RESUMO
Positron emission tomography (PET)/computed tomography (CT) using sodium [18F]fluoride (Na[18F]F) has been proven to be a promising hot-spot imaging modality for myocardial infarction (MI). We investigated Na[18F]F uptake in ischemia-reperfusion injury (IRI) of rats and humans. Sodium [18F]fluoride PET/CT was performed in Sprague-Dawley rats that had IRI surgery, and it readily demonstrated prominent Na[18F]F uptake in the infarct area post-IRI. Sodium [18F]fluoride uptake was matched with negative 2,3,5-triphenyl-2 H-tetrazolium chloride staining results, accompanied by myocardial apoptosis and associated with positive calcium staining results. Furthermore, area at risk was negative for Na[18F]F uptake. Cyclosporine A (CysA) treatment reduced standardized uptake value of 18F over the infarct area, and a significant decrease in infarct size was also observed by the CysA treatment. In humans, Na[18F]F PET/CT readily demonstrated increased Na[18F]F uptake in the 2 patients with MI post-percutaneous coronary intervention. In conclusion, this study sheds light on the potential utility of Na[18F]F PET/CT as a hot-spot imaging modality for myocardial IRI.
Assuntos
Fluordesoxiglucose F18/análise , Imagem Multimodal/métodos , Infarto do Miocárdio/diagnóstico por imagem , Traumatismo por Reperfusão Miocárdica/diagnóstico por imagem , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , Animais , Humanos , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
A proof-of-concept study is presented using dual gene therapy that employed a small hairpin RNA (shRNA) specific for mammalian target of rapamycin (mTOR) and a herpes simplex virus-thymidine kinase (HSV-TK) gene to inhibit the growth of tumors. Recombinant adeno-associated virus (rAAV) vectors containing a mutant TK gene (sc39TK) were transduced into HeLa cells, and the prodrug ganciclovir (GCV) was administered to establish a suicide gene-therapy strategy. Additionally, rAAV vectors expressing an mTOR-targeted shRNA were employed to suppress mTOR-dependent tumor growth. GCV selectively induced death in tumor cells expressing TK, and the mTOR-targeted shRNA altered the cell cycle to impair tumor growth. Combining the TK-GCV system with mTOR inhibition suppressed tumor growth to a greater extent than that achieved with either treatment alone. Furthermore, HSV-TK expression and mTOR inhibition did not mutually interfere with each other. In conclusion, gene therapy that combines the TK-GCV system and mTOR inhibition shows promise as a novel strategy for cancer therapy.
Assuntos
Terapia Genética/métodos , Neoplasias Experimentais/terapia , Simplexvirus/genética , Serina-Treonina Quinases TOR/genética , Timidina Quinase/genética , Proteínas Virais/genética , Animais , Antineoplásicos/farmacologia , Antivirais/farmacologia , Western Blotting , Dependovirus , Ganciclovir/farmacologia , Genes Transgênicos Suicidas , Vetores Genéticos , Células HeLa , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , RNA Interferente Pequeno , Transdução Genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Sodium [(18)F]fluoride (Na[(18)F]F) positron emission tomography with integrated computed tomography (PET/CT) has not been used for imaging myocardial infarction (MI). Here, we aimed to investigate the Na[(18)F]F PET/CT features of MI in a rat model. PROCEDURES: MI was induced by coronary artery ligation in 8-week-old male Spraque-Dawley rats (300 ± 10 g) and confirmed by triphenyl tetrazolium chloride (TTC) staining. Na[(18)F]F PET/CT images were obtained using an animal-dedicated PET/CT scanner (NanoPET/CT, Mediso) in vivo and ex vivo. Uptake of Na[(18)F]F was quantitated using the standardized uptake value (SUV). Myocardial apoptosis was evaluated using histone-1 targeted peptide (ApoPep-1) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining, while calcium accumulation was investigated using von Kossa's staining. Na[(18)F]F PET/CT was compared with (99m)Tc-methoxyisobutylisonitrile (MIBI) or (99m)Tc-hydroxymethylenediphosphonate (HMDP) single photon emission computed tomography/computed tomography (SPECT/CT) in rats with day 1 MI. RESULTS: The rats showed strong Na[(18)F]F uptake both in vivo and ex vivo; the maximal uptake occurred 1 day after MI (SUV ratio of infarct to lung = 4.56 ± 0.74, n = 7, P = 0.0183 vs the control). The Na[(18)F]F uptake area perfectly matched the apoptotic area, determined by ApoPep-1 uptake and TUNEL assay. However, calcification, assessed by von Kossa's staining, was absent in the infarct. Na[(18)F]F PET/CT showed an increased uptake at the perfusion deficit area in [(99m)Tc]MIBI SPECT/CT and an equivalent signal to [(99m)Tc]HMDP SPECT/CT in rats with day 1 MI. CONCLUSIONS: Na[(18)F]F PET/CT is a promising hot-spot imaging modality for MI.
