RESUMEN
A vertical external cavity surface emitting laser (VECSEL) has been developed for a sodium guide star application. Stable single frequency operation with 21 W of output power near 1178â nm with multiple gain elements while lasing in the TEM00 mode has been achieved. Higher output power results in multimode lasing. For the sodium guide star application, the 1178â nm can be frequency doubled to 589â nm. The power scaling approach used involves using multiple gain mirrors in a folded standing wave cavity. This is the first demonstration of a high power single frequency VECSEL using a twisted-mode configuration and multiple gain mirrors located at the cavity folds.
RESUMEN
Dacarbazine (DTIC) is one of the most popular alkylating agents used for the treatment of malignant melanoma. DTIC induces apoptosis of melanoma cells via double-strand breaks (DSBs). Melanoma cells, however, tend to increase their expression of DNA repair molecules in order to be resistant to DTIC. Here, we show that DTIC increases expression of Rad51, but not Ku70, in a cultured B16-F10 mouse melanoma cell line in dose- and time-dependent manners. On introducing Rad51 short interfering RNA (siRNA) with the hemagglutinating virus of Japan envelope (HVJ-E) to B16-F10 cells, DSBs induced by DTIC treatment were not efficiently repaired and resulted in enhanced apoptotic cell death. Colony formation of B16-F10 cells that received Rad51 siRNA was significantly decreased by DTIC treatment as compared with cells that received scramble siRNA. In melanoma-bearing mice, the combination of three intratumoral injections of HVJ-E containing Rad51 siRNA and five intraperitoneal injections of DTIC at a clinical dose synergistically suppressed the tumors. Moreover, HVJ-E demonstrated anti-tumor immunity by inducing cytotoxic T lymphocytes to B16-F10 cells on administration of DTIC. These results suggest that the combination of chemotherapy with HVJ-E containing therapeutic molecules will provide a promising therapeutic strategy for patients bearing malignant tumors resistant to chemotherapeutic agents.
Asunto(s)
Antineoplásicos Alquilantes/administración & dosificación , Dacarbazina/administración & dosificación , Melanoma Experimental/terapia , Recombinasa Rad51/genética , Neoplasias Cutáneas/terapia , Animales , Antígenos Nucleares/metabolismo , Roturas del ADN de Doble Cadena , Reparación del ADN/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Femenino , Autoantígeno Ku , Melanoma Experimental/tratamiento farmacológico , Melanoma Experimental/genética , Ratones , Ratones Endogámicos C57BL , ARN Interferente Pequeño/farmacología , Recombinasa Rad51/metabolismo , Virus Sendai/genética , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/genéticaRESUMEN
The transcriptional factor nuclear factor-kappaB (NFkappaB) plays a pivotal role in the coordinated transactivation of cytokine and adhesion molecule genes that might be involved in myocardial damage after ischemia and reperfusion. Therefore, we hypothesized that synthetic double-stranded DNA with high affinity for NFkappaB could be introduced in vivo as "decoy" cis elements to bind the transcriptional factor and to block the activation of genes mediating myocardial infarction, thus providing effective therapy for myocardial infarction. Treatment before and after infarction by transfection of NFkappaB decoy, but not scrambled decoy, oligodeoxynucleotides before coronary artery occlusion or immediately after reperfusion had a significant inhibitory effect on the area of infarction. Here, we report the first successful in vivo transfer of NFkappaB decoy oligodeoxynucleotides to reduce the extent of myocardial infarction following reperfusion, providing a new therapeutic strategy for myocardial infarction.
