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BACKGROUND: Psoriasis is an autoimmune disease that is caused by a shift in the Th1/Th2 balance toward Th1-dominant immunity. It has been established as an effective treatment to counteract psoriasis by subcutaneous injection of recombinant interleukin (IL)-4, and IL-4 gene therapy by topical transdermal penetration has shown its antipsoriatic effect in mice. Retinoic acid (RA) and dimethylsulfoxide can increase the efficiency of gene transfection in the topical transdermal delivery system. OBJECTIVE: We investigated whether RA could improve anti-psoriasis efficiency using IL-4 expression plasmid pORF-mIL-4 (pIL-4) via transdermal delivery system in K14-vascular endothelial growth (K14-VEGF) factor transgenic mice. METHODS: After pretreatment with RA, plasmid pIL-4 in 10% dimethylsulfoxide was applied to the ear skin by topical transdermal penetration. Hematoxylin- eosin staining and immunohistochemistry were performed with ear samples to evaluate anti-psoriasis efficiency in mice. RESULTS: The psoriasis pathological features were relieved and psoriasis-associated factors were significantly reduced. CONCLUSION: Our results reveal that topical application of pIL-4 in dimethylsulfoxide by transdermal delivery with RA pretreatment can improve psoriasis significantly.
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BACKGROUND: The use of adenoviral vector for gene therapy is still an important strategy for advanced cancers, however, the lack of the requisite coxsackie-adenovirus receptor in cancer cells and host immune response to adenovirus limit the application of adenoviral vector in vivo. METHOD: We designed the antiangiogenic gene therapy with recombinant PEDF adenovirus (Ad-PEDF) encapsulated in cationic liposome (Ad-PEDF/Liposome), and investigated the anti-tumor efficacy of Ad-PEDF/Liposome complex on inhibition of tumor metastasis. RESULTS: We found that systemic administration of Ad-PEDF/liposome was well tolerated and resulted in marked suppression of tumor growth, and was more potent than uncoated Ad-PEDF to induce apoptosis in B16-F10 melanoma cells and inhibit murine pulmonary metastases in vivo. After Ad-luciferase was encapsulated with liposome, its distribution decreased in liver and increased in lung. The anti-Ad IgG level of Ad-PEDF/Liposome was significantly lower than Ad-PEDF used alone. CONCLUSION: The present findings provide evidences of systematic administration of cationic liposome-encapsulated Ad-PEDF in pulmonary metastatic melanoma mice model, and show an encouraging therapeutic effect for further exploration and application of more complexes based on liposome-encapsulated adenovirus for more cancers.
Assuntos
Adenoviridae/genética , Proteínas do Olho/genética , Terapia Genética/métodos , Lipossomos/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/terapia , Melanoma/patologia , Melanoma/terapia , Fatores de Crescimento Neural/genética , Serpinas/genética , Animais , Cátions , Feminino , Vetores Genéticos , Imunoglobulina G/química , Lipossomos/química , Melanoma Experimental , Camundongos , Camundongos Endogâmicos C57BL , Metástase NeoplásicaRESUMO
Thermotherapy and thermochemotherapy have been used in clinics to treat patients with malignant diseases, including colon cancer, and their efficacy has been well proved. Heat shock proteins (HSPs), especially Hsp70, play important roles in neutralizing their efficacy. It has been reported that quercetin can suppress cancer by inhibiting the intratumoral expression of Hsp70. This study was designed to investigate whether quercetin could enhance sensitivity to thermotherapy and thermochemotherapy. Soluble quercetin liposome was used in this study. The effects of quercetin were investigated in vitro and in mouse colon cancer models of subcutaneous tumor and peritoneal carcinomatosis. The results showed that quercetin liposome inhibited the upregulation of Hsp70 and enhanced apoptosis induced by hyperthermia and thermochemotherapy. Systemic administration of quercetin liposome can sensitize CT26 cells to thermotherapy and chemothermotherapy. This study suggests that quercetin liposome might be potentially applied for clinical cancer therapy.
