医学分子生物学杂志
醫學分子生物學雜誌
의학분자생물학잡지
FOREIGN MEDICAL SCIENCES
2014年
3期
169-174
,共6页
冯翠娟%陆炜洪%欧金来%李沙
馮翠娟%陸煒洪%歐金來%李沙
풍취연%륙위홍%구금래%리사
壳聚糖%纳米细胞%pEGFP-N1质粒%HeLa细胞%转染率%细胞毒性
殼聚糖%納米細胞%pEGFP-N1質粒%HeLa細胞%轉染率%細胞毒性
각취당%납미세포%pEGFP-N1질립%HeLa세포%전염솔%세포독성
chitosan%nanocell%pEGFP-N1%HeLa cells%transfection efficiency%cytotocixity
目的：构建核壳型壳聚糖（chitosan， CTS）纳米细胞（nanocell， NC），探讨其作为基因转染载体的可行性。方法以壳聚糖为载体材料， pEGFP-N1为模型质粒，制备质粒复合物pEGFP-N1/CTS-NP。以pEGFP-N1/CTS-NP为核心，用复乳法外包含十八胺（stearylamine， SA）的脂质体膜制备纳米细胞pEG-FP-N1/CTS-SANC，用CTS对其进行包覆制成CTS-（ pEGFP-N1/CTS-SANC），测定其形态、粒径与电位。用MTT法测定NC的细胞毒性，用荧光显微镜及流式细胞仪定性、定量地评价其在HeLa细胞中的转染率。结果所制备的样品多呈类球形，粒径与电位分别分布在120～220 nm与40～65 mV之间。 pEGFP-N1/CTS-SANC与CTS-（ pEGFP-N1/CTS-SANC）均可将质粒转染到HeLa细胞并表达绿色荧光蛋白。 CTS-（ pEGFP-N1/CTS-SANC）可降低pEGFP-N1/CTS-SANC的毒性，在相同质粒用量下，使HeLa细胞存活率从81．8％增至100．9％，转染率从3．90％增至8．13％，达市售脂质体转染试剂的76％。结论壳聚糖纳米细胞有望作为基因转染的载体。
目的：構建覈殼型殼聚糖（chitosan， CTS）納米細胞（nanocell， NC），探討其作為基因轉染載體的可行性。方法以殼聚糖為載體材料， pEGFP-N1為模型質粒，製備質粒複閤物pEGFP-N1/CTS-NP。以pEGFP-N1/CTS-NP為覈心，用複乳法外包含十八胺（stearylamine， SA）的脂質體膜製備納米細胞pEG-FP-N1/CTS-SANC，用CTS對其進行包覆製成CTS-（ pEGFP-N1/CTS-SANC），測定其形態、粒徑與電位。用MTT法測定NC的細胞毒性，用熒光顯微鏡及流式細胞儀定性、定量地評價其在HeLa細胞中的轉染率。結果所製備的樣品多呈類毬形，粒徑與電位分彆分佈在120～220 nm與40～65 mV之間。 pEGFP-N1/CTS-SANC與CTS-（ pEGFP-N1/CTS-SANC）均可將質粒轉染到HeLa細胞併錶達綠色熒光蛋白。 CTS-（ pEGFP-N1/CTS-SANC）可降低pEGFP-N1/CTS-SANC的毒性，在相同質粒用量下，使HeLa細胞存活率從81．8％增至100．9％，轉染率從3．90％增至8．13％，達市售脂質體轉染試劑的76％。結論殼聚糖納米細胞有望作為基因轉染的載體。
목적：구건핵각형각취당（chitosan， CTS）납미세포（nanocell， NC），탐토기작위기인전염재체적가행성。방법이각취당위재체재료， pEGFP-N1위모형질립，제비질립복합물pEGFP-N1/CTS-NP。이pEGFP-N1/CTS-NP위핵심，용복유법외포함십팔알（stearylamine， SA）적지질체막제비납미세포pEG-FP-N1/CTS-SANC，용CTS대기진행포복제성CTS-（ pEGFP-N1/CTS-SANC），측정기형태、립경여전위。용MTT법측정NC적세포독성，용형광현미경급류식세포의정성、정량지평개기재HeLa세포중적전염솔。결과소제비적양품다정류구형，립경여전위분별분포재120～220 nm여40～65 mV지간。 pEGFP-N1/CTS-SANC여CTS-（ pEGFP-N1/CTS-SANC）균가장질립전염도HeLa세포병표체록색형광단백。 CTS-（ pEGFP-N1/CTS-SANC）가강저pEGFP-N1/CTS-SANC적독성，재상동질립용량하，사HeLa세포존활솔종81．8％증지100．9％，전염솔종3．90％증지8．13％，체시수지질체전염시제적76％。결론각취당납미세포유망작위기인전염적재체。
Objective To develop a core-shell type chitosan nanocell ( CTS-NC) and to in-vestigate its feasibility as a vector of gene transfection .Methods The plasmid complex , pEGFP-N1/CTS-NP was prepared by using chitosan as the carrier material and pEGFP-N1 as the model plasmid.The pEGFP-N1/CTS-NP core was coated with a liposome membrane containing stearyl-amine ( SA) to obtain nanocell pEGFP-N1/CTS-SANC by double emulsifying method .The pEGFP-N1/CTS-SANC was additionally coated with chitosan outside to prepare CTS-( pEGFP-N1/CTS-SANC ) .The morphology , size and zeta potential of all these nano-size particles were meas-ured.The cytotoxicity was measured in HeLa cells by the MTT method .The gene transfection effi-ciency was evaluated in Hela cells by fluorescence inverted microscopy and flow cytometry qualita-tively and quantitatively .Results Most of the prepared nano-size particles were spherical .The size and zeta potential was within 120-220 nm and 40-65 mV, respectively.Both pEGFP-N1/CTS-SANC and CTS-( pEGFP-N1/CTS-SANC ) could be transfected into Hela cells , which afterwards ex-pressed green fluorescence protein .CTS-( pEGFP-N1/CTS-SANC ) reduced the cytotoxicity of pEGFP-N1/CTS-SANC by additional chitosan coating .With the same amount of plasmid used , the viability of Hela cells was increased from 81.8% to 100.9% and the transfection efficiency en-hanced from 3.90%to 8.13%, which was about 76%of that of commercial DNAFect transfection reagent.Conclusion The chitosan nanocell is expected to become a candidate vector for gene trans-fection.
