中华胰腺病杂志
中華胰腺病雜誌
중화이선병잡지
CHINESE JOURNAL OF PANCREATOLOGY
2013年
3期
179-182
,共4页
薛成俊%周忠海%陈复兴%吕小婷%李莹%费素娟
薛成俊%週忠海%陳複興%呂小婷%李瑩%費素娟
설성준%주충해%진복흥%려소정%리형%비소연
氢化可的松%杀伤细胞,天然%胰腺肿瘤%细胞增殖%杀伤活性
氫化可的鬆%殺傷細胞,天然%胰腺腫瘤%細胞增殖%殺傷活性
경화가적송%살상세포,천연%이선종류%세포증식%살상활성
Hydrocortisone%Killer cell,natural%Pancreatic neoplasms%Cell proliferation%Killing activity
目的 探讨氢化可的松对人外周血NK细胞增殖及其对胰腺癌SW1990细胞杀伤力的影响.方法 分离健康人外周血单个核细胞加入到含IL-15细胞因子的NK细胞培养基中诱导培养NK细胞.当NK细胞纯度达到70％以上时,加入10-6、10-5、10-4、10-3 μmol/L氢化可的松继续培养7d.以未加氢化可的松的NK细胞作为对照组.采用锥虫蓝染色计数细胞；采用流式细胞术检测CD3-CD56+ NK细胞含量及其穿孔素、颗粒酶B和IFN-γ的表达；以20∶1的效靶比将NK细胞与SW1990细胞共培养,用细胞增殖-毒性检验法(CCK-8)检测NK细胞对SW1990细胞的杀伤效应.结果 用氢化可的松处理7d后NK细胞量平均达到70.72％ ～ 76.39％,与对照组的(72.61±3.76)％差异无统计学意义；10-6、10-5、10-4、10-3 μmol/L氢化可的松处理组NK细胞的增殖倍数分别为(9.13±0.94)、(9.67±1.51)、(10.33± 1.07)、(8.40±1.47)倍,均显著高于对照组的(4.23±0.82)倍(P值均＜0.01)；NK细胞对SW1990细胞的杀伤效率分别为(58.58±4.89)％、(62.27±5.63)％、(64.02±5.79)％、(63.88±3.61)％,均较对照组的(57.46±5.11)％增强,其中10-4μmol/L组的差异具有统计学意义(P ＜0.05)；10-4、10-3μmol/L氢化可的松处理组NK细胞穿孔素表达分别为(96.71±3.04)％、(97.56±2.18)％,显著高于对照组的(92.40±3.53)％(P ＜0.05或0.01)；10-5 μmol/L氢化可的松处理组NK细胞的颗粒酶B表达为(78.23±2.94)％,显著高于对照组(73.68±3.52)％(P＜0.05)；10-5、10-4、10-3 μmol/L氢化可的松处理组NK细胞IFN-γ表达分别为(96.61±2.04)％、(97.58±2.17)％、(98.00±1.77)％,均显著高于对照组的(92.44±2.74)％(P值均＜0.01).结论 氢化可的松可促进IL-15活化的NK细胞增殖,适当浓度条件下增强对SW1990细胞杀伤活性,其机制可能与其穿孔素、颗粒酶B和IFN-γ表达上调有关.
目的 探討氫化可的鬆對人外週血NK細胞增殖及其對胰腺癌SW1990細胞殺傷力的影響.方法 分離健康人外週血單箇覈細胞加入到含IL-15細胞因子的NK細胞培養基中誘導培養NK細胞.噹NK細胞純度達到70％以上時,加入10-6、10-5、10-4、10-3 μmol/L氫化可的鬆繼續培養7d.以未加氫化可的鬆的NK細胞作為對照組.採用錐蟲藍染色計數細胞；採用流式細胞術檢測CD3-CD56+ NK細胞含量及其穿孔素、顆粒酶B和IFN-γ的錶達；以20∶1的效靶比將NK細胞與SW1990細胞共培養,用細胞增殖-毒性檢驗法(CCK-8)檢測NK細胞對SW1990細胞的殺傷效應.結果 用氫化可的鬆處理7d後NK細胞量平均達到70.72％ ～ 76.39％,與對照組的(72.61±3.76)％差異無統計學意義；10-6、10-5、10-4、10-3 μmol/L氫化可的鬆處理組NK細胞的增殖倍數分彆為(9.13±0.94)、(9.67±1.51)、(10.33± 1.07)、(8.40±1.47)倍,均顯著高于對照組的(4.23±0.82)倍(P值均＜0.01)；NK細胞對SW1990細胞的殺傷效率分彆為(58.58±4.89)％、(62.27±5.63)％、(64.02±5.79)％、(63.88±3.61)％,均較對照組的(57.46±5.11)％增彊,其中10-4μmol/L組的差異具有統計學意義(P ＜0.05)；10-4、10-3μmol/L氫化可的鬆處理組NK細胞穿孔素錶達分彆為(96.71±3.04)％、(97.56±2.18)％,顯著高于對照組的(92.40±3.53)％(P ＜0.05或0.01)；10-5 μmol/L氫化可的鬆處理組NK細胞的顆粒酶B錶達為(78.23±2.94)％,顯著高于對照組(73.68±3.52)％(P＜0.05)；10-5、10-4、10-3 μmol/L氫化可的鬆處理組NK細胞IFN-γ錶達分彆為(96.61±2.04)％、(97.58±2.17)％、(98.00±1.77)％,均顯著高于對照組的(92.44±2.74)％(P值均＜0.01).結論 氫化可的鬆可促進IL-15活化的NK細胞增殖,適噹濃度條件下增彊對SW1990細胞殺傷活性,其機製可能與其穿孔素、顆粒酶B和IFN-γ錶達上調有關.
