中华地方病学杂志
中華地方病學雜誌
중화지방병학잡지
Chinese Journal of Endemiology
2015年
11期
785-789
,共5页
氟化物中毒%小胶质细胞%细胞核因子κB
氟化物中毒%小膠質細胞%細胞覈因子κB
불화물중독%소효질세포%세포핵인자κB
Fluoride poisoning%Microglial%Nuclear factor κB
目的 观察氟对小胶质细胞炎症因子表达及细胞核因子κB(NF-κB)信号通路的影响.方法 以体外培养人急性单核细胞白血病细胞(THP-1)作为小胶质细胞模型,用不同浓度[0(阴性对照组)、10、50、100、500、1 000、5 000 μmol/L]氟化钠(NaF)培养48 h,CCK8法检测细胞存活率.根据CCK8法检测结果,将THP-1细胞分为3组:对照组和低、高剂量染氟组,分别用0、500、5 000μmol/L NaF染氟处理48 h,以酶联免疫吸附实验(ELISA)检测细胞培养液上清中炎性细胞因子白细胞介素(IL)-1β和肿瘤坏死因子(TNF)-α,蛋白质免疫印迹(Western blot)法检测培养细胞IκBo、Phospho-NF-κB p65 (P-P65)和Phospho-IκB-α(P-I κBα)蛋白表达水平.结果 500、1 000、5 000 μmol/L染氟组THP-1细胞存活率[(73.21±3.67)％、(31.40±4.56)％、(0.40±0.24)％]明显低于阴性对照组[(100.00±0.00)％,P均＜0.01].低、高剂量染氟组炎性因子IL-1β[(1.42±0.79)、(19.47±2.90)ng/L]、TNF-α含量[(61.06±2.20)、(172.72±2.29)ng/L]均高于对照组[(0.36±0.07)、(31.07±0.81)ng/L,P均＜0.05];高剂量染氟组IκBα蛋白表达水平[(63.53±9.67)％]低于对照组[(100.00±10.99)％,P＜ 0.01];低、高剂量染氟组P-P65蛋白表达水平[(113.71±8.99)％、(134.74±1.93)％]明显高于对照组[(100.00±5.48)％,P均＜0.05];低、高氟剂量组P-IκBα蛋白表达水平[(152.61±14.16)％、(176.91±7.95)％]高于对照组[(100.00±14.82)％,P均＜0.01].线性回归分析表明,TNF-α和IL-1β含量与P-P65蛋白表达水平之间呈线性正相关(r=0.74、0.72,P均＜0.05).结论 过量氟可诱导小胶质细胞产生炎性因子并激活NF-κB信号通路,炎性因子释放及信号通路的激活可能是氟导致中枢神经系统损伤的机制之一.
目的 觀察氟對小膠質細胞炎癥因子錶達及細胞覈因子κB(NF-κB)信號通路的影響.方法 以體外培養人急性單覈細胞白血病細胞(THP-1)作為小膠質細胞模型,用不同濃度[0(陰性對照組)、10、50、100、500、1 000、5 000 μmol/L]氟化鈉(NaF)培養48 h,CCK8法檢測細胞存活率.根據CCK8法檢測結果,將THP-1細胞分為3組:對照組和低、高劑量染氟組,分彆用0、500、5 000μmol/L NaF染氟處理48 h,以酶聯免疫吸附實驗(ELISA)檢測細胞培養液上清中炎性細胞因子白細胞介素(IL)-1β和腫瘤壞死因子(TNF)-α,蛋白質免疫印跡(Western blot)法檢測培養細胞IκBo、Phospho-NF-κB p65 (P-P65)和Phospho-IκB-α(P-I κBα)蛋白錶達水平.結果 500、1 000、5 000 μmol/L染氟組THP-1細胞存活率[(73.21±3.67)％、(31.40±4.56)％、(0.40±0.24)％]明顯低于陰性對照組[(100.00±0.00)％,P均＜0.01].低、高劑量染氟組炎性因子IL-1β[(1.42±0.79)、(19.47±2.90)ng/L]、TNF-α含量[(61.06±2.20)、(172.72±2.29)ng/L]均高于對照組[(0.36±0.07)、(31.07±0.81)ng/L,P均＜0.05];高劑量染氟組IκBα蛋白錶達水平[(63.53±9.67)％]低于對照組[(100.00±10.99)％,P＜ 0.01];低、高劑量染氟組P-P65蛋白錶達水平[(113.71±8.99)％、(134.74±1.93)％]明顯高于對照組[(100.00±5.48)％,P均＜0.05];低、高氟劑量組P-IκBα蛋白錶達水平[(152.61±14.16)％、(176.91±7.95)％]高于對照組[(100.00±14.82)％,P均＜0.01].線性迴歸分析錶明,TNF-α和IL-1β含量與P-P65蛋白錶達水平之間呈線性正相關(r=0.74、0.72,P均＜0.05).結論 過量氟可誘導小膠質細胞產生炎性因子併激活NF-κB信號通路,炎性因子釋放及信號通路的激活可能是氟導緻中樞神經繫統損傷的機製之一.
