中国免疫学杂志
中國免疫學雜誌
중국면역학잡지
CHINESE JOURNAL OF IMMUNOLOGY
2015年
8期
1040-1044,1048
,共6页
杨惠%李银聚%刘一尘%程相朝%杨丹芳%韩海锋%金修哲
楊惠%李銀聚%劉一塵%程相朝%楊丹芳%韓海鋒%金脩哲
양혜%리은취%류일진%정상조%양단방%한해봉%금수철
IFN-α-IL-18%酵母表达%淋巴细胞转化试验%核因子κB
IFN-α-IL-18%酵母錶達%淋巴細胞轉化試驗%覈因子κB
IFN-α-IL-18%효모표체%림파세포전화시험%핵인자κB
Interferon-α-interleukin-18%Yeast expression%The lymphocyte transformation test%NF-κB
目的::探讨鸡IFN-α-IL-18融合蛋白对鸡外周血淋巴细胞转化及核因子κB( NF-κB)的影响。方法:构建重组酵母表达质粒pPICZ-IFN-α、pPICZ-IL-18和pPICZ-IFN-α-IL-18,电转化酵母菌X-33后以1%甲醇诱导,SDS-PAGE和Western blot鉴定表达IFN-α、IL-18及IFN-α-IL-18蛋白,淋巴细胞转化试验初步检测表达产物生物学活性,双抗体夹心法测定不同时段淋巴细胞中总NF-κB和细胞核中NF-κB的含量。结果:酵母培养上清液中存在分子量分别为22、23和43 kD的目的蛋白;表达的IFN-α-IL-18和IL-18均能明显促进鸡外周血淋巴细胞转化(P<0.05),IFN-α对淋巴细胞转化有促进作用,但总体差异不显著(P>0.05)。与对照组相比,经IFN-α-IL-18和IL-18作用的鸡淋巴细胞中总NF-κB含量明显增加(P<0.05),细胞核中NF-κB含量差异极显著(P<0.01),IFN-α作用的淋巴细胞中总NF-κB含量与对照组相比差异不显著(P>0.05),细胞核中NF-κB含量显著增加(P<0.05)。结论:IFN-α-IL-18及IL-18能够明显促进淋巴细胞转化,IFN-α促进淋巴细胞转化活性较弱,这种促淋转机制与NF-κB的表达、活化及核质转运有关,为进一步研究IFN-α-IL-18融合蛋白的功能及其在疫病防控中的作用奠定了基础。
目的::探討鷄IFN-α-IL-18融閤蛋白對鷄外週血淋巴細胞轉化及覈因子κB( NF-κB)的影響。方法:構建重組酵母錶達質粒pPICZ-IFN-α、pPICZ-IL-18和pPICZ-IFN-α-IL-18,電轉化酵母菌X-33後以1%甲醇誘導,SDS-PAGE和Western blot鑒定錶達IFN-α、IL-18及IFN-α-IL-18蛋白,淋巴細胞轉化試驗初步檢測錶達產物生物學活性,雙抗體夾心法測定不同時段淋巴細胞中總NF-κB和細胞覈中NF-κB的含量。結果:酵母培養上清液中存在分子量分彆為22、23和43 kD的目的蛋白;錶達的IFN-α-IL-18和IL-18均能明顯促進鷄外週血淋巴細胞轉化(P<0.05),IFN-α對淋巴細胞轉化有促進作用,但總體差異不顯著(P>0.05)。與對照組相比,經IFN-α-IL-18和IL-18作用的鷄淋巴細胞中總NF-κB含量明顯增加(P<0.05),細胞覈中NF-κB含量差異極顯著(P<0.01),IFN-α作用的淋巴細胞中總NF-κB含量與對照組相比差異不顯著(P>0.05),細胞覈中NF-κB含量顯著增加(P<0.05)。結論:IFN-α-IL-18及IL-18能夠明顯促進淋巴細胞轉化,IFN-α促進淋巴細胞轉化活性較弱,這種促淋轉機製與NF-κB的錶達、活化及覈質轉運有關,為進一步研究IFN-α-IL-18融閤蛋白的功能及其在疫病防控中的作用奠定瞭基礎。
목적::탐토계IFN-α-IL-18융합단백대계외주혈림파세포전화급핵인자κB( NF-κB)적영향。방법:구건중조효모표체질립pPICZ-IFN-α、pPICZ-IL-18화pPICZ-IFN-α-IL-18,전전화효모균X-33후이1%갑순유도,SDS-PAGE화Western blot감정표체IFN-α、IL-18급IFN-α-IL-18단백,림파세포전화시험초보검측표체산물생물학활성,쌍항체협심법측정불동시단림파세포중총NF-κB화세포핵중NF-κB적함량。결과:효모배양상청액중존재분자량분별위22、23화43 kD적목적단백;표체적IFN-α-IL-18화IL-18균능명현촉진계외주혈림파세포전화(P<0.05),IFN-α대림파세포전화유촉진작용,단총체차이불현저(P>0.05)。여대조조상비,경IFN-α-IL-18화IL-18작용적계림파세포중총NF-κB함량명현증가(P<0.05),세포핵중NF-κB함량차이겁현저(P<0.01),IFN-α작용적림파세포중총NF-κB함량여대조조상비차이불현저(P>0.05),세포핵중NF-κB함량현저증가(P<0.05)。결론:IFN-α-IL-18급IL-18능구명현촉진림파세포전화,IFN-α촉진림파세포전화활성교약,저충촉림전궤제여NF-κB적표체、활화급핵질전운유관,위진일보연구IFN-α-IL-18융합단백적공능급기재역병방공중적작용전정료기출。
Objective:To discuss the effects of recombinant IFN-α-IL-18 fusion protein on chicken lymphocyte transformation and the chicken nuclear factor kappa B ( NF-κB ) activity.Methods: The recombinant plasmids pPICZ-IFN-α, pPICZ-IL-18 and pPICZ-IFN-α-IL-18 were constructed,and transformed into P.Pastoris X-33 strain by electroporation.The recombinant proteins were ex-pressed under the induction of 1% methanol,and detected by SDS-PAGE and Western blot.The biological activities of the expressed products were detected by the lymphocyte transformation test,the NF-κB concentration in chicken lymphocyte and lymphocytic nucleus were detected at different times with ELISA.Results:SDS-PAGE and Western blot showed that the expressed products existed in super-natant,and the molecular weights were about 22 kD,23 kD and 43 kD,respectively.The expressed products IL-18 and IFN-α-IL-18 could stimulate chicken lymphocyte transformation (P<0.05),the biological activity of IFN-αstimulating lymphocyte transform was feeble (P>0.05).Compared with the control group,the total NF-κB concentration in chicken lymphocyte induced by IL-18 and IFN-α-IL-18 were increased (P<0.05),and the NF-κB in lymphocytic nucleus was increased significantly (P<0.01).Otherwise,the total NF-κB in lymphocyte induced by IFN-αincrease was limited (P>0.05),but the NF-κB in lymphocytic nucleus showed the remarkable increase ( P<0.05 ) .Conclusion: IFN-α-IL-18 and IL-18 can promote lymphocyte transformation significantly, the activity of IFN-αinduced lymphocyte transformation is imperfect.The biological activity of stimulating lymphocyte transformation is associated with the NF-κB expression,activation and nucleo-cytoplasmic transport.The study built foundation for the function of IFN-α-IL-18 fusion protein and the exploration of the role of controlling epidemic diseases.