Asunto(s)
Infarto del Miocardio/prevención & control , FN-kappa B/metabolismo , Oligonucleótidos/administración & dosificación , Transfección , Animales , Sitios de Unión , Células Cultivadas , Fluoresceína-5-Isotiocianato , Regulación de la Expresión Génica/genética , Infarto del Miocardio/genética , Miocardio/metabolismo , Oligonucleótidos/genética , Ratas , Ratas Sprague-DawleyRESUMEN
Graft coronary arteriosclerosis, which limits the long-term survival of allograft recipients, is characterized by diffuse intimal thickening composed of proliferative smooth muscle cells. We observed that messenger RNA of the cell cycle regulatory enzyme cyclin-dependent kinase (cdk) 2 kinase, which mediates smooth muscle cell proliferation, was elevated in the thickened intima of coronary arteries of murine heterotopic cardiac allografts. We studied the effects of antisense phosphorothioate oligodeoxynucleotide (ODN) against this enzyme using gene transfer mediated by a hemagglutinating virus of Japan (HVJ)-liposome complex intraluminally delivered to inhibit the intimal hyperplasia. At 30 days after transplantation, antisense cdk2 kinase ODN treatment had dramatically inhibited neointimal formation in the allografts. Expression of vascular cell adhesion molecule-1 was also suppressed by antisense cdk2 kinase. However, these effects were not observed in the sense or scrambled ODN-treated allografts. Thus, an intraluminal administration of antisense ODN directed to a specific cell cycle regulatory gene can inhibit neointimal formation after cardiac transplantation.
Asunto(s)
Quinasas CDC2-CDC28 , Enfermedad de la Arteria Coronaria/prevención & control , Quinasas Ciclina-Dependientes/genética , Enfermedad Injerto contra Huésped/prevención & control , Trasplante de Corazón/efectos adversos , Oligonucleótidos Antisentido/uso terapéutico , Proteínas Serina-Treonina Quinasas/genética , Animales , Enfermedad de la Arteria Coronaria/etiología , Enfermedad de la Arteria Coronaria/patología , Vasos Coronarios/enzimología , Vasos Coronarios/patología , Quinasa 2 Dependiente de la Ciclina , Técnicas de Transferencia de Gen , Vectores Genéticos , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Oligonucleótidos Antisentido/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Respirovirus/genéticaRESUMEN
There are currently no effective therapies for progressive fibrotic diseases. Recent evidence has implicated overproduction of transforming growth factor-beta1 (TGF-beta1) as a major cause of tissue fibrosis. Furthermore, this evidence implies that inhibitors of TGF-beta1 may be clinically useful as antifibrotic agents. The proteoglycan decorin is a known inhibitor of TGF-beta1. In a rat model of glomerulonephritis we have shown that fibrosis is mediated by TGF-beta1. We report here that transfer of decorin cDNA into rat skeletal muscle increases the amount of decorin messenger RNA and protein present in skeletal muscle and decorin present in kidney, where it has a marked therapeutic effect on fibrosis induced by glomerulonephritis. Transfected glomerulonephritic rats showed a significant reduction in levels of glomerular TGF-beta1 mRNA and TGF-beta1 protein, extracellular matrix accumulation and proteinuria. These results demonstrate the potential of gene therapy as a novel treatment for fibrotic diseases caused by TGF-beta1.
Asunto(s)
Glomerulonefritis/prevención & control , Riñón/patología , Músculo Esquelético/metabolismo , Proteoglicanos/genética , Animales , Decorina , Proteínas de la Matriz Extracelular , Fibrosis/genética , Fibrosis/metabolismo , Fibrosis/prevención & control , Expresión Génica , Técnicas de Transferencia de Gen , Terapia Genética , Glomerulonefritis/genética , Glomerulonefritis/metabolismo , Proteoglicanos/biosíntesis , Proteoglicanos/uso terapéutico , Ratas , Ratas Sprague-DawleyRESUMEN
Closure of the ductus arteriosus requires prenatal formation of intimal cushions, which occlude the vessel lumen at birth. Survival of newborns with severe congenital heart defects, however, depends on ductal patency. We used a gene transfer approach to create a patent ductus arteriosus by targeting the fibronectin-dependent smooth muscle cell migration required for intimal cushion formation. Fetal lamb ductus arteriosus was transfected in utero with hemagglutinating virus of Japan liposomes containing plasmid encoding 'decoy' RNA to sequester the fibronectin mRNA binding protein. Fibronectin translation was inhibited and intimal cushion formation was prevented. We thus established the essential role of fibronectin-dependent smooth muscle cell migration in intimal cushion formation in the intact animal and the feasibility of incorporating biological engineering in the management of congenital heart disease.