Assuntos
Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/terapia , Hipertermia Induzida/métodos , Lipossomos/farmacologia , Quercetina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Terapia Combinada , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genéticaRESUMO
Hepatocellular carcinoma is one of the most common cancers in worldwide. We previously reported a novel thienopyridine derivative 3-amino-6-(3,4-dichlorophenyl) thieno[2,3-b]pyridine-2-carboxamide (SKLB70359) which possesses anticancer activity against hepatocellular carcinoma. In present study, we further investigated its anticancer activity and possible mechanism. The SKLB70359 treatment decreased the viability of a panel of hepatocellular carcinoma cell lines in a concentration- and time-dependent manner with IC(50) 0.4 ~ 2.5 µM. The mechanism study showed that SKLB70359 induced G0/G1 cell cycle arrest and then led to apoptotic cell death of HepG2 cell. The SKLB70359 induced G0/G1 cell cycle arrest was characterized by down-regulation of cyclin-dependent kinase 2 (CDK2), CDK4, CDK6 expression and up-regulation of p53, p21(WAF1). Activating of caspase-3 and caspase-9 was also observed. Meanwhile, proliferation inhibitory effect of SKLB70359 was associated with decreased level of phosphorylated p44/42 mitogen activated protein kinase (p44/42 MAPK) and phosphorylated retinoblastoma protein (Rb). Moreover, SKLB70359 exhibit less toxicity to non-cancer cells than tumor cells. In conclusion, the findings in this study suggested that SKLB70359 have potential anticancer efficacy via G0/G1 cell cycle arrest and apoptosis induction. Its potential to be a candidate of anticancer agent is worth being further investigated.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Piridinas/farmacologia , Tiofenos/farmacologia , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Caspase 3/metabolismo , Caspase 9/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Células HCT116 , Células HEK293 , Células HeLa , Células Hep G2 , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação , Piridinas/química , Piridinas/uso terapêutico , Proteína do Retinoblastoma/metabolismo , Tiofenos/química , Tiofenos/uso terapêutico , Proteína Supressora de Tumor p53/metabolismo , Regulação para CimaRESUMO
AIM: To investigate the therapeutic effect of the plasmid pcDNA3.1-IL15 complexed with cationic liposome (CL-IL15) in the B16-F10 melanoma lung metastasis model. METHODS: A plasmid with high secretive efficiency of IL-15 was constructed and the optimum mix ratio was determined to formulate cationic liposome-plasmid complex with the optimal encapsulation. The CHO-K1 cell line was transfected by CL-IL15. The secretion of transfected IL-15 gene was detected by Western blot and its biological function was measured through the proliferation response of CTLL-2 cytotoxic T cell line of murine by MTT assay. The C57BL/6 mice were inoculated intravenously (i.v.) with B16-F10 melanoma lung metastasis cells then treated (i.v.) by CL-IL15 in a therapeutic setting to determine the tumorigenesis and research the corresponding mechanisms. RESULTS: The pcDNA3.1-IL15 plasmid was successfully constructed and the mass-ratio of optimal condition of cationic liposome-plasmid with perfect entrapment was 1:5 (plasmid: cationic liposome). Western blot analysis displayed the detection of IL-15 both in the medium and the pcDNA3.1-IL15 transfected cells. MTT assay showed that CTLL-2 cells could proliferate with the medium obtained from CHO-K1 cells transfected by CL-IL15. And the administration of CL-IL15 complexes led to the significant inhibition lung metastasis of malignant melanoma (P<0.05). CONCLUSION: CL-IL15 could inhibit the metastasis of malignant melanoma and the cationic liposome delivered plasmid pcDNA3.1-IL-15 complexes may be an efficient therapeutic strategy for the treating of lung metastasis. And the effective splenic cell-mediated cytotoxicity and the obvious NK cells recruitment may be involved.
