ABSTRACT
This study is to investigate the tumor invasion and metastasis inhibition effects of the immunoconjugate composed of lidamycin and anti-type IV collagenase monoclonal antibody Fab' fragment. Boyden chamber assay was used to evaluate the influence of Fab'-LDM on HT-1080 cells invasion ability, gelatinase spectrum was used to measure the change of invasion factor MMP-2 and MMP-9's secretion, and RT-PCR was adopted to determine TIMP-1 mRNA expression level. The immunoconjugate inhibition of tumor in situ metastasis was also tested in nude mice. The Fab'-LDM conjugates had dose-dependent inhibition effect on HT-1080 cells' invasion. At the concentrations of 5 and 10 nmol L(-1), the Fab'-LDM inhibited the invasion by (60 +/- 12) % and (79 +/- 11) % respectively. At the concentration of 5 and 10 nmol L(-1), the Fab'-LDM inhibited the secretion of MMP-2 by (42 +/- 8) % and (54 +/- 6) % and that of MMP-9 by (57 +/- 3) % and (87 +/- 1) %, respectively. RT-PCR indicated that conjugates increased the anti-invasion factor TIMP-1 level. The in vivo experiment showed that, compared with the control group, the tumor inhibition rate in Fab', Fab'-LDM, and LDM group equaled to (30 +/- 13) %, (86 +/- 26) %, (74 +/- 22) % respectively. In conclusion, Fab'-LDM could inhibit the invasion and metastasis of tumor and it might be a new tumor biotherapy agent.
Subject(s)
Animals , Humans , Mice , Aminoglycosides , Pharmacology , Antibiotics, Antineoplastic , Pharmacology , Antibodies, Monoclonal , Allergy and Immunology , Cell Line, Tumor , Enediynes , Pharmacology , Fibrosarcoma , Metabolism , Pathology , Immunoconjugates , Pharmacology , Immunoglobulin Fab Fragments , Pharmacology , Matrix Metalloproteinase 2 , Allergy and Immunology , Bodily Secretions , Matrix Metalloproteinase 9 , Allergy and Immunology , Bodily Secretions , Mice, Inbred BALB C , Mice, Nude , Neoplasm Invasiveness , Neoplasm Metastasis , Neoplasm Transplantation , RNA, Messenger , Metabolism , Tissue Inhibitor of Metalloproteinase-1 , Genetics , Metabolism , Tumor BurdenABSTRACT
To investigate the antitumor activities of the immunoconjugates composed of anti-type IV collagenase monoclonal antibody Fab' fragment and lidamycin (LDM) prepared with different linkers. The immunoconjugates were prepared by linking Fab' to lysine-69 of LDM apoprotein by SPDP, LCSPDP, SMBS or SSMPB as the intermediate drug linkers. Immunoreactivities of the conjugates were determined by ELISA. The cytotoxicities of the conjugates were examined by clonogenic assay. In vivo antitumor effects of the conjugates were evaluated in nude mice bearing subcutaneously implanted HT-1080 tumor. ELISA assay showed that the conjugates retained part of the immunoreactivity of 3G11 against the antigen. The cytotoxicities of the Fab'-SMBS-LDM and Fab'-SSMPB-LDM to HT-1080 cells were significantly potent, compared with Fab'-SPDP-LDM, Fab'-LCSPDP-LDM and free LDM. In animal models at the same condition, free LDM, Fab'-SPDP-LDM and Fab'-LCSPDP-LDM inhibited the growth of HT-1080 tumor by 70.9%, 74.8% and 72.3%, while Fab'-SMBS-LDM and Fab'-SSMPB-LDM reached 78.0% and 87.7%, respectively. The median survival time of the mice treated with free LDM, Fab'-SPDP-LDM and Fab'-LCSPDP-LDM were prolonged by 71.9%, 82.2% and 107.5%, respectively, compared with that of untreated group. Whereas, the median survival time of Fab'-SMBS-LDM and Fab'-SSMPB-LDM were prolonged by 145.2% and 165.8%, respectively, indicating that Fab'-SSMPB-LDM was more effective than Fab'-SMBS-LDM in tumor suppression and life span prolongation. Fab'-SSMPB-LDM has more marked selective antitumor efficacy and lower toxicity, and might be a novel candidate for cancer therapy.
