ABSTRACT
Aim To analyze differential expression and interaction of β-catenin/ICAT proteins in HL60 cells when they were induced into monocytic differentiation, and to figure out the mechanism of NSC67657 in cellu-lar induction. Methods HL60 cells were treated by 10 μmol · L-1 NSC67657 , and cellular differentiation could be observed by cytochemical staining and flow cytometry. Then, RT-PCR and Western blot were em-ployed to determine the differential expression of β-catenin/ICAT genes and proteins. Co-immunoprecipi-tation assay was used to confirm the interaction of β-catenin/ICAT proteins, and laser co-focus light mi-croscopy technology was used to co-indentify proteins differential expression and intracellular location. Re-sults HL60 cells could be induced into monocytic dif-ferentiation after 5 days treatment using 10μM NSC67657 . The CD14 ( +)% cells could be up to o-ver 90%, and cytochemical staining reports were con-sistent with this result. The expressions of ICAT gene and protein were up-regulated significantly ( P <0. 01 ) , but the expressions ofβ-catenin gene and pro-tein, on the contrary, were down-regulated(P<0. 05) when HL60 cells were induced into monocytic differen-tiation. From co-immunoprecipitation assay findings, ICAT protein interacted with β-catenin protein, and the absorbance of protein electrophoresis bands in-creased in differentiated cells. From laser co-focus light microscopy assay findings, the fluorescence of ICAT and β-catenin protein could be both observed in cytoplasm and nucleus. In drug treated HL60 cells, the fluorescence of ICAT protein was enhanced both in cytoplasm and nucleus, however, the fluorescence ofβ-catenin protein, which looked like transferring into different organelles, decreased significantly in nucleus, but increased in cytoplasm. Conclusions HL60 cells could be induced into monocytic differentiation by NSC67657 and β-catenin/ICAT proteins differentially expressed during cellular differentiation. The enhanced interaction of β-catenin/ICAT proteins and β-catenin protein transferring from nucleus into cytoplasm indi-cates that NSC67657 probably induces HL60 cells into monocytic differentiation through down-regulating β-catenin protein and blockingβ-catenin protein from nu-cleus.
ABSTRACT
Objective To investigate the etiologic roles of apoptosis-associated genes,environmental factors and their interactions in lumbar disc herniation (LDH).Methods A case-control trial was conducted.We recruited 128 outpatients with LDH as case group and 132 normal people matched by age and gender as control group.Peripheral venous blood samples were collected and DNA was extracted from leukocytes.By using a modified Brucker Autoflex MALDI-TOF mass spectrometer,we analyzed 3 genes with 9 polymorphic sites,namely,Fas-1377G/A rs2234767,Fas-670G/A rs1800682,Fas rs2147420,Fas rs2296603,Fas rs7901 656,Fas rs1 57101 9,FasL-844C/T rs7631 10,CASP-9-1263A > G rs4645978,and CASP-9-712C > T rs4645981.The correlations between polymorphism of Fas,FasL and CASP-9 genes and the risk of LDH were evaluated by non-conditional Logistic regression model.Multiple Logistic regression model was performed to assess the interaction between apoptosis-associated genes and environment factors,such as lumbar vertebral loads,bed type,spare-time exercises and spare-time activities. Results There were preferable balances in case and control groups in age and gender without significant differences.However,the two groups differed significantly (P G (rs4645978),and FasL-844C/T TT and CASP-9-1263A>G GG genotypes might be the high risk genotypes of LDH.The gene-environment interaction analysis revealed that super-multiplicative and sub-multiplicative interactions respectively between FasL-844TT genotype and lumbar vertebral loads (3-4 level),and between CASP-9-rs4645978 GG and lumbar vertebral loads (3-4 level).Conclusion FasL,CASP-9 genes and lumbar vertebral loads and their interactions play important roles in the pathogenesis of LDH.It suggests that the risk of LDH may be codetermined by environmental factors and inherited susceptibility genes,and that the mechanisms of interactions vary in different genotypes and the same or different environmental factors.