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Interleucina-15/genética , Neoplasias Pulmonares/secundário , Neoplasias Pulmonares/terapia , Plasmídeos/genética , Animais , Células CHO , Cátions , Cricetinae , Citotoxicidade Imunológica/imunologia , Modelos Animais de Doenças , Terapia Genética , Interleucina-15/imunologia , Interleucina-15/metabolismo , Células Matadoras Naturais/imunologia , Lipossomos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Melanoma Experimental/secundário , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Plasmídeos/administração & dosagemRESUMO
Inducing apoptosis is a promising therapeutic approach to overcome cancer. Here we described that a novel synthesized compound, 3-amino-N-(4-chlorobenzyl)-6-(3-methoxyphenyl)thieno[2,3-b]pyridine-2-carboxamide (SKLB703), exhibits antitumor activity via inducing apoptosis both invitro and invivo. Our results showed that SKLB703 inhibited the proliferation of a panel of human cancer cell lines, and human hepatocellular carcinoma cell line HepG2 was the most sensitive. The proliferation inhibitory effect of SKLB703 was associated with its apoptosis-inducing effect by activating caspase-3 and caspase-9 rather than caspase 8. Exposure of HepG2 to SKLB703 also resulted in Bax upregulation, Bcl-2 downregulation, cytochrome c release and mitochondrial transmembrane potential change in mitochondrial apoptotic pathway. Moreover, the decrease of phosphorylated p 44/42 mitogen-activated protein kinase and phosphorylated Akt was observed. SKLB703 suppressed the growth of established tumors in xenograft models in mice, whereas no toxicity was exhibited. TUNAL analysis showed that SKLB703 induced HepG2 tumor apoptosis. Taken together, the present study demonstrates that SKLB730 exhibits its antitumor activity through inducing apoptosis via mitochondrial apoptotic pathway. Its potential to be a candidate of anticancer agent is worth being further investigated.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Piridinas/farmacologia , Tiofenos/farmacologia , Animais , Antineoplásicos/química , Western Blotting , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Caspases/metabolismo , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Feminino , Células Hep G2 , Humanos , Células K562 , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/fisiologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosforilação/efeitos dos fármacos , Piridinas/química , Ratos , Ratos Sprague-Dawley , Tiofenos/química , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Antagonizing angiogenesis-related receptor tyrosine kinase is a promising therapeutic strategy in oncology. In present study, we designed and synthesized a novel vascular endothelial growth factor receptor (VEGFR) inhibitor N-methyl-4-(4-(3-(trifluoromethyl) benzamido) phenoxy) picolinamide SKLB610 that potently suppresses human tumor angiogenesis. SKLB610 inhibited angiogenesis-related tyrosine kinase VEGFR2, fibroblast growth factor receptor 2 (FGFR2) and platelet-derived growth factor receptor (PDGFR) at rate of 97%, 65% and 55%, respectively, at concentration of 10µM in biochemical kinase assays. In vitro, SKLB610 showed more selective inhibition of VEGF-stimulated human umbilical vein endothelial cells (HUVECs) proliferation, and this proliferation inhibitory effect was associated with decreased phosphorylation of VEGFR2 and p42/44 mitogen-activated protein kinase (p42/44 MAPK). Antiangiogenic evaluation showed that SKLB610 inhibited the HUVECs capillary-tube formation on Matrigel in vitro and the sub-intestinal vein formation of zebrafish in vivo. Moreover, SKLB610 inhibited a panel of human cancer cells proliferation in a concentration-dependent manner and human non-small cell lung cancer cell line A549 and human colorectal cancer cell line HCT116 were most sensitive to SKLB610 treatment. In vivo, chronic intraperitoneally administration of SKLB610 at dose of 50mg/kg/d resulted in significant inhibition in the growth of established human A549 and HCT116 tumor xenografts in nude mice without exhibit toxicity. Histological analysis showed significant reductions in intratumoral microvessel density (CD31 staining) of 43-55% relative to controls depending on the specific tumor xenografts. In conclusion, the present study demonstrated that SKLB610 exhibited its antitumor activity as a multi-targeted inhibitor with more potent inhibition of VEGFR2 activity. Its potential to be a candidate of anticancer agent is worth being further investigated.
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Inibidores da Angiogênese/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Colorretais/tratamento farmacológico , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Neovascularização Patológica/tratamento farmacológico , Ácidos Picolínicos/farmacologia , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptores do Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Amidas/síntese química , Amidas/farmacologia , Inibidores da Angiogênese/síntese química , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Colágeno/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Combinação de Medicamentos , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Feminino , Humanos , Laminina/metabolismo , Camundongos , Camundongos Nus , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Fosforilação/efeitos dos fármacos , Ácidos Picolínicos/síntese química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Proteoglicanas/metabolismo , Receptor Tipo 2 de Fator de Crescimento de Fibroblastos/metabolismo , Receptores do Fator de Crescimento Derivado de Plaquetas/metabolismo , Veias Umbilicais/citologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-ZebraRESUMO
PURPOSE: Patients with localized prostate cancer can usually achieve initial response to conventional treatment. However, most of them will inevitably progress to advanced disease stage. There is a clear need to develop innovative and effective therapeutics for prostate cancer. Mouse survivin T34A (mS-T34A) is a phosphorylation-defective Thr34 â Ala dominant negative mutant, which represents a potential promising target for cancer gene therapy. This study was designed to determine whether mS-T34A plasmid encapsuled by DOTAP-chol liposome (Lip-mS) has the anti-tumor activity against prostate cancer, if so, to further investigate the possible mechanisms. METHODS: In vitro, TRAMP-C1 cells were transfected with Lip-mS and examined for apoptosis by PI staining and flow cytometric analysis. In vivo, subcutaneous prostate cancer models were established in C57BL/6 mice, which were randomly assigned into three groups to receive i.v. administrations of Lip-mS, pVITRO2-null plasmid complexed with DOTAP-chol liposome (Lip-null) or normal saline every 2 days for eight doses. Tumor volume was measured. Tumor tissues were inspected for apoptosis by TUNEL assay. Microvessel density (MVD) was determined by CD31 immunohistochemistry. Alginate-encapsulated tumor cell test was conducted to evaluate the treatment effect on angiogenesis. RESULTS: Administration of Lip-mS resulted in significant inhibition in the growth of mouse TRAMP-C1 tumors. The anti-tumor response was associated with increased tumor cell apoptosis and decreased microvessel density. CONCLUSIONS: The present study may be of importance in the exploration of the potential application of Lip-mS in the treatment of a broad spectrum of tumors.