Asunto(s)
Conducto Arterioso Permeable/genética , Fibronectinas/genética , Fibronectinas/fisiología , Terapia Genética/métodos , Transfección/métodos , Animales , Movimiento Celular/genética , Modelos Animales de Enfermedad , Conducto Arterioso Permeable/embriología , Conducto Arterioso Permeable/cirugía , Femenino , Vectores Genéticos , Cardiopatías Congénitas/mortalidad , Cardiopatías Congénitas/patología , Cardiopatías Congénitas/terapia , Liposomas , Músculo Liso Vascular/citología , Plásmidos , Embarazo , Biosíntesis de Proteínas , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Respirovirus , OvinosRESUMEN
Liver cirrhosis is the irreversible end result of fibrous scarring and hepatocellular regeneration, characterized by diffuse disorganization of the normal hepatic structure of regenerative nodules and fibrotic tissue. It is associated with prominent morbidity and mortality, and is induced by many factors, including chronic hepatitis virus infections, alcohol drinking and drug abuse. Hepatocyte growth factor (HGF), originally identified and cloned as a potent mitogen for hepatocytes, shows mitogenic, motogenic and morphogenic activities for a wide variety of cells. Moreover, HGF plays an essential part in the development and regeneration of the liver, and shows anti-apoptotic activity in hepatocytes. In a rat model of lethal liver cirrhosis produced by dimethylnitrosamine administrations, repeated transfections of the human HGF gene into skeletal muscles induced a high plasma level of human as well as enodogenous rat HGF, and tyrosine phosphorylation of the c-Met/HGF receptor. Transduction with the HGF gene also suppressed the increase of transforming growth factor-beta1 (TGF-beta1), which plays an essential part in the progression of liver cirrhosis, inhibited fibrogenesis and hepatocyte apoptosis, and produced the complete resolution of fibrosis in the cirrhotic liver, thereby improving the survival rate of rats with this severe illness. Thus, HGF gene therapy may be potentially useful for the treatment of patients with liver cirrhosis, which is otherwise fatal and untreatable by conventional therapy.
Asunto(s)
Terapia Genética , Factor de Crecimiento de Hepatocito/genética , Cirrosis Hepática Experimental/terapia , Animales , Apoptosis , Northern Blotting , Humanos , Hígado/patología , Cirrosis Hepática Experimental/patología , Ratas , Ratas Sprague-Dawley , Transfección , Factor de Crecimiento Transformador beta/biosíntesisRESUMEN
SUMMARY: The main component of the metabolic by-products of fermentation by Propionibacterium freudenreichii ET-3 is 1,4-dihydroxy-2-naphthoic acid (DHNA), which has a naphthoquinone skeleton, as in vitamin K2. This study showed that DHNA improved bone mass reduction with osteoporosis model mice caused by FK506. INTRODUCTION: Growth of the intestinal bacterium Lactobacillus bifidus is specifically facilitated by DHNA. The present study used osteoporosis model mice to investigate the effects of DHNA on bone remodeling. METHODS: FK506, an immunosuppressant, was used to prepare osteoporosis model mice. Thirty mice were divided into three groups: FK group, FK+DHNA group, and control group. In the FK group, FK506 was administered to induce bone mass reduction. In the FK-DHNA group, FK506 and DHNA were administered concurrently to observe improvements in bone mass reduction. To ascertain systemic and local effects of DHNA, we investigated systemic pathological changes in colon, kidney function and cytokine dynamics, and morphological and organic changes in bone and osteoclast dynamics as assessed by culture experiments. RESULTS: Compared to the FK group without DHNA, colon damage and kidney dysfunction were milder for FK+DHNA group, and production of inflammatory cytokines (interleukin (IL)-1beta, IL-6 and tumor necrosis factor (TNF)-alpha) was more suppressed. Furthermore, compared to the group without DHNA, histological analyses and radiography showed that bone resorption was suppressed for the DHNA group. Culture experiments using osteoclasts from murine bone marrow showed osteoclast suppression for the DHNA group compared to the group without DHNA. CONCLUSION: These results show that DHNA has some effects for improving bone mass reduction caused by FK506.
