CAJ | 학술논문

目的筛选能与集落细胞刺激因子(granulocyte-colony stimulatingfactor,G-CSF)结合,并能调节血细胞生长的活性物质,为研究药理作用提供实验依据.方法采用毛细管电泳(capillary ehetrophoresis,CE)方法研究15种不同链长和不同硫酸基含量的硫酸寡聚糖(Dextran sulfate,DexS)和G-CSF的相互作用.采用细胞生物学实验观察被筛选出的寡聚糖与G-CSF结合后对小鼠白血病细胞(bipotential murine hemopoietic cellline,NFS-60)增生、细胞周期、细胞表面抗原的表达和细胞形态的影响,以证明毛细管电泳所筛选出的DexS具有促进细胞分化等生物学活性.结果除肝素寡聚糖、降解的卡拉胶、低分子量右旋Dexs、非硫酸化的寡聚糖以及碱性壳寡糖外,其余10种寡聚糖与G-CSF均有相互作用.DexS500、λ卡拉胶、ι卡拉胶、κ卡拉胶和具有抗凝活性的低分子量肝素(LMWH)-1能抑制NFS-60细胞生长,其作用NFS-60细胞后相对吸光度值分别为31.33±2.08、37.67±2.08、45.00±1.20、43.00±2.65、74.67±1.52,与对照组100.00±3.00比较,差异有统计学意义(t值分别为27.94、26.71、26.42、26.61、10.54,P均<0.01).形态学结果表明,LMWH、λ卡拉胶和相对分子质量为500 000的硫酸葡聚糖能有效地诱导早幼粒细胞过渡到中性晚幼粒细胞.流式细胞术分析结果表明,用LMWH和λ卡拉胶分别处理细胞后,其表面的CD11b和CD18表达量增加.在硫酸葡聚糖(DexSS00)和λ或ι卡拉胶作用后,细胞表面CD34含量增加.用不同的寡聚糖作用后,其CD34表达量降低.用壳寡糖处理后,CDs含量几乎保持不变.细胞周期分析结果表明,在LMWH、DexS500和λ或ι卡拉胶作用后,S期NFS-60细胞减少,而壳寡糖处理后其细胞增加.结论毛细管电泳技术能有效地筛选出与G-CSF有相互作用的酸性寡聚糖,相互作用的强弱与寡聚糖的碳链长度和酸性基团的含量及其基团位置有关.
목적 사선능여집락세포자격인자(granulocyte-colony stimulatingfactor,G-CSF)결합,병능조절혈세포생장적활성물질,위연구약리작용제공실험의거.방법 채용모세관전영(capillary ehetrophoresis,CE)방법연구15충불동련장화불동류산기함량적류산과취당(Dextran sulfate,DexS)화G-CSF적상호작용.채용세포생물학실험관찰피사선출적과취당여G-CSF결합후대소서백혈병세포(bipotential murine hemopoietic cellline,NFS-60)증생、세포주기、세포표면항원적표체화세포형태적영향,이증명모세관전영소사선출적DexS구유촉진세포분화등생물학활성.결과 제간소과취당、강해적잡랍효、저분자량우선Dexs、비류산화적과취당이급감성각과당외,기여10충과취당여G-CSF균유상호작용.DexS500、λ잡랍효、ι잡랍효、κ잡랍효화구유항응활성적저분자량간소(LMWH)-1능억제NFS-60세포생장,기작용NFS-60세포후상대흡광도치분별위31.33±2.08、37.67±2.08、45.00±1.20、43.00±2.65、74.67±1.52,여대조조100.00±3.00비교,차이유통계학의의(t치분별위27.94、26.71、26.42、26.61、10.54,P균<0.01).형태학결과표명,LMWH、λ잡랍효화상대분자질량위500 000적류산포취당능유효지유도조유립세포과도도중성만유립세포.류식세포술분석결과표명,용LMWH화λ잡랍효분별처리세포후,기표면적CD11b화CD18표체량증가.재류산포취당(DexSS00)화λ혹ι잡랍효작용후,세포표면CD34함량증가.용불동적과취당작용후,기CD34표체량강저.용각과당처리후,CDs함량궤호보지불변.세포주기분석결과표명,재LMWH、DexS500화λ혹ι잡랍효작용후,S기NFS-60세포감소,이각과당처리후기세포증가.결론 모세관전영기술능유효지사선출여G-CSF유상호작용적산성과취당,상호작용적강약여과취당적탄련장도화산성기단적함량급기기단위치유관.
Objective To screen out the bioactive sulfate saccharides interacted with granulocyte colony-stimulating factor (G-CSF) and provide some scientific data on their pharmacology. Methods The interactions between G-CSF and 15 sulfate saccharides with various chain lengths and extent of sulfation were studied by capillary electrophoresis. Furthermore, the effects of these G-CSF binding saccharides on the proliferation and differentiation of leukemia cell line and NFS-60 were also investigated. Results Except for the hepatosaccharide, degradable carrageenan, Dextran sulfate (DexS) with a low molecular mass the nonsulfated sancharide and alkaline chitosan oligosaccharide, the rest of saccharides had interact with G-CSF.DexS500, λ carrageenan, ι carrageenan, κ carrageenan and low molecular weight heparin-1(LMWH-1)with anticoagulant activity could inhibit the NFS-60 growth. Compared with the absorbency of the control group (100.00±3.00) after these saccharides interacted with the NPS-60 cells, their relative absorbencies were 31.33±2.08,37.67±2.08,45.00±1.20,43.00±2.65 and 74.67±1.52, respectively. Their differences had a statistical significance(t=27.94,26.71,26.42,26.61,10.54,P<0.01). The morphological analytic results showed that the LMWH and lambda carrageenan as well as DexS with a molecular mass of 500 000 induce effectively promyclocyte to transit metamyelocyte. Flow cytometry results showed enhanced expressions of CD11b, CD18 and CD45 after treatment with the LMWH, lambda carrageenan and DexS500,respectively. Cell cycle analysis indicated that the S phase population decreased after treatment with these saccharides. Conclusions Capillary electrophoresis is an effective medicine-screening technique that is used to screen some of these saccharides which could bind to G-CSF. And it is found that the interactions are correlated with the chain length, the extent and the position of sulfation of these saccharides.