Assuntos
Proteínas Inibidoras de Apoptose/genética , Mutação , Neoplasias da Próstata/terapia , Proteínas Repressoras/genética , Animais , Linhagem Celular Tumoral , Ácidos Graxos Monoinsaturados/administração & dosagem , Terapia Genética , Proteínas Inibidoras de Apoptose/uso terapêutico , Lipossomos/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microscopia de Fluorescência , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Compostos de Amônio Quaternário/administração & dosagem , Proteínas Repressoras/uso terapêutico , SurvivinaRESUMO
Metastatic breast cancers are hard to treat and almost always fatal. Chloroquine diphosphate, a derivative of quinine, has long been used as a potent and commonly used medicine against different human diseases. We therefore investigated the effects of chloroquine diphosphate on a highly metastatic mouse mammary carcinoma cell line. In vitro treatment of 4T1 mouse breast cancer cells with chloroquine diphosphate resulted in significant inhibition of cellular proliferation and viability, and induction of apoptosis in 4T1 cells in a time- and dose-dependent manner. Further analysis indicated that induction of apoptosis was associated with the loss of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-9 and caspase-3, and cleavage of poly(ADP-ribose) polymerase. The effect of chloroquine diphosphate was then examined using a mice model in which 4T1 cells were implanted subcutaneously. Chloroquine diphosphate (25mg/kg and 50mg/kg, respectively) significantly inhibited the growth of the implanted 4T1 tumor cells and induced apoptosis in the tumor microenvironment. Moreover, the metastasis of tumor cells to the lungs was inhibited significantly and the survival of the mice enhanced. These data suggested that chloroquine diphosphate might have chemotherapeutic efficacy against breast cancer including inhibition of metastasis.
Assuntos
Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Cloroquina/análogos & derivados , Neoplasias Mamárias Experimentais/tratamento farmacológico , Metástase Neoplásica/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cloroquina/farmacologia , Cloroquina/uso terapêutico , Citocromos c/metabolismo , Feminino , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Poli Adenosina Difosfato Ribose/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Colon carcinoma is one of the common malignant tumors and has high morbidity and mortality in the world. Pigment epithelial-derived factor (PEDF) has been found to be the most potent natural inhibitor of angiogenesis and PEDF gene has been extensively used for the therapy of tumors, which suggests a potential approach to the therapy of colon carcinoma. However, the transfer of PEDF gene largely depends on the effective gene delivery systems. Poly (lactic-co-glycolic acid) nanoparticles (PLGANPs) have been extensively used for gene therapy due to its low-toxicity, biocompatibility and biodegradability, due to its potential to be an excellent carrier of the PEDF gene. We investigated the effect of PEDF gene loaded in PLGA nanoparticles (PEDF-PLGANPs) on the mouse colon carcinoma cells (CT26s) in vitro and in vivo. Blank PLGANPs (bPLGANPs) showed lower cytotoxicity than PEI to the CT26s. In vitro, PEDF-PLGANPs directly induced CT26 apoptosis and inhibit human umbilical vein endothelial cell (HUVEC) proliferation. In vivo, PEDF-PLGANPs inhibited CT26 tumors growth by inducing CT26 apoptosis, decreasing MVD and inhibiting angiogenesis. Our present study demonstrates the inhibitory effect of PEDF-PLGANPs on the growth of CT26s in vitro and in vivo for the first time. PLGANP-mediated PEDF gene could provide an innovative strategy for the therapy of colon carcinoma.
