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Investigation on the Proliferation,Differentiation and Apoptosis Effect and Mechanism of LfcinB in Jurkat Cell

Author: HanZhaoLian
Tutor: XuXiaoZuo
School: Northeast Agricultural University
Course: Of Food Science
Keywords: LfcinB Jurkat cells Apoptosis P21waf1 Notch1 AKT
CLC: R151
Type: Master's thesis
Year: 2013
Downloads: 40
Quote: 0
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Abstract


Lactoferricin B (LfcinB) is a microbial activity peptide of high antimicrobial activity, and hasextensive source. It has no effects to normal cells at concentration range of antineoplastic activity,but has pertinence that show cytotoxicity to most cancer cells of human. Synthetic LfcinB wasacted on acute T cell leukemia cells (Jurkat) in this experiment, and flow cytometry (FCM) andwestern blotting were used to study and discuss the effects and mechanisms of LfcinB to Jurkatcells on proliferation, differentiation and apoptosis.All this experiment has four parts:(1) Cell counting kit-8was used to measure the effect ofLfcinB to Jurkat cells on proliferation.100nmol/mL sirolimus as the blank group and LfcinB of0μg/mL,50μg/mL,100μg/mL,200μg/mL and400μg/mL as the experimental groups to measurethe proliferation ratios of Jurket cells. As the results, the effects of LfcinB on proliferation of Jurkatcells were enhanced by the increase of LfcinB concentration and action time and appeareddependence to concentration and action time.(2) Morphological variation of Jurkat cell nucleusthat was effect by LfcinB of0μg/mL,50μg/mL,100μg/mL,200μg/mL and400μg/mL after24hwas analysed through inverted flurescence microscopy. From the results, cell nucleus of theexperimental groups were concentrated and appeared dense dyed state, however, the blank groupswere appeared uniform dyed state. The cells of experimental groups were apoptosis.(3) Thechange rule of early apoptosis that LfcinB effect on Jurket cells was measured byAnnexinⅤ-FITC/PI double marker flow cytometry.100nmol/mL sirolimus as the blank group andLfcinB of0μg/mL,50μg/mL,100μg/mL,200μg/mL and400μg/mL as the experimental groupsto measure the apoptosis ratios of Jurket cells. According to the results, early apoptosis ratios ofJurkat cells were increased by LfcinB concentration at the same action time and at the same LfcinBconcentration when action time at72h, the early apoptosis ratio decreased obviously andproportion of late apoptosis or death cells was increased.(4) The changes of phosphorylated levelof P21waf1, AKT and mTOR of Jurket cells and the expressions of protein of Nocth1, C-MYC,Bcl-2, Bax and Cyclin D1. This experiment has eight parts which were100nmol/mL sirolimus,LfcinB of0μg/mL,50μg/mL,100μg/mL,200μg/mL and400μg/mL, sirolimus+PI3K inhibitor and LfcinB+PI3K inhibitor. The phosphorylation of P21waf1, AKT and mTOR of Jurket cells weredecreased by increasing of LfcinB concentration and prolonging of action time. And theexpressions of Nocth1, C-MYC, Bcl-2and Cyclin D1were negative correlation with the LfcinBconcentration and action time, but the expressions of Bax were appeared positive correlation withLfcinB concentration and action time.

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