ug/ml,50ug/ml,75ug/ml,recorded as LNT-I group,LNT-II group,LNT-III group,LNT-IV Group,LNT-V group,LNT-VI group,the remaining 6 groups were control group,recorded as LNT-I* group,LNT-II* group,LNTIII* group,LNT-IV* group,LNT-V* group,LNT-VI* group,then,the cells were inoculated in a 96-well plate according to the concentration gradient of lentinan,and the cells were further added,after 72 hours,the target cells were added to the cells for 24 hours,and the absorbance at 490 nm of each well of the 96-well plate was measured by an enzyme-linked immunosorbent assay(absorbance,A)Value,and the results were recorded.The cytotoxic activity of ??T cells induced by the concentration of each drug(lentipolysaccharide)in each sample was determined,and then analyzed by spss statistical software.Results(1)On the10 th day of culture,the expression of Anti-TCR-?? surface markers of ??T cells was detected by flow cytometry(41.30±12.80)%,the expression of CD44+ surface markers was(39.93±5.68)%,and the expression of CD107a+ surface markers was(18.46±4.95)%,the CD3+ surface marker was expressed as(96.15±0.49)%.(2)The ??T cells on the 10 th day ofculture were grouped and added to different concentrations of lentinan for 72 hours,and then added to the target cells for 24 hours,the ??T cells of each experimental group showed significant differences in the killing activity against lung cancer A549 cells,there was no significant difference in the killing activity between the LNT-I group and the LNT-V group(P>0.05),the difference between the other experimental groups was extremely statistically significant(P<0.001).(3)The killing activities of the LNT-I group(0 ug/ml),LNT-II group(5ug/ml),LNT-III group(10 ug/ml),LNT-IV group(25 ug/ml),LNT-Vgroup(50ug/ml)and LNT-VI group(75ug/ml)were respectively(34.70±2.14)%,(54.04±5.27)%,(90.73±3.68)%,(67.86±2.90)%,(40.14±2.33)%and(14.76±2.13)%.Conclusion:Different concentrations of lentinan-induced ??T cells have a killing effect on lung cancer A549 cells,LNT-II group(5ug/ml),LNT-III group(10 ug/ml),LNT-IV group(25 ug/ml)concentration Compared with LNT-I(0 ug/ml)group,??T cells induced by lentinan were significantly enhanced in lung cancer A549 cells(P<0.001),LNT-II group(5 ug/ml)and LNT-III group,the differences between the LNT-II group(5 ug/ml),the LNT-III group(10 ug/ml),and the LNT-IV group(25ug/ml)were extremely statistically significant,and the killing activity was dosedependent;There was no significant difference in the killing effect of ??T cells induced by lentinan in LNT-V group(50 ug/ml)compared with LNT-I(0 ug/ml)group(P>0.05);LNT-VI group(75 ug/ml)concentration group compared with LNT-I(0 ug/ml)concentration group,the killing effect of ??T cells induced by lentinan on lung cancer A549 cells was significantly decreased(P<0.001).In summary,lentinan can enhance the killing effect of ??T cells on lung cancer A549 cells within a certain concentration range,if it exceeds this range,it may inhibit its killing effect,this experiment suggests that lentinan can be used for clinical ??T cell adoptive immunization,however,it is necessary to determine the safe dose range,the effective dose range and the optimal dose through more clinical experiments to benefit tumor patients.