Assuntos
Carcinoma/terapia , Neoplasias do Colo/terapia , Proteínas do Olho/genética , Técnicas de Transferência de Genes , Terapia Genética/métodos , Ácido Láctico/química , Nanopartículas , Fatores de Crescimento Neural/genética , Ácido Poliglicólico/química , Serpinas/genética , Animais , Apoptose/efeitos dos fármacos , Carcinoma/irrigação sanguínea , Carcinoma/genética , Carcinoma/metabolismo , Carcinoma/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Dependovirus/genética , Células Endoteliais/efeitos dos fármacos , Proteínas do Olho/biossíntese , Vetores Genéticos , Humanos , Ácido Láctico/toxicidade , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/prevenção & controle , Fatores de Crescimento Neural/biossíntese , Ácido Poliglicólico/toxicidade , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Serpinas/biossíntese , Fatores de Tempo , Carga Tumoral/efeitos dos fármacosRESUMO
Basic fibroblast growth factor (bFGF) is a mitogen for endothelial cells, which participates in tumor angiogenesis. Active immunity against bFGF could be a promising approach for the biotherapy of cancer. Because bFGF is abundant in normal and malignant tissues, it is presumably difficult for normal bFGF to induce immunity due to self-tolerance. In addition, previous studies have shown that a complex consisting of a cationic liposome and a non-coding plasmid DNA can be used to stimulate innate immunity. This stimulation initiates a potent cytokine response, which can inhibit tumor growth. To investigate the effects of immunity against bFGF on murine colon carcinomas, we employed an N-, C-terminally truncated basic fibroblast growth factor (tbFGF, of human origin) as an antigen and a liposome-DNA complex as an adjuvant. After six immunizations, a robust bFGF-specific immune response was elicited. Subsequently, inhibition of tumor growth and a significant reduction in tumor vasculature were observed. The antitumor effect was confirmed by adoptive therapy of activated spleen cells from the immunized mice. In vitro, a CTL assay revealed that bFGF-specific cytotoxic T lymphocytes (CTL) resulted in the lysis of mouse microvascular endothelial cells (MS1) rather than that of the CT26 colorectal cancer cells. These results suggest that anti-angiogenesis treatment induced by a bFGF-specific CTLs against microvascular endothelial cells may be a useful method for cancer therapy.
Assuntos
Vacinas Anticâncer/administração & dosagem , Neoplasias do Colo/imunologia , Neoplasias do Colo/terapia , Fator 2 de Crescimento de Fibroblastos/imunologia , Sequência de Aminoácidos , Animais , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Neoplasias do Colo/patologia , Neoplasias do Colo/prevenção & controle , DNA/administração & dosagem , Feminino , Fator 2 de Crescimento de Fibroblastos/administração & dosagem , Fator 2 de Crescimento de Fibroblastos/genética , Vetores Genéticos/genética , Humanos , Imunidade Inata/imunologia , Lipossomos/administração & dosagem , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Fragmentos de Peptídeos/administração & dosagem , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Plasmídeos/genéticaRESUMO
BACKGROUND: Psoriasis is a chronic and relapsing inflammatory skin disease associated with various immunologic abnormalities. Repeated subcutaneous injection of interleukin-4 (IL-4) has been established as an effective treatment to counteract psoriasis. OBJECTIVE: We investigated whether gene therapy using IL-4 expression plasmid (pIL-4) via transdermal delivery was an alternative treatment for psoriasis. In our experiment, dimethylsulfoxide (DMSO) was used as a penetration enhancer. METHODS: At first, the penetration efficiency of the complex of reporter plasmid accompanied by DMSO was investigated both in vitro and in vivo. Then, the antipsoriasis efficiency of the treatment with pIL-4-DMSO was tested in mice. RESULTS: The expression of the reporter gene was detected in epidermis and dermis both in vitro and in vivo. More importantly, the psoriasis symptoms were relieved, and significant reductions in some psoriasis-associated factors were observed after pIL-4-DMSO treatment. CONCLUSION: We conclude that the topical application of pIL-4-DMSO can treat psoriasis to a significant extent.