Asunto(s)
Conservadores de la Densidad Ósea/uso terapéutico , Naftoles/uso terapéutico , Osteoporosis/prevención & control , Animales , Peso Corporal/efectos de los fármacos , Densidad Ósea/efectos de los fármacos , Células Cultivadas , Colon/efectos de los fármacos , Colon/patología , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos/métodos , Ingestión de Alimentos/efectos de los fármacos , Fémur/efectos de los fármacos , Fémur/patología , Fémur/fisiopatología , Inmunosupresores , Mediadores de Inflamación/metabolismo , Riñón/efectos de los fármacos , Riñón/fisiopatología , Masculino , Ratones , Ratones Endogámicos ICR , Osteoclastos/efectos de los fármacos , Osteoclastos/patología , Osteoporosis/inducido químicamente , TacrolimusRESUMEN
OBJECTIVE: The aim of this study is to understand whether the responsiveness of the hypothalamic-pituitary-adrenal (HPA) axis to stress increases excessively with aging in senescence-accelerated mice-prone 10 (SAMP10) and to investigate the role of arachidonic acid (ARA) in this process. MATERIALS AND METHODS: The area under the curve of CORT concentration (CORT-AUC), an index of the HPA axis responsiveness to stress, was assessed in SAMP10 subjected to a 30-minute restraint stress up to 120 minutes after the restraint stress onset. Furthermore, the HPA axis responsiveness was evaluated in aged SAMP10 fed 0.4% ARA-containing diet (ARA group) or control diet (CON group) for 4 weeks. Three weeks later, these mice were divided into a group with a 30-minute restraint stress (CON-S or ARA-S group) and a group without restraint stress (CON-NS or ARA-NS group). Hippocampi were collected after stress release and fatty acid and glucocorticoid receptor (GR) protein levels were evaluated in the nucleus and cytosol. RESULTS: The CORT-AUC of aged SAMP10 was 21% significantly higher than that of young SAMP10. In the ARA group, hippocampal ARA was 0.5% significantly higher than that in the CON group. CORT-AUC in the ARA group was 24% significantly lower than that in the CON group. The ratio of GR protein levels in the nucleus and cytosol in the ARA-S group was 1.72 times significantly higher than that in the ARA-NS group but no difference was observed between the CON-S and CON-NS groups. CONCLUSIONS: Dietary ARA seems to suppress age-related excessive enhancement of the HPA axis responsiveness via attenuation of age-related decline in hippocampal GR translocation into the nucleus after stress loading, which may contribute to an improvement of mental health.
Asunto(s)
Envejecimiento/efectos de los fármacos , Ácido Araquidónico/farmacología , Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Sistema Hipófiso-Suprarrenal/efectos de los fármacos , Estrés Fisiológico/efectos de los fármacos , Envejecimiento/metabolismo , Animales , Ácido Araquidónico/administración & dosificación , Suplementos Dietéticos , Sistema Hipotálamo-Hipofisario/metabolismo , Ratones , Sistema Hipófiso-Suprarrenal/metabolismoRESUMEN
Hemagglutinating virus of Japan envelope (HVJ-E) vector with inactivated replication-defective Sendai virus was originally developed as a versatile drug delivery system. Recently, we have shown the direct tumor-killing activity of HVJ-E itself without therapeutic molecules in prostate cancer cells. Although human glioblastoma cells were also sensitive to HVJ-E treatment, complete eradication was not achieved using HVJ-E alone. Here, to develop more effective therapeutic strategy of glioblastoma, we enhanced the anti-tumor activity by incorporating Short interfering RNA (siRNA) of mitotic motor protein Eg5 into HVJ-E. Treatment with HVJ-E-containing Eg5 siRNA induced monopolar spindle formation and resulted in cell-cycle arrest and apoptosis. When HVJ-E-containing Eg5 siRNA was directly injected into an intradermal tumor xenograft, all mice became tumor-free. Similar results were observed in the intracranial tumor xenografts. The survival time of treated mice was significantly prolonged when HVJ-E was used. Histological examination showed that tumors remained in the brain after treatment with HVJ-E-containing negative control siRNA, but no tumors were detected after treatment with HVJ-E-containing Eg5 siRNA. This remarkable anti-tumor response was likely due to a synergistic effect of Eg5 siRNA and HVJ-E. Thus, this combination shows promise as an attractive novel therapy for glioblastoma.
