胃内肠嗜铬样细胞的分离与培养及功能测定

IIsolation and culture of enterochromaffin like cell and its functionaldetermination

目的 体内研究已提示胃内肠嗜铬样 (ECL)细胞在调节胃酸分泌中起到重要的作用。但以往由于缺乏分离、培养ECL细胞的方法 ,对它的生理调节、病理变化的研究受到了限制。方法 采用链霉蛋白酶的消化 ,淘析法和颗粒梯度离心分离纯化ECL细胞。结果 经电镜 ,组胺含量测定 ,纯化率达 90 %～ 95 %。经锥虫蓝染色细胞成活率大于 95 %。分离纯化后的ECL细胞短期培养 2 4h后 ,胃泌素对ECL细胞的作用 ,经酶标免疫测定组胺分泌的EC50 为 3× 10 -10 mol/L。经BrdU掺入试验测定DNA增生的EC50 为 2× 10 -11mol/L。胃泌素受体拮抗剂L3652 66抑制胃泌素释放组胺的作用 (IC50 10 -8mol/L)。结论 ECL细胞在培养系统中生长良好 。

Objective Invivostudieshavesuggestedanimportantroleforgastricenterochromaffin like (ECL)cellinmediatingacidsecretion .ThefurtherstudyofECLcellinitsphysiologicalfunctionand pathologicalchangehasbeenlimitedsincenopurepreparationandculturesystemwereavailable .Methods ThepurposeofthisstudyistodevelopapureisolatedECLcellpreparationanditsculturesystem .ECLcell wasisolatedbypronaseandpurifiedbyacombinationofelutriationanddensity gradientcentrifugation .En richmentwasdeterminedbytotalhistaminecontentandelectronmicroscope .Intheculturesystemfor 2 4 hoursthefunctionofECLcellwasdeterminedbydosedependentgastrinstimulatedhistaminesecretionand DNAsynthesis.Results ThepurityofECLpreparationwas 90 %～ 95 % .Trypanblueexclusiondemon strated >95 %viability .Histaminesecretionismeasuredbyenzymeimmunoassay .Basalsecretionwas (0 .6 ± 0 .2 )nmol/ 10 3cell.GastrinstimulatedhistaminesecretionwithEC50 of 3× 10 -10 mol/Landwasinhibited bypretreatmentofgastrinreceptorantagonistL3652 66,(IC50 10 -8mol/L) .TheDNAsynthesisofECLcell wasdeterminedbyBrdU .GastrinstimulatedECLcellDNAsynthesiswithEC50 of 2× 10 -11mol/L . ConclusionAboveresultsindicatedthatECLcellcouldlivewellinculturesystemandprovideagoodmeansfor investigatingcentralroleofECLcellfunction .