Assuntos
Terapia Genética/métodos , Interleucina-4/genética , Psoríase/terapia , Transdução Genética/métodos , Administração Cutânea , Animais , Dimetil Sulfóxido/química , Feminino , Camundongos , Camundongos Transgênicos , Plasmídeos , Psoríase/patologiaRESUMO
OBJECTIVE: To explore the application of MHC-Ig/peptide polymer technique for detecting antigen-specific cytotoxic T lymphocytes (CTLs) in mice with experimental autoimmune uveitis (EAU). METHODS: B10RIII mice were immunized with an interphotoreceptor retinal-binding protein (IRBP) synthetic peptide (IRBP161-180). The in vivo primed T cells were separated and stained with MHC-Ig polymer combined with a panel of truncated peptides derived from IRBP161-180. The level of IRBP-specific CTLs cells was determined by FACS analysis. The CD8+ T cells were isolated from the primed T cells and stimulated with complex polymers containing MHC-Ig and various IRBP-derived peptides. The proliferation of CD8+ T cells was measured by H thymidine incorporation. The production of interferon- (IFN-) in the cell suspensions was measured by ELISA. RESULTS: The IRBP-specific CTLs were detected by MHC-Ig/peptide polymers. The MHC-Ig/IRBP168-177 peptide polymer dtected 12.3% specific CTLs, showing greater ability in stimulating proliferation of CTLs and production of IFN--than the other MHC-Ig/ peptide polymers (P < 0.01). The truncated 10-mer peptide, IRBP168-177, was the major antigenic epitope for the IRBP-specific CTLs. The MHC-Ig/IRBP168-177 peptide polymer detected the highest level(4.9% +/- 1.1%) of specific CTLs from peripheral blood mononuclear cells (PBMCs) at the acute stage of EAU. CONCLUSION: The MHC-Ig polymer technique is an effective instrument for detecting antigen-specific CTLs, with good sensitivity and specificity in EAU studies.
Assuntos
Doenças Autoimunes/imunologia , Proteínas do Olho/imunologia , Oligopeptídeos/imunologia , Proteínas de Ligação ao Retinol/imunologia , Linfócitos T Citotóxicos/imunologia , Uveíte/imunologia , Animais , Feminino , Imunoglobulina G/imunologia , Interferons/biossíntese , Camundongos , Camundongos Mutantes , Células Fotorreceptoras de Vertebrados/imunologia , PolímerosRESUMO
Zinc finger proteins have been implicated as transcription factors in the differentiation and development of cells and tissues in higher organisms. The classical C2H2 zinc finger motif is one main type of motif of zinc finger proteins. Our previous studies have shown that Zfp637, which comprises six consecutively typical and one atypical C2H2 zinc finger motifs, is highly expressed in undifferentiated or poorly differentiated cell lines, but is moderately or slightly expressed in normal tissues. We have also demonstrated that Zfp637 can promote cell proliferation. However, its role in the regulation of cell differentiation remains unknown. We report here that endogenous Zfp637 as well as mTERT is expressed in proliferating C2C12 myoblasts and that their expression is downregulated during myogenic differentiation. Constitutive expression of Zfp637 in C2C12 myoblasts increased mTERT expression and telomerase activity, and promoted the progression of the cell cycle and cell proliferation. By contrast, endogenous repression of Zfp637 expression by RNA interference downregulated the mTERT gene and the activity of telomerase, and markedly reduced cell proliferation. Overexpression of Zfp637 also inhibited the expression of myogenic differentiation-specific genes such as MyoD and myogenin, and prevented C2C12 myoblast differentiation. Our results suggest that Zfp637 inhibits muscle differentiation through a defect in the cell cycle exit by potentially regulating mTERT expression in C2C12 myoblasts. This may provide a new research line for studying muscle differentiation.
Assuntos
Diferenciação Celular/fisiologia , Proteínas de Ligação a DNA/fisiologia , Músculo Esquelético/citologia , Dedos de Zinco , Animais , Sequência de Bases , Western Blotting , Linhagem Celular , Proliferação de Células , Primers do DNA , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Camundongos , Músculo Esquelético/enzimologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Telomerase/metabolismoRESUMO
Radiosensitivity of tumors is due to a complex interaction of various factors, it has been reported that survivin also acts as a constitutive and inducible radioresistance factor in a panel of tumor cells and approaches designed to inhibit survivin expression or function may lead to tumor sensitisation to chemical and physical agents. Previously, we found that the plasmid encoding the phosphorylation-defective mouse survivin threonine 34-->alanine mutant complexed to DOTAP-chol liposome (Lip-mS) can suppress murine primary breast carcinoma. However, little is known regarding the biological effect of Lip-mS combined with radiation. The present study was designed to determine whether Lip-mS could enhance the anti-tumor activity of radiation. The Lewis Lung Carcinoma (LLC) cells treated with a combination of Lip-mS and radiation displayed apparently increased apoptosis compared with those treated with Lip-mS or radiation alone. Mice bearing LLC tumors were treated with intravenous injections of Lip-mS and radiation, the combined treatment significantly reduced mean tumor volume compared with either treatment alone. Moreover, the anti-tumor effect of Lip-mS combined with radiation was greater than their additive effect when compared with the expected effect of the combined treatment. These data suggest that inhibition of survivin using a dominant-negative mutant, survivin T34A, could sensitize LLC cells to radiation efficiently and the synergistic anti-tumor activity may in part result from increasing the apoptosis of tumor cells, inhibiting tumor angiogenesis and inducing a tumor-protective immune response in the combined treatment.