Asunto(s)
Terapia Genética/métodos , Glioblastoma/terapia , Cinesinas/genética , Viroterapia Oncolítica/métodos , ARN Interferente Pequeño , Virus Sendai/genética , Proteínas del Envoltorio Viral/genética , Animales , Apoptosis , Ciclo Celular , Femenino , Técnicas de Transferencia de Gen , Vectores Genéticos , Masculino , Ratones , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Ehrlich ascites tumor cells were found to be very insensitive to diphtheria toxin. We formed 37 hybrids from Ehrlich tumor cells and diphtheria toxin-sensitive human fibroblasts. The effects of diphtheria toxin on protein synthesis in those hybrids were examined. The hybrids were divided into three groups on the basis of toxin sensitivity. Group A hybrids were as sensitive to diphtheria toxin as human fibroblasts, Group C were as resistant as Ehrlich tumor cells, and Group B had intermediate sensitivity. Group A hybrids had diphtheria toxin-binding sites but Group B and C had no detectable binding sites. Elongation factor-2 of all the hybrids was susceptible to ADP-ribosylation by fragment A of diphtheria toxin. Cells of Group A and B became more sensitive to CRM 45 (cross-reacting material 45 of diphtheria toxin) after they were exposed to low pH (pH = 4.5). The resistance of Group C to CRM 45 was not affected by the same treatment. Group A and B hybrids and human fibroblasts had similar sensitivities to a hybrid toxin composed of wheat germ agglutinin and fragment A of diphtheria toxin, but Group C and Ehrlich tumor cells were resistant to this hybrid toxin. All the hybrids and Ehrlich tumor cells were more sensitive to a hybrid toxin composed of wheat germ agglutinin and subunit A of ricin than were human fibroblasts. On subcloning of Group B hybrids, one Group C hybrid was obtained, but no Group A hybrid. These facts suggest that Ehrlich ascites tumor cells differ from human fibroblasts in the expression of a factor(s) that is involved in entry of fragment A of diphtheria toxin into the cytoplasm after the toxin binds to its surface receptors.
Asunto(s)
Carcinoma de Ehrlich/metabolismo , Toxina Diftérica/metabolismo , Células Híbridas/metabolismo , Receptores de Superficie Celular , Receptores Colinérgicos/metabolismo , Adenosina Difosfato Ribosa/metabolismo , Animales , Membrana Celular/metabolismo , Toxina Diftérica/toxicidad , Fibroblastos/metabolismo , Factor de Crecimiento Similar a EGF de Unión a Heparina , Humanos , Péptidos y Proteínas de Señalización Intercelular , Cinética , Masculino , Ratones , Factor 2 de Elongación Peptídica , Factores de Elongación de Péptidos/metabolismo , Biosíntesis de Proteínas/efectos de los fármacos , Piel/metabolismoRESUMEN
A monoclonal antibody that recognizes antigenic determinants on the nucleus of cultured mammalian cells was isolated. Immunofluorescence studies using this antibody showed that the recognized antigen was present not only on the nucleus but also in cytoplasmic vesicles of interphase cells and in the perichromosomal region of mitotic cells. Premature chromosome condensation analysis showed that the reactive site for this monoclonal antibody could be detected in the perichromosomal region during the G2 and M phases, but not during the G1 and S phases. Finally, immunoblot analysis showed that this monoclonal antibody prepared against the nucleus recognized a protein of approximately 40 kD both in the cytoplasm and in the perichromosomal regions.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Núcleo Celular/inmunología , Gránulos Citoplasmáticos/inmunología , Animales , Especificidad de Anticuerpos , Núcleo Celular/ultraestructura , Cromosomas/inmunología , Técnica del Anticuerpo Fluorescente , Humanos , Inmunoglobulina M/inmunología , Técnicas de Inmunoadsorción , Interfase , Membranas Intracelulares/inmunología , Mitosis , Peso Molecular , Membrana Nuclear/inmunología , Proteínas/inmunologíaRESUMEN
DNA and nuclear proteins were transferred into cells simultaneously at more than 95% efficiency by means of vesicle complexes. The DNA was rapidly transported into the nuclei of cultured cells, and its expression reached a maximum within 6 to 8 hours after its introduction. Moreover, when the plasmid DNA and nuclear protein were cointroduced into nondividing cells in rat liver by injection into the portal veins of adult rats, the plasmid DNA was carried into liver cell nuclei efficiently by nuclear protein. The expression of the DNA in adult rat liver, on introduction of the DNA with nuclear protein, was more than five times as great as with nonnuclear protein.
