| Objective: Malignant lymphoma is a kind of malignant tumor in thesource of lymphaden and (or) lymph tissue. Malignant lymphoma occurredmostly with lymphocyte proliferation differentiation during the process of theimmune response which is related to the immune cell malignanttransformation, and is a malignant tumor of the immune system. Due to theinfluence of the genetic factors, physical and chemical factors in theenvironment, etc.There is an upward trend incidence of malignant lymphomain children, Therefore children’s health and life is serious threated. Pediatricmalignant lymphoma in clinical manifestation, histopathological types andrespective treatments are different from adults malignant lymphoma,whichhave its unique biological characteristics, Such as: Short incubation period、rapid growth、strong invasive and so on. Currently,combination chemotherapyis still the principal treatment for malignant lymphoma, but the side effects ofchemotherapy and multi-resistantance is one of the important factors affectingthe long-term survival of children. A variety of targeted drugs are discovered:Rituximab（Anti-CD20）、Anti-CD22McAb for example, which clinicalapplication increased the rate of remission of type B cell lymphoma. Howeverthe targeted drugs of T cell lymphoma hasn’t been found, the clinicalprognosis is still poor. Cytokine induced killer cells is a new kind of immuneactive cells, Which is the human peripheral blood mononuclear cells in vitrounder a wide variety of cytokines stimulated, get a group of heterogeneouscells, Which also has a strong anti tumor activity of T lymphocytes and naturalkiller cells’s major histocompatibility and the limited killing tumor twocharacteristics.Survivin is a member of family of apoptosis protein inhibitors,Which can inhibit cell apoptosis, adjust the function of the cell cycle and celldivision. Livin which is discovered a new members of apoptosis inhibiting protein family, the antiapoptotic proteins whcih independed of the Bcl-2. Livinmainly exists in cytoplasm and cytoplasmic Livin in a filamentous expression,at the same time is also expressed in the nucleus. Livin anti-poptotic effectthrough the following way to completed:(1) antagonism death receptorpathway;(2) against mitochondria mediated apoptosis effect and resistance tochemotherapy drugs mediated apoptosis. Livin can directly combine withCaspases, thus inhibiting the activity of antagonism of apoptosis, Smac mayalso through decomposition, preventing the antagonism between XIAP andindirect inhibition of Caspases to regulate apoptosis.Research shows, Livin innormal adult tissue such as the spleen, thymus gland, prostate, testis, smallintestine, colon, ovarian, liver, lungs, brain, skeletal muscle, and do notexpress or low expression in lymphocytes and peripheral blood.Most tumorswith high expression, such as melanoma, breast cancer, cervical cancer,bladder cancer, prostate cancer, stomach cancer, colon cancer and leukemiaetc.This experiment through the study of umbilical cord blood CIK cells forLivin, the influence of Survivin and study its possible molecular mechanismof anti-tumor effect of umbilical cord blood CIK cells to provide theoreticalbasis for the clinical application of treatment of malignant lymphoma.Methods: In vitro cultivation of Jurkat T cell lymphoma cells andumbilical cord blood CIK cells.Divided into cytarabine, umbilical cord bloodCIK cells group and cytarabine, umbilical cord blood CIK cell group and thecontrol group. Application of RT-PCR method to detect tumor cell specificityapoptosis protein Livin Survivin mRNA expression level; Determined byMTT colorimetry to detect damage rate; Flow cytometry (FCM) to detectapoptosis rate after culturing the supernatant fluid of umbilical cord blood CIKcells Jurkat cells.All measurement data showed by means±standard deviation (X±S), andused SPSS13.0statistical software for statistical analysis. The means datebetween groups tested by one-way ANOVA measure, and the means datewithin groups tested by SNK measure. There was a statistical difference if P value less than0.05.Results:1RT-PCR detection results show: in the role of CIK cells, cytarabineJurkat cells after24h,48h, tumor cells apoptosis inhibit factors of specificmRNA expression of Livin, Survivin quantity are lower than the control group,One of CIK cells joint cytarabine group is significantly lower than the controlgroup.And after each experimental group the Livin-αA/β-actin A、Livin-βA/β-actin A、Survivin A/β-actin A48h is less than24h.(A is the greyvalue for electrophoresis bands), they have statistically difference (P<0.05).CIK cell promote the apoptosis of Jurkat cells by down-regulated theexpression of Livin, Survivin apoptosis inhibit factor, and it has timedependence.2Determined by MTT colorimetric test result display: Preliminaryexperiments CIK cells and effect target than Jurkat cells5:1、10:1、15:1、20:1after12h、24h、48h, compared with the control group, each group absorbancevalue (OD) all have varying degrees of decline. Each experimental groupcompared with the control group were statistically significant (P <0.05).Namely after CIK cells in Jurkat cells as the target higher than and longeraction time, CIK cell killing effect of Jurkat gradually strengthen, assumes theconcentration-time dependence. The experimental group show cytarabine、CIKcells、Cytarabine combined with CIK cells, each group absorbance value(OD) all have varying degrees of decline, including cytarabine absorbance hadthe greatest reduction combined with CIK cells group, the experimental groupand control group were statistically significant (P <0.05). Kill rate calculationresults show that cytarabine combined with umbilical cord blood-CIK groupup to(61.23±4.6)％, is1.41times that of cytarabine alone, is1.34times ofCIK alone group (P <0.05).3Flow cytometry detection according to the results: See Jurkat cellmicroscopic observation, together with CIK cells supernatant after12htraining, there are a few lymphoma cell shrinkage, apoptosis, and apoptosispeak after24h,48h after several lymphoma cells apoptosis significantly increased, the control group did not appeared apoptosis peak. Compared withthe control group, experimental group tumor cell apoptosis rate wassignificantly higher (P <0.05), The apoptosis rate after24h was (31.2±2.9)％,after48h was (38.6±3.6)％.Comparison between the experimental group,along with the role of the extension of time, the apoptosis rate is on the rise,the significant difference between groups (P <0.05). CIK cells can be inducedby various cytokines released into the extracellular cell apoptosis to kill tumorcells, and time dependence.Conclusion:1Umbilical cord blood CIK can induce lymphoma Jurkat cells apoptosis,and can make the Jurkat cell Livin, Survivin mRNA expression decreased,the obvious dose and time dependent relationship, think that umbilical cordblood CIK by down-regulated Livin, Survivin the expression to inducelymphoma cells apoptosis.2Umbilical cord blood CIK cells in vitro for T cell lymphoma (Jurkatcell lines) growth has obvious inhibitory effect, and the effect of target thanwithin the scope of5:1–20:1me and dose dependent.3CIK cells can be induced by various cytokines released into theextracellular cell apoptosis to kill tumor cells, and time dependence.4Umbilical cord blood CIK combination chemotherapy drugs(cytarabine) acted with lymphoma cells the anti-tumor effect obviouslystronger than the one of the antineoplastic effect, for the clinical drugtreatment with combination chemotherapy for lymphoma provides experimentbasis. |
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