Assuntos
Carcinoma Pulmonar de Lewis/tratamento farmacológico , Carcinoma Pulmonar de Lewis/radioterapia , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/fisiologia , Tolerância a Radiação/genética , Animais , Apoptose , Colesterol/farmacologia , Terapia Combinada/métodos , Ácidos Graxos Monoinsaturados/farmacologia , Genes Dominantes , Proteínas Inibidoras de Apoptose , Lipossomos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Plasmídeos , Molécula-1 de Adesão Celular Endotelial a Plaquetas/biossíntese , Compostos de Amônio Quaternário/farmacologia , Radioterapia/métodos , Proteínas Repressoras , Survivina , Sais de Tetrazólio/farmacologia , Tiazóis/farmacologiaRESUMO
Lung cancer is one of the most highly malignant tumors, and a significant threat to human health. Lung cancer patients often exhibit tumor cell invasion and metastasis, which often render current treatments ineffective. Recently, the beneficial effects of low molecular weight heparin (LMWH) on cancer metastasis were reported in pre-clinical research studies. LMWH may be a potential drug for cancer therapy. However, the mechanism of LMWH on the invasion and metastasis of cancer has yet to be determined. This study investigated the effects of Fraxiparine on the proliferation, invasion and metastasis of the human lung adenocarcinoma A549 cell line. MTT assay and flow cytometry showed that Fraxiparine slightly inhibited the cell viability dose- and time-dependently, but did not arrest the A549 cells in the G1 phase nor induce early apoptosis. The transwell chamber assay showed that Fraxiparine significantly suppressed the invasion and migration of the A549 cells in vitro. Fraxiparine also markedly inhibited the adhesion of the A549 cells to Matrigel. The RT-PCR assay demonstrated that the reduction in invasion and metastasis may be related to the up-regulation of nm23-H1 and the down-regulation of the heparanase expression. Moreover, the RT-PCR assay and Western blot analysis demonstrated that down-regulation of the expression of integrin ß1 and ß3, as well as that of matrix metalloproteinase-2 and -9 may be responsible for the inhibition of the invasion and metastasis of A549 cells by Fraxiparine.
RESUMO
OBJECTIVE: To study the mechanisms of nongenomic effect of 17beta-estradiol on human spermatozoa. METHODS: The intracellular calcium ([Ca2+]i) in the spermatozoa was measured by flow cytometry after the spermatozoa was treated with the inhibitors of trans-membrane signaling transduction pathways and impermeable 17beta-estradiol (E2-BSA). Western blot was used to detect the activation of the signal proteins after the spermatozoa was treated with 1 x 10(-6) mol/L E2-BSA and tamoxifen, an estrogen receptor inhibitor. RESULTS: Adenylyl cyclase (AC) inhibitor SQ22536, phospholipase C (PLC) inhibitor U73122 and protein tyrosine kinase (TPK) inhibitor Genistein all deterred the increase of [Ca2+]i caused by E2-BSA. E2-BSA also increased the PLC protein and PKC protein significantly. Tamoxifen, an antagonist of estrogen receptor, did not inhibit the activation of PLC caused by E2-BSA. CONCLUSION: The E2-BSA has an effect on human spermatozoa in a nongenomic pathway, possibly through the transmembrane signal transduction in relation to AC, PLC and TPK.
Assuntos
Cálcio/metabolismo , Estradiol/farmacologia , Transdução de Sinais , Espermatozoides/efeitos dos fármacos , Adenina/análogos & derivados , Adenina/farmacologia , Inibidores de Adenilil Ciclases , Adulto , Sítios de Ligação , Estrenos/farmacologia , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirrolidinonas/farmacologia , Tamoxifeno/farmacologia , Fosfolipases Tipo C/antagonistas & inibidoresRESUMO
PURPOSE: Human neutrophil peptides (HNP1-3), small molecular antimicrobial peptides, are expressed within tumors and associated with tumor necrosis and inhibition of angiogenesis. Recent investigations have suggested that HNP1-3 are likely to be involved in the host immune responses to tumors. EXPERIMENTAL DESIGN: We used recombinant pSec-HNP1, which expresses a secretable form of HNP1, to obtain expression of HNP1 in the tumor milieu in immunocompetent mice to explore the possible roles of HNP1 in tumor immunity. The antitumor effects were investigated in established CT26 colon cancer and 4T1 breast cancer models. RESULTS: HNP1-mediated chemotactic and activating effects on immature dendritic cells were detected both in vitro and in vivo. Intratumoral expression of HNP1 resulted in not only significant tumor growth inhibition but also increased CTL infiltration within tumors. Adoptive transfer of splenocytes and a (51)Cr release assay revealed specific cellular immune responses. Furthermore, increased antibodies were also found in sera from pSec-HNP1-treated mice supporting specific humoral immune responses. Increased apoptosis and decreased angiogenesis were also shown in treated tumors. CONCLUSIONS: These findings indicate that HNP1 can exert multiple antitumor effects through different mechanisms; more importantly, HNP1 mediates host immune responses to tumors in situ through the recruitment and subsequent activation of immature dendritic cells and thus shows promising potential in cancer therapy.