Subject(s)
Animals , Humans , Mice , Aminoglycosides , Pharmacology , Antibiotics, Antineoplastic , Pharmacology , Antibodies, Monoclonal , Allergy and Immunology , Cell Line, Tumor , Cell Proliferation , Collagenases , Allergy and Immunology , Enediynes , Pharmacology , Fibrosarcoma , Pathology , Immunoconjugates , Pharmacology , Immunoglobulin Fab Fragments , Allergy and Immunology , Matrix Metalloproteinase Inhibitors , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Tumor BurdenABSTRACT
In this study, the antitumor activities of VEGF shRNA and tubulin inhibitors on human prostate cancer DU145 cells was investigated, and shRNA transient expression plasmid pCSH1-VEGF targeting VEGF mRNA was constructed. The silence efficiency of pCSH1-VEGF was detected by RT-PCR assay, Western blotting, and Matrigel invasion assay. The sensitivity change of DU145 cells to Taxol and vincristine (VCR) was measured by MTT assay. To detect the effects of pCSH1-VEGF and Taxol in vivo, nude mice model of DU145 xenograft tumor was established by subcutaneous inoculation. The results showed that transcription and expression of VEGF were knocked by pCSH1-VEGF in DU145 cells. Matrigel invasion assay results showed that pCSH1-VEGF significantly reduced the migration of DU145 cells with inhibitory rate of 56.1%. Furthermore, pCSH1-VEGF enhanced the sensitivity of DU145 cells to Taxol and vincristine, and the values of IC50 decreased by 77.3% and 92.6%, respectively. In vivo experiment showed that Taxol, pCSH1-VEGF, combination of pCSH1-VEGF and Taxol inhibited tumor growth by the rates of 48.8%, 56.2% and 81.8%, respectively. The coefficient of drug interaction (CDI) of pCSH1-VEGF and Taxol was 0.82. The data suggested that VEGF shRNA could significantly enhance the sensitivity of human prostate cancer to tubulin inhibitors.
Subject(s)
Animals , Humans , Male , Mice , Antineoplastic Agents, Phytogenic , Pharmacology , Cell Line, Tumor , Cell Movement , Cell Proliferation , Genetic Vectors , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Paclitaxel , Pharmacology , Plasmids , Prostatic Neoplasms , Metabolism , Pathology , RNA Interference , RNA, Small Interfering , Genetics , Transfection , Tubulin Modulators , Pharmacology , Tumor Burden , Vascular Endothelial Growth Factor A , Genetics , Metabolism , Vincristine , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To induce hepatic differentiation of human adipose-derived stem cells (hADSCs) in vitro.</p><p><b>METHODS</b>hADSCs were isolated from human adipose tissue and treated with improved hepatic medium containing HGF, bFGF and FGF4. After 7 days of culture, OSM was added to the culture media. Cell growth during hepatic differentiation was evaluated by CCK8 assay. Morphology of differentiation was examined under light microscope. Liver specific genes and proteins were detected by RT-PCR analysis and immunohistochemical staining, respectively. And functional characteristics of hepatocytes were also examined.</p><p><b>RESULTS</b>The number of hADSCs cultured in the improved hepatic media was increased significantly in comparison to hADSCs cultured in control media from 5 days to 21 days (t=6.59, 8.69, 15.94 and 24.64, respectively, P<0.05). The hADSCs-derived hepatocyte-like cells exhibited hepatocyte morphology, expressed hepatocyte markers, possessed hepatocyte-specific activities, such as uptake and excretion of indocyanine green, glycogen storage and albumin production.</p><p><b>CONCLUSION</b>hADSCs can be induced into hepatocyte-like cells in this differentiation system. And this differentiation system promoted the growth of hADSCs.</p>
Subject(s)
Humans , Adipose Tissue , Cell Biology , Albumins , Metabolism , Cell Culture Techniques , Cell Differentiation , Cell Proliferation , Cell Separation , Cells, Cultured , Culture Media , Fibroblast Growth Factor 2 , Pharmacology , Hepatocyte Growth Factor , Pharmacology , Hepatocytes , Cell Biology , Metabolism , Mesenchymal Stem Cells , Cell Biology , Reverse Transcriptase Polymerase Chain Reaction , alpha-Fetoproteins , MetabolismABSTRACT
This study is to investigate the antitumor activities of the immunoconjugates composed of anti-type IV collagenase monoclonal antibody 3G11 and lidamycin (LDM) prepared by different methods. The immunoconjugates were prepared by linking 2-iminothiolane modified 3G11 to lysine-69 of LDM apoprotein by SPDP and SMBS as the intermediate drug linker. Immunoreactivity of the conjugates was determined by ELISA. The cytotoxicity of the conjugates was examined by clonogenic assay. Antitumor effects of the conjugates in vivo were evaluated in nude mice bearing subcutaneously implanted HT-1080 tumor. ELISA assay showed that the immunoconjugates retained the immunoreactivity of 3G11 against type IV collagenase. The cytotoxicity of the 3G11-SMBS-LDM to HT-1080 cells was significantly more potent than that of free LDM and 3G11-SPDP-LDM. In animal model at the same condition, free LDM inhibited the growth of HT-1080 tumor by 71.2%, while 3G11-SPDP-LDM and 3Gl1-SMBS-LDM reached 77.1% and 86.1%, respectively. The median survival time of the mice treated with free LDM was prolonged by 71.9% compared with that of untreated group. Whereas, the median survival time of 3G11-SPDP-LDM and 3G11-SMBS-LDM was prolonged by 125.3% and 163.7%, respectively, indicating that 3G11-SMBS-LDM was more effective than 3G11-SPDP-LDM in tumor suppression and life span prolongation. 3Gll-SMBS-LDM has more selective antitumor efficacy and lower toxicity, and might be a novel candidate for cancer therapy. LDM was more effective than 3G11-SPDP-LDM in tumor suppression and life span prolongation. 3Gll-SMBS-LDM has more selective antitumor efficacy and lower toxicity, and might be a novel candidate for cancer therapy.
