K562细胞规模化培养及硒酸多糖诱导其凋的研究-食品工程专业毕业论文.docx

ABSTRACTIn ABSTRACT In thjs study，t11e baSic culture conditions of cl们11ic myelogenous leul(emia(CML) K562 cells were researched by bio-reactor．Simultalleously， t11e apoptosis of K562 cells inducedby selenic acid canila萨polysacch撕de were also studied．The cell mo叫ological chal．acteristics were obserVed by HE a11d Hoechst 33342／PI．The cell cycle剐陌est aIld mitochondrial membrane potential(△、王，m)were aSsessed by Flow Cytome臼呵(FCM)．The expression of印optosis—related protein and gene were allalyzed by inunuIlonuorescence a11d Real．time PCR． Firstly’tlle basic culture conditions of bio-reactor including iIlitial cell density，pH a11d stirring speed were researched．The results showed tllat me iIlitial cell dens时2．5×1 0’／mL， the pH 7．4 a11d the stirring speed 70 r／min were me best conditions of K562 cells scale culture．111e l鹕est dens时could reaCh 1．57×1 06 cell妇L under the condition． Secondly，tlle proliferation inllibition and apoptosis induction of selenic acid c甜ilage polysacch乏Lride to K562 ceUs were assessed．MTT sho、Ⅳed that t11e proliferation of K562 cells was inhibited by selellic acid cartilage polysaccharide with dose-dependent and time-dependent manner a11d the inllibition rate could reach 80％aRer K562 cells were exposed to l 00pg／mL selenic acid canilage polysaccharide f．or 48h．1’ypical apoptotic characteristics of ceU VoluIne reduction aIld c11r。omatin shrirll(age、Vere observed by inVerted microscope，HE and Hoechst 33342／PI． ARer廿e绷[1lent of selenic acid carrtilage polysaccharide，FCM analysis indicated that sele证c acid c砒ilage polySaccharide could increase me number of cells in S phase盘om 40．75％to 62．93％，a11d reduce t11e△、王，m by l 8％．It was reVealed t11at Cyclin A，CDK2 and Bcl-2 were decreaSed sigIlificantly but p2 1 and caspase一9 were i11creaSed si鲥ficaIltly by iInmunofluorescence a11d ELISA．ImPCR sho、ved tllat t11e genes of these proteins had t11e s锄e仃e：nds．AU the results proVed t11at selellic acid cartilage polysaccharide could inhibit K562 cells proliferation aI