Assuntos
Antineoplásicos/farmacologia , Neoplasias/imunologia , alfa-Defensinas/biossíntese , Animais , Apoptose , Linhagem Celular Tumoral , Citocinas/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos SCID , Transplante de Neoplasias , Neoplasias/química , Baço/citologia , TransfecçãoRESUMO
Barbigerone is a naturally occurring isoflavone with antioxidant activity. In present study, we investigated the antitumor activity of barbigerone against murine lung cancer cells LL/2 and the possible mechanism in vitro. Our results showed that barbigerone inhibited LL/2 cells proliferation in a concentration- and time-dependent manner and caused apoptotic death of LL/2 cells. Barbigerone-induced apoptosis was characterized by enhanced mitochondrial cytochrome c release, activation of caspase-3,-9, but not caspase-8. Exposure of LL/2 cells to barbigerone resulted in upregulation of Bcl-2-associated protein (Bax) and down-regulation of Bcl-2. In addition, proliferation inhibitory effect of barbigerone was associated with decreased level of phosphorylated p42/44 mitogen-activated protein kinase (p42/44 MAPK) and phosphorylated Akt. Moreover, barbigerone exhibit less toxicity to non-cancer cells than tumor cells. In conclusion, our results indicated that barbigerone can inhibit murine lung cancer cell proliferation by inducing apoptosis via mitochondrial apoptotic pathway and by decreasing phosphorylated p42/44 MAPK and Akt. Its potential to be a candidate of anti-cancer agent is worth being further investigated.
Assuntos
Antineoplásicos/toxicidade , Apoptose , Isoflavonas/toxicidade , Neoplasias Pulmonares/patologia , Mitocôndrias/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica , Isoflavonas/química , Isoflavonas/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fatores de Tempo , Proteína X Associada a bcl-2/metabolismoRESUMO
PURPOSE: PNAS-4, a novel pro-apoptotic gene activated during the early response to DNA damage, can inhibit proliferation via apoptosis when overexpressed in some tumor cells. The objectives of this study were to determine whether PNAS-4 could enhance apoptosis induced by cisplatin besides its induction of apoptosis, and to evaluate the usefulness of combined treatment with mouse PNAS-4 (mPNAS-4) gene therapy and low-dose cisplatin chemotherapy in the inhibition of tumor growth in colon carcinoma (CT26) and Lewis lung carcinoma (LL/2) murine models. METHODS: In this study, the in vitro growth-inhibitory and pro-apoptotic effects of PNAS-4 and/or cisplatin on CT26, LL/2, and SKOV3 cancer cells were assessed by MTT assay, flow cytometric analysis, DNA fragmentation, and morphological analysis, respectively. The in vivo antitumor activity of combined treatment with mPNAS-4 gene therapy and low-dose cisplatin were evaluated in the inhibition of tumor growth in colon carcinoma (CT26) and Lewis lung carcinoma (LL/2) murine models. Tumor volume and survival time were observed. Induction of apoptosis was also assessed in tumor tissues. RESULTS: In vitro, PNAS-4 inhibited proliferation of colon carcinoma (CT26), Lewis lung carcinoma (LL/2) and human ovarian cancer (SKOV3) cell lines via apoptosis, and significantly enhanced the apoptosis of CT26, LL/2, and SKOV3 cells induced by cisplatin. In vivo systemic administration of expression plasmid encoding mPNAS-4 (pcDNA3.1-mPS) and cisplatin, significantly decreased tumor growth through increased tumor cell apoptosis compared to treatment with mPNAS-4 or cisplatin alone. CONCLUSIONS: Our data suggests that the combined treatment with mPNAS-4 plus cisplatin may augment the induction of apoptosis in tumor cells in vitro and in vivo, and that the augmented antitumor activity in vivo may result from the increased induction of apoptosis. The present study may provide a novel way to augment the antitumor efficacy of cytotoxic chemotherapy.