목적 관찰불대소효질세포염증인자표체급세포핵인자κB(NF-κB)신호통로적영향.방법 이체외배양인급성단핵세포백혈병세포(THP-1)작위소효질세포모형,용불동농도[0(음성대조조)、10、50、100、500、1 000、5 000 μmol/L]불화납(NaF)배양48 h,CCK8법검측세포존활솔.근거CCK8법검측결과,장THP-1세포분위3조:대조조화저、고제량염불조,분별용0、500、5 000μmol/L NaF염불처리48 h,이매련면역흡부실험(ELISA)검측세포배양액상청중염성세포인자백세포개소(IL)-1β화종류배사인자(TNF)-α,단백질면역인적(Western blot)법검측배양세포IκBo、Phospho-NF-κB p65 (P-P65)화Phospho-IκB-α(P-I κBα)단백표체수평.결과 500、1 000、5 000 μmol/L염불조THP-1세포존활솔[(73.21±3.67)％、(31.40±4.56)％、(0.40±0.24)％]명현저우음성대조조[(100.00±0.00)％,P균＜0.01].저、고제량염불조염성인자IL-1β[(1.42±0.79)、(19.47±2.90)ng/L]、TNF-α함량[(61.06±2.20)、(172.72±2.29)ng/L]균고우대조조[(0.36±0.07)、(31.07±0.81)ng/L,P균＜0.05];고제량염불조IκBα단백표체수평[(63.53±9.67)％]저우대조조[(100.00±10.99)％,P＜ 0.01];저、고제량염불조P-P65단백표체수평[(113.71±8.99)％、(134.74±1.93)％]명현고우대조조[(100.00±5.48)％,P균＜0.05];저、고불제량조P-IκBα단백표체수평[(152.61±14.16)％、(176.91±7.95)％]고우대조조[(100.00±14.82)％,P균＜0.01].선성회귀분석표명,TNF-α화IL-1β함량여P-P65단백표체수평지간정선성정상관(r=0.74、0.72,P균＜0.05).결론 과량불가유도소효질세포산생염성인자병격활NF-κB신호통로,염성인자석방급신호통로적격활가능시불도치중추신경계통손상적궤제지일.
Objective To investigate fluoride-induced inflammation and nuclear factor-κB (NF-κB) signaling pathway in cultured human acute monocytic leukemia cells (THP-1).Methods In vitro cultured THP-1 cells were used as a model of microglia.After cultured with different concentrations of [0 (negative control group),10,50,100,500,1 000 and 5 000 μmol/L] sodium fluoride (NaF) for 48 h,the survival of cells was detected by CCK8.THP-1 cells were divided into 3 groups:control group,low dose and high dose fluoride groups according to the results of CCK8 assay,and then treated with different concentrations of sodium fluoride (0,500,5 000 μmol/L) for 48 h,concentrations of inflammatory cytokines,such as Interleukin-lβ (IL-1β) and tumor necrosis factor-α (TNF-α) were measured by enzyme linked immunosorbent assay (ELISA) in THP-1 mononuclear cell culture medium.The protein levels of IκBα,phospho-NF-κB p65 and phospho-IκB-α were detected by Western blotting.Results THP-1 cells were treated with different concentrations of sodium fluoride (500,1 000,5 000 μ mol/L) for 48 h.Fluoride group THP-1 cell survival rate [(73.21 ± 3.67)％,(31.40 ± 4.56)％,(0.40 ± 0.24)％] was lower than that of the negative control group [(100.00 ± 0.00)％,all P ＜ 0.01].Compared to the control groups [(0.36 ± 0.07),(31.07 ± 0.81)ng/L],significant increases of the inflammatory cytokines IL-1β [(1.42 ± 0.79),(19.47 ± 2.90)ng/L] and TNF-α [(61.06 ± 2.20),(172.72 ± 2.29)ng/L] were detected in culture medium in low-fluoride and high fluoride groups,respectively.Interestingly,compared to the control groups [(100.00 ± 5.48)％,(100.00 ± 14.82)％],significant increases of phospho-NF-κB p65 [(113.71 ± 8.99)％,(134.74 ± 1.93)％] and phospho-IκB-α [(152.61 ± 14.16)％,(176.91 ± 7.95)％] were observed in both low-fluoride and high fluoride groups.Meanwhile,the protein level of IκBα in high fluoride group [(63.53 ± 9.67)％] was significantly lower than that of the control group [(100.00 ± 10.99)％,P ＜ 0.01].Furthermore,significant positive correlation was detected between increased IL-1β,TNF-α and phospho-NF-κB p65 (r =0.74,0.72,all P ＜ 0.05).Conclusions Excessive fluoride can induce microglial cells to release inflammatory cytokines and activate nuclear factor-κB signaling pathway.The release of inflammatory cytokines and activation of the signaling pathway may be one of the mechanisms of the damage of the central nervous system caused by sodium fluoride.