Assuntos
Infarto do Miocárdio/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Fluoreto de Sódio , Tomografia Computadorizada por Raios X , Animais , Apoptose , Cálcio/metabolismo , Vasos Coronários/diagnóstico por imagem , Radioisótopos de Flúor , Histonas/metabolismo , Ligadura , Masculino , Ratos Sprague-Dawley , Medronato de Tecnécio Tc 99m/análogos & derivados , Tecnécio Tc 99m Sestamibi , Tomografia Computadorizada de Emissão de Fóton ÚnicoRESUMO
We have previously reported that interleukin-1 (IL-1) receptor-associated kinase (IRAK1) is essential for Epstein-Barr virus (EBV) latent infection membrane protein 1 (LMP1)-induced p65/RelA serine 536 phosphorylation and NF-κB activation but not for IκB kinase α (IKKα) or IKKß activation (Y. J. Song, K. Y. Jen, V. Soni, E. Kieff, and E. Cahir-McFarland, Proc. Natl. Acad. Sci. U. S. A. 103:2689-2694, 2006, doi:10.1073/pnas.0511096103). Since the kinase activity of IRAK1 is not required for LMP1-induced NF-κB activation, IRAK1 is proposed to function as a scaffold protein to recruit a p65/RelA serine 536 kinase(s) to enhance NF-κB-dependent transcriptional activity. We now report that Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) interacts with IRAK1 and is critical for LMP1-induced p65/RelA serine 536 phosphorylation and NF-κB activation. CaMKII bound the death domain of IRAK1 and directly phosphorylated p65/RelA at serine 536 in vitro. Downregulation of CaMKII activity or expression significantly reduced LMP1-induced p65/RelA serine 536 phosphorylation and NF-κB activation. Furthermore, LMP1-induced CaMKII activation and p65/RelA serine 536 phosphorylation were significantly reduced in IRAK1 knockout (KO) mouse embryonic fibroblasts (MEFs). Thus, IRAK1 may recruit and activate CaMKII, which phosphorylates p65/RelA serine 536 to enhance the transactivation potential of NF-κB in LMP1-induced NF-κB activation pathway.
Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Quinases Associadas a Receptores de Interleucina-1/metabolismo , NF-kappa B/metabolismo , Fator de Transcrição RelA/metabolismo , Proteínas da Matriz Viral/metabolismo , Animais , Western Blotting , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Células Cultivadas , Embrião de Mamíferos/citologia , Ativação Enzimática , Fibroblastos/citologia , Fibroblastos/metabolismo , Células HEK293 , Humanos , Quinases Associadas a Receptores de Interleucina-1/genética , Camundongos , Camundongos Knockout , Mutação , NF-kappa B/genética , Fosforilação , Ligação Proteica , Interferência de RNA , Serina/genética , Serina/metabolismo , Fator de Transcrição RelA/genética , Proteínas da Matriz Viral/genéticaRESUMO
We have previously reported that seventy percent ethanol extract of Chrysanthemum indicum Linne (CIE) strongly reduces Epstein-Barr virus (EBV)-transformed lymphoblastoid cell line (LCL) survival by inhibiting virus-encoded latent infection membrane protein 1 (LMP1)-induced NF-κB activation. To identify an active compound(s) in CIE that inhibits LMP1-induced NF-κB activation, activity-guided fractionation was employed. The CH2Cl2 fraction of CIE strongly reduced LMP1-induced NF-κB activation and LCL viability with relatively low cytotoxic effects on primary human foreskin fibroblast (HFF), HeLa or Burkitt's lymphoma (BL41) cells. Furthermore, lupeol, a pentacyclic triterpene, was identified in the CH2Cl2 fraction of CIE to attenuate LMP1-induced NF-κB activation and LCL viability. This study demonstrates that lupeol is one of active compounds in the CH2Cl2 fraction of CIE that inhibits LMP1-induced NF-κB activation and reduces NF-κB-dependent LCL viability.