The Effect On The Interaction Between Mid-Late Fetus And Mouse Hepatocarcinoma Cell H₂₂

Malignant tumor is the uncontrolled proliferation and dysdifferentiation of cells.Striking similarity can be found between tumorigenesis and embryonic development.The microenvironment of early embryo and the embryo implantation not only played a fine and orderly control for embryonic organizations but also to effectively control the behaiviors of tumor cells.This study focused on mid-late embryo,from the perspective of developmental biology approach the interaction between mid-late fetus and tumors.To provide new experimental evidence for the studies mid-late fetus'microenvironment reverse the malignant tumor phenotype.Objective (1)Observation of mice's birth,development,cancer bearing after translation with H22 cells ,we approach the interaction between mid-late fetus and tumor cells(2)Mouse fetus'abdominal cavity and its subcutaneouly after delivery were injected with H22 cells twice to approach on the effect of embryonic microenvironment.(3)To investigate the effects of mouse amniotic fluid on the proliferation and apoptosis of mouse hepatoma cell H22.we discussed whether the amniotic fluid Participate the effect of embryonic microenvironment on tumor cells.Methods (1)Every mouse fetus'abdominal cavity (D14~D19) was injected with 0.02ml of tumor suspension (1×106cells) using intrauterine transplantation technique in vivo.The fetus transplanted tumor cells observed its cancer ascites and development after operation before delivery and after birth.The physiological developmental indices were examined including the newborn appearance,opening-eyes time,descensus testiculorum and ostium vaginae openning time,the weight at 30 and 60 days of age, the function of concordance,movement,reflection,feeling,cognition and reproductive behavior were still examined during development. The HE staining revealed that the major organs,including the lung ,the liver and the kidney,wer histologically normal.(2)Every mouse fetus'abdominal cavity (D15) was injected with 0.02ml of tumor suspension (1×106cells) using IUT in vivo,and subcutaneouly with H22 cells after delivery.Nomarl newborn mice subcutaneouly were injected H22 cells as control group;Mouse fetus'abdominal cavity was injected with H22 cells as blank group.The fetus transplanted tumor cells observed its cancer ascites and development before delivery and after birth.The physiological developmental indices were examined including the newborn appearance,opening-eyes time,descensus testiculorum and ostium vaginae openning time,the weight at 1~4 weeks of age,the function of concordance,movement,reflection,feeling,cognition and reproductive behavior were still examined during development. The HE staining revealed that the major organs,including the lung ,the liver and the kidney.Messuring the long and short diameter of mass calculated gross tumor volume and relative growth rate.(3)Treated with the mixture of RPMI 1640 medium and 10% mouse amniotic fluid collected respectively on gestational day 11,13,15 and 17 or RPMI 1640 medium as control .the proliferation of H22 cell was detected with MTT and the apoptosis of H22 cells with Annexin V-FITC,H22 cells wer observed morphologically under electron microscope;The expressions of proliferation cell nulcear antigen(PCNA) and P53 in H22 cell were investigated with immunofluorescence staining and laser conlocal microscopy.Results (1)The later transplating time and pregnant mice'delivery was more.There were no significant difference in the delivery rate or fetal mortality before delivery between control group(P>0.05).The mice injected with H22 cells into their abdominal cavities on D14~D19developed successfully(57.9%,53.7%,51.8%,39.4%,18.7% and 8.3%)without deformities or retardation of growth .The delivered new born mice grew into adults;their developmental time series,and the parameters of physiological functions were not aberrant compared with that of the control group ( P<0.05 ) .Cancer bearing mice'concordance,movement,reflection, cognition,forced grasping wire and escarpment avoidance decreased; acouesthesi startle,opening-eyes time extended and the body weight increased with that of no cancer bearing mice.Furthermore,(D14~D16)all major organs developed well.The HE staining revealed that the major organs,including the lung ,the liver and the kidney,wer histologically normal.The experimental group (D17~D19) were 5.6%, 40.3% and 46.9% of tumorigenic rate (P <0.05);we coule see ascites and block at planting sites after delivery 7 days. D19 group was the largest.The HE staining revealed that the lung and the liverwere histologically abnormal,but the kindey was not abnormal.(2)The developmental indexes indicated that there were significant difference between experimental groups,control group and blank group;Furthermore,the body weight of control group and expemental group were more than blank group;control group was the most. The total mortality and weekly mortality of experimental groups and control group both were higher than that of bland group(P<0.05). we coule see ascites and block at planting sites after delivery 7 days. The experimental group and 4days control group,furthermore, There were no significant difference in the tumor growth rate between experimental group and control group(P>0.05).The HE staining revealed that the lung and muscle were histologically cancer bearing ,but the liver and the kindey were not abnormal.The lung's cancer bearing rates in the experimental group was lower than that of control group.(3)Amniotic fluid significantly inhibited the cell proliferation as well as the PCNA expressions ,increased the P53 expressions and induced cell apoptosis.The amniotic fluid collected on D11 was the most effective ,while that of D17 was the least effective (P<0.05).There was a negative corrected. Between PCNA and P53 expressions(r=-0.808,P<0.01). Conclusions ( 1 ) Mid-late embryonic microenvironment can significantly reduce invasiveness and tumorigenicity of the tumor H22 cells in vivo.Furthermore,tumor H22 cells can affect embryonic development. ( 2 ) Mice transplanted with H22 cells twice were later in tumorigenesis and survival time were longer than mice transplanted H22 cells one time which may be related that embryonic microenvironment could inhibit the tumor cells.(3)Amniotic fluid on gestational day 11,13,15and 17can inhibit the proliferation of H22 cells and induce their apoptosis. Cues associated with PCNA and P53 expressions possibly, furthermore, the earlier pregnancy and the longer interrention is more significant the effects of amniotic fluid are.