목적 탐토경화가적송대인외주혈NK세포증식급기대이선암SW1990세포살상력적영향.방법 분리건강인외주혈단개핵세포가입도함IL-15세포인자적NK세포배양기중유도배양NK세포.당NK세포순도체도70％이상시,가입10-6、10-5、10-4、10-3 μmol/L경화가적송계속배양7d.이미가경화가적송적NK세포작위대조조.채용추충람염색계수세포；채용류식세포술검측CD3-CD56+ NK세포함량급기천공소、과립매B화IFN-γ적표체；이20∶1적효파비장NK세포여SW1990세포공배양,용세포증식-독성검험법(CCK-8)검측NK세포대SW1990세포적살상효응.결과 용경화가적송처리7d후NK세포량평균체도70.72％ ～ 76.39％,여대조조적(72.61±3.76)％차이무통계학의의；10-6、10-5、10-4、10-3 μmol/L경화가적송처리조NK세포적증식배수분별위(9.13±0.94)、(9.67±1.51)、(10.33± 1.07)、(8.40±1.47)배,균현저고우대조조적(4.23±0.82)배(P치균＜0.01)；NK세포대SW1990세포적살상효솔분별위(58.58±4.89)％、(62.27±5.63)％、(64.02±5.79)％、(63.88±3.61)％,균교대조조적(57.46±5.11)％증강,기중10-4μmol/L조적차이구유통계학의의(P ＜0.05)；10-4、10-3μmol/L경화가적송처리조NK세포천공소표체분별위(96.71±3.04)％、(97.56±2.18)％,현저고우대조조적(92.40±3.53)％(P ＜0.05혹0.01)；10-5 μmol/L경화가적송처리조NK세포적과립매B표체위(78.23±2.94)％,현저고우대조조(73.68±3.52)％(P＜0.05)；10-5、10-4、10-3 μmol/L경화가적송처리조NK세포IFN-γ표체분별위(96.61±2.04)％、(97.58±2.17)％、(98.00±1.77)％,균현저고우대조조적(92.44±2.74)％(P치균＜0.01).결론 경화가적송가촉진IL-15활화적NK세포증식,괄당농도조건하증강대SW1990세포살상활성,기궤제가능여기천공소、과립매B화IFN-γ표체상조유관.
Objective To investigate the effects of hydrocortisone (HC) on proliferation and killing activity of NK cells against pancreatic cancer SW1990 cells in vitro.Methods Peripheral blood mononuclear cells of healthy people were isolated and cultured with NK cells medium containing IL-1S.When the purity of NK cells reached above 70％,different concentrations of HC (10-6,10-5,10-4,10-3 μmol/L) were added and co-cultured with NK cells for 7 days.And NK cells without HC were used as control.CD3-CD56 + NK cell numbers of each group were countered by trypan blue staining.Perforin,granzyme B and IFN-γ expression of CD3-CD56+ + NK cells were verified by flow cytometry.NK cells and SW1990 cells were co-cultured with a 20∶1effector to target ratio,then the cytotoxic activity of NK cells against SW1990 cells were analyzed by CCK-8 kit.Results After treatment with different concentration of HC for 7 days,NK cells purity of each group reached 70.72％ ～ 76.39％,and it was not significantly different with that in control group [(72.61 ± 3.76) ％].The proliferation folds of NK cells treated with 10-6,10-5,10-4,10-3 μmol/L HC were (9.13 ± 0.94),(9.67 ±1.51),(10.33±1.07),(8.40±1.47) times,respectively,while it was (4.23 ±0.82) times in control group (all P ＜0.01).The killing effects of NK cells on SW1990 cells were (58.58 ± 4.89) ％,(62.27 ± 5.63) ％,(64.02 ± 5.79) ％,(63.88 ± 3.61) ％,which were higher than that in control group [(57.46 ± 5.11) ％],moreover,the difference between NK cells of 10-4 μmol/L HC treatment group and control group was statistically significant(P ＜ 0.05).The expressions of perforins of 10-4,10-3 μmol/L HC treatment group were (96.71 ± 3.04) ％,(97.56 ± 2.18) ％,which were significantly higher than that in control group [(92.40 ±3.53)％,P ＜0.05 or 0.01].The expression of granzyme B in 10-5 μmol/L HC treatment group was (78.23 ±2.94)％,which were significantly higher than that in control group [(73.68 ±3.52) ％,P ＜0.05].The expressions of IFN-γ in 10-5,10-4,10-3 μmol/L HC treatment group were (96.61 ±2.04)％,(97.58 ± 2.17)％,(98.00 ± 1.77)％,which were significantly higher than that in control group [(92.44 ± 2.74)％,P＜0.01].Conclusions HC can promote IL-15 activated NK cells proliferation and enhance NK cells mediated killing activity against SW1990 cells with proper concentration,and up-regulation of perforin,granzyme B and IFN-γ expression may be the main mechanisms.
