ABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of T lymphoma invasion and metastasis inducing factor 1 antisense oligodeoxynucleotides (Tiam 1 ASODN) transfection on the morphology and invasive migration potential of gastric cancer cells.</p><p><b>METHODS</b>The higher invasive and migratory subgroup (M(H)) were separated from human gastric cancer cell line MKN-45 (M(0)) by laminin adhesion method in vitro. Tiam 1 ASODN was transfected into M(H) cells with liposome, and the expression of Tiam 1 mRNA and protein was determined by RT-PCR and flowcytometry respectively. The changes in morphology, the invasive and migratory potential between Tima 1 ASODN transfected M(H) cells and no transfected M(H) cells were observed by HE stain, cytoskeletal protein stain, scanning electronic microscope (SEM) and Boyden chamber test.</p><p><b>RESULTS</b>Compared with the control, the expression of Tiam 1 mRNA and protein in M(H) cells was significantly decreased after transfected with 0.43 micromol/L ASODN(P< 0.01). The invasive and migratory potential of M(H) cells in vitro was also much more decreased than that of no transfected cells (P< 0.05 or P< 0.01). At the same time, transfected M(H) cells had less membrane surface projections, fewer or shorter pseudopodia, less irregular cytoskeletal network and less spotted-like actin bodys than no transfected M(H) cells did.</p><p><b>CONCLUSION</b>Tiam 1 ASODN transfection can effectively suppress the expression of Tiam 1 in gastric cancer cells and impair its invasive and migratory potential in vitro, which may be fulfilled through modulating the reconstruction of cytoskeleton and decreasing the deforming and migratory potential of gastric cancer cells.</p>
Subject(s)
Animals , Humans , Mice , Cell Line, Tumor , Flow Cytometry , Guanine Nucleotide Exchange Factors , Genetics , Mice, Inbred BALB C , Neoplasm Invasiveness , Oligonucleotides, Antisense , Genetics , RNA, Messenger , Genetics , Stomach Neoplasms , Genetics , Pathology , T-Lymphoma Invasion and Metastasis-inducing Protein 1 , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To explore the proliferation-promoting effect of sensory neuropeptide substance P (SP) on the cultured granulation tissue fibroblasts in vitro and its regulative effect on the gene expression of basic fibroblast growth factor (bFGF) mRNA.</p><p><b>METHODS</b>The proliferation-promoting effect of cultured granulation tissue fibroblasts was observed by means of MTT; the regulative effect of SP on gene expression of fibroblast bFGF by RT-PCR. The time and dose-efficiency relations were also observed.</p><p><b>RESULTS</b>There was a significant proliferation-promoting effect of SP on the cultured granulation tissue fibroblasts in vitro in a remarkable dose-dependent fashion. However, bFGF antibody only partly exerted its inhibitive effect. SP could induce the bFGF mRNA expression of the fibroblasts at the 3rd and 6th hour (P < 0.01). SP could promote the bFGF mRNA expression of the fibroblasts in the concentration of 10(-9) - 10(-5) mol/L and peaked in the concentration of 10(-7) mol/L.</p><p><b>CONCLUSIONS</b>SP has a significant proliferation-promoting effect on the granulation tissue fibroblasts, which is correlated with SP inducing bFGF mRNA expression of fibroblasts.</p>
Subject(s)
Animals , Male , Rats , Cell Division , Dose-Response Relationship, Drug , Fibroblast Growth Factor 2 , Genetics , Fibroblasts , Cell Biology , Metabolism , Gene Expression , Granulation Tissue , Cell Biology , Metabolism , RNA, Messenger , Genetics , Metabolism , Rats, Wistar , Reverse Transcriptase Polymerase Chain Reaction , Substance P , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of substance P (SP) on gene expression of transforming growth factor beta-1 (TGFbeta-1), transforming growth factor receptor-1 (TGFR-1) and transforming growth factor receptor-2 (TGFR-2) in fibroblasts cultured in vitro from rat's granulation tissues.</p><p><b>METHODS</b>The fibroblasts from the granulation tissues in the skeletal muscle of rat's hind limbs injured by formaldehyde were cultured in vitro. When different concentrations (10(-9)-10(-5) mol/L) of SP were added into the culture medium, the changes of gene expression of TGFbeta-1, TGFR-1 and TGFR-2 in the cultured fibroblasts were observed with reverse transcription polymerase chain reaction at different intervals (0, 3, 6, 12 and 24 hours after incubation).</p><p><b>RESULTS</b>The gene expression of TGFbeta-1, TGFR-1 and TGFR-2 in the fibroblasts cultured from rat's granulation tissues was up-regulated by SP. The peak level of the mRNA expression was found at 10(-8) mol/L SP and the up-regulation effect was not found at 10(-5) mol/L and 10(-6) mol/L. The peak levels of gene expression of TGFbeta-1, TGFR-1 and TGFR-2 in the fibroblasts treated with SP were achieved at 6 and 12 hours, respectively.</p><p><b>CONCLUSIONS</b>SP has up-regulation effect on the gene expression of TGFbeta-1, TGFR-1 and TGFR-2 in fibroblasts from rat's granulation tissues in vitro, and the effect is related to different stimulating concentrations of SP. It may be concerned with proliferation and differentiation of fibroblasts and formation of scar tissues during wound healing.</p>
Subject(s)
Animals , Female , Male , Rats , Analysis of Variance , Base Sequence , Cells, Cultured , Fibroblasts , Models, Animal , Molecular Sequence Data , Polymerase Chain Reaction , Methods , Probability , RNA, Messenger , Rats, Wistar , Receptors, Transforming Growth Factor beta , Genetics , Sensitivity and Specificity , Substance P , Pharmacology , Wound Healing , PhysiologyABSTRACT
<p><b>OBJECTIVE</b>To explore the regulative effects and significance of neuropeptide substance P (SP) on the expression of basic fibroblast growth factor (bFGF) of granulation tissue fibroblasts in vitro.</p><p><b>METHODS</b>A local aseptic inflammation was induced by injection of formaldehyde in rats, and its granulation tissue was cultured. RT-PCR was employed to observe expression of bFGF mRNA after inducement of SP at different concentrations and time points in the granulation tissue, and western blot to assay expression of bFGF protein.</p><p><b>RESULTS</b>The expression of bFGF mRNA was markedly increased significantly 3 and 6 hours after inducement with SP in 10(-7) mol/L, compared with control group (P < 0.01). The expression of bFGF protein was markedly higher than the control group after 12 hours, and it reached the peak at the 24th hour and declined gradually after 48 hours. SP at concentrations of 10(-9) - 10(-5) mol/L could significantly promote the expression of bFGF mRNA, and that at 10(-8) - 10(-5) mol/L induce the expression of bFGF protein. Both expressions reached the peak when SP concentration was 10(-7) mol/L (P < 0.01).</p><p><b>CONCLUSION</b>SP can induce the expressions of bFGF mRNA and bFGF protein of granulation tissue fibroblasts in vitro, which may possess an important significance in wound healing.</p>