Study On Apoptosis Of Leukemia Cells Induced By Lycorine And The Caspase Pathway

Alkaloids exists widely in nature,it is nitrogen organic compounds containing a kind of alkaline,most alkaloids containing complex nitrogen ring structure,such as pyridine,indole,quinoline,purine,etc,and a few of them are amine compound.The discovery of alkaloid began in the early 19th century it is the most natural organic compounds and the earliest for people to study.Alkaloids have been found more than 6000 kinds until now,and it is increasing at a speed of about 100 kinds each year.Most alkaloids compounds have physiological activity,they are often effective components in many Chinese herbal medicine.The important basis for the extraction and purification of alkaloids is its solubility,the dissolution of their solubility and solvent type,the number of polar groups and the presence of nitrogen have a close relationship with the form of the atom in alkaloid molecules.According to the solubility of alkaloids in water and lipid solvents,it can be divided into alkaloids of fat soluble and water-soluble alkaloids.Lycorine widely exists in Amaryllidaceae plants,it is pyrrole and anthridine alkaloid.It has Inhibition of tumor,antihypertensive,antiviral,inhibition of acetylcholinesterase and other pharmacological activities.There are more and more research on the anti tumor effect of lycorine in recent years,But research on inhibition of leukemia cell and mechanism is still not found within the country.So the experiment mainly studied the cycle and apoptotic mechanism of lycorine block chronic myelogenous leukemia cell line K-562.In this experiment,MTT method was used to observe the HCTP inhibit K-562,the results showed,HCTP has certain inhibitory effect on the growth of K-562.In this experiment,MTT method was used to observe the lycorine inhibit human chronic myelogenous leukemia K-562,whether it has the inhibition effect on the growth of human chronic myelogenous leukemia cell line K-562.The results showed,lycorine hydrochloride has certain inhibitory effect on the growth of human chronic myelogenous leukemia K-562,24h,48h and 72h,the IC50 was 30.427μg/mL,22.506μg/ml and 16μg/mL.Further observations of lycorine treated chronic myelogenous leukemia cell line K-562 by inverted microscope and fluorescence microscope cell morphology,and use flow cytometry quantitative experimental determination of tumor cell apoptosis rate.In lycorine 5μg/mL,10μg/mL,20μg/mL chronic myelogenous leukemia cell line K-562 after 48h,the cell morphology was observed by inverted microscope,the negative control group cells grew well,closely arranged,shape intact.With the dosage increasing,the cells finally presents the morphological characteristics of apoptosis,irregular shape,crushing,cell growth gradually become sparse,adherent cells gradually become less.After Hoechst33258 staining of chronic myelogenous leukemia cell line K-562 were observed under fluorescence microscopy morphology,in lycorine 2.5μg/mL,5μg/mL,10μg/mL 48h,negative control group uniform staining,the cell growth in good condition and morphological rules.With the increase of the dose,Lycorine hydrochloride group come apoptotic bodies,uneven dyeing,decrease in the number of cells one and another and other phenomena.Lycorine hydrochloride human chronic myelogenous leukemia K-562 cells after 48h,the flow cytometry was used to detect the apoptosis rate display,with the dosage increased,the apoptosis rate also increased.To sum up,the lycorine can be induced in human chronic myelogenous leukemia K-562 cell apoptosis.Study of laser scanning confocal microscopy,Caspase-3 kit to observe the lycorine hydrochloride human chronic myelogenous leukemia K-562 mechanisms are employed,to study the impact of the mitochondrial membrane potential and Caspase-3.Lycorine 5μg/mL,10μg/mL,20μg/mL effect of human chronic myelogenous eukemia K-562 cells after 48h,using laser scanning confocal scanning microscope qualitative to detect mitochondrial membrane potential change.The results showed,lycorine in human chronic myelogenous leukemia after K-562,mitochondrial membrane potential decreased,and has relied on dosage.Caspase-3 kit for detection of lycorine to human chronic myelogenous leukemia cell K-562 protein changes,the results show,lycorine inhibit human chronic myelogenous leukemia cell K-562,Caspase-3 protein expression increased a lot.The study indicated that,lycorine can inhibit the growth of human chronic myelogenous leukemia K-562 cell,decreased mitochondrial membrane potential,mitochondrial damage,membrane permeability changes,thereby promoting the release of Caspase-3,induced chronic myelogenous leukemia K-562 cells occurred apoptosis.It illustrate mitochondrial pathway is likely to be a pathway in human chronic myelogenous leukemia K-562 cell apoptosis induced by lycorine.