Subject(s)
Animals , Humans , Mice , Aminoglycosides , Therapeutic Uses , Antibiotics, Antineoplastic , Therapeutic Uses , Antibodies, Monoclonal , Allergy and Immunology , Cell Line, Tumor , Cell Proliferation , Collagenases , Allergy and Immunology , Enediynes , Therapeutic Uses , Fibrosarcoma , Pathology , Therapeutics , Immunoconjugates , Therapeutic Uses , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Tumor BurdenABSTRACT
<p><b>OBJECTIVES</b>To analyze the relationship between the fetus infection and HBV M, HBV DNA in amniotic fluid, umbilical cord blood, maternal blood and placenta, and to explore the mechanism of vertical transmission of HBV.</p><p><b>METHODS</b>Immunonetric assay and nucleic acid amplification hybri-comb were used. Both HBV M and HBV DNA were detected in amniotic fluid, vein blood, umbilical cord blood for each of 65 HBV-positive women in their different gestational periods, while immunohistochemical analysis was carried out on the tissue of placenta, liver, lung or heart from each abortive fetus/dead infant in the case.</p><p><b>RESULTS</b>For all of the 65 HBsAg-positive women in their different gestational periods, the detected positive rate of HBsAg was 21.50% in amniotic fluid, and 20.00% in umbilical blood. The positive rate of HBsAg, HBeAg, Anti-HBc and HBV DNA detected in blood, amniotic fluid and umbilical blood was 6.15%. The cases with positive HBsAg, Anti-HBe, Anti-HBc and negative HBV DNA were in a percentage of 13.85%. Immunohistochemical analysis on placentas after birth/abortion as well as the tissues of livers, lungs, hearts of the fetuses/dead infants in 4 cases of pregnant women with positive HBsAg, HBeAg, Anti-HBc or HBV DNA in blood, amniotic fluid or umbilical blood showed that HBsAg, HBcAg positive cells in the scope could be seen in every layer of the tissue of placenta, in the hepatic/pulmonary tissue, but not in the cardiac tissue.</p><p><b>CONCLUSION</b>The infection in amniotic fluid or placenta relates to HBV infection in fetus; intrauterine HBV may result in infection in organs such as blood, liver, or lung of a fetus; infection in the amniotic fluid may be another key route of the intrauterine infection of fetus, and the detection on HBV M or HBV DNA in amniotic may be used as one of diagnostic proofs of HBV infection of fetus in its early stage.</p>
Subject(s)
Adult , Female , Humans , Pregnancy , Amniotic Fluid , Virology , DNA, Viral , Hepatitis B , Diagnosis , Immunohistochemistry , Infectious Disease Transmission, Vertical , Placenta , Virology , Pregnancy Complications, Infectious , DiagnosisABSTRACT
@#ObjectiveTo investigate the application of CT inspection during the recovery period to cerebral infarct patients. Methods53 recurrence cases were taken out from random 100 cases of first reexamination CT films and analyzed while comparing the cases′ recurrence times,contents and the relationship between recurrence times and the original deceases.ResultsThere was no close connection between the rate of cerebral infarct recurrence and recurrence times(P>0.05).But cerebral infarct recurrence mainly appeared within the first year and the recurrence time caused by the presence of hypertension was shorter than the time caused by diabetes(P<0.05).92% cerebral infarct recurrence were cavity cerebral infarct recurrence and 11% were silent brain infarction.Conclusions It is significant for the patients to take CT reexamination within the first recovery year and also important for the no symptom patients.