Asunto(s)
ADN/metabolismo , Proteínas del Grupo de Alta Movilidad/metabolismo , Hígado/metabolismo , Animales , Northern Blotting , Compartimento Celular , Núcleo Celular/metabolismo , Células Cultivadas , ADN/farmacocinética , Ratones , Ratas , Transformación GenéticaAsunto(s)
Terremotos , Salud Mental , Humanos , Anciano , Trastornos por Estrés Postraumático/epidemiología , Desastres , MasculinoRESUMEN
For the first time, we demonstrate injection locking and single frequency operation of a multi-core Yb-doped phosphate fiber laser (MCF). The 19 MCF laser cores operated in CW mode at 1030 nm. Each laser core was locked to the frequency and polarization of the single-frequency master laser, and produced milliwatts of power with similar lasing thresholds. The pump beam was homogenized with a simple technique to increase uniform lasing behavior of the cores. This behavior was verified using a MCF laser model developed in-house. This unique MCF laser can be useful for applications of coherent, coupled oscillator networks, for example in an all-optical coherent Ising machine configuration.
RESUMEN
OBJECTIVES AND DESIGN: The expressions of human beta defensin-1 (HBD-1), -2 (HBD-2) and -3 (HBD-3) in non-inflamed pseudocysts such as mucoceles were investigated immunohistochemically in this study. MATERIALS AND METHODS: Mucocele specimens were obtained from 21 patients. The expression of HBDs was studied immunohistochemically by using antibodies directed against HBD-1, -2, and -3. Statistical analyses were carried out on serial sections stained with antibodies. RESULTS: Cells expressing HBDs were found in mucoceles. The expression of HBD-2 was observed in floating cells in all the specimens, whereas HBD-1 and HBD-3-expressing cells were detected in 93% and 73% of the mucoceles, respectively. The HBD-2 signal was the most intense and the HBD-3 signal intensity was weaker than that of HBD-1. HBDs were expressed in neutrophils and in other floating cells. Interestingly, the signal intensity and the population of positive cells located close to the centers of cysts were higher than those located in the peripheral areas of cysts. CONCLUSION: The expression of HBDs was found even in non-inflamed pseudocysts such as mucoceles. These results suggest that an unknown mechanism not involved in biophylaxis for the expression of HBDs may exist.
Asunto(s)
Enfermedades de los Labios/metabolismo , Mucocele/metabolismo , beta-Defensinas/biosíntesis , Adolescente , Adulto , Niño , Preescolar , Femenino , Expresión Génica , Humanos , Inmunohistoquímica , Masculino , Adulto JovenAsunto(s)
Infecciones por Papillomavirus , Vacunas contra Papillomavirus , Neoplasias del Cuello Uterino , Humanos , Femenino , Japón , Confianza , Infecciones por Papillomavirus/prevención & control , Estigma Social , Vacunación , Conocimientos, Actitudes y Práctica en Salud , Neoplasias del Cuello Uterino/prevención & controlRESUMEN
Vascular renin angiotensin system (RAS) has been reported to exist in vascular wall. However, there is no direct evidence whether the vascular RAS per se can modulate growth of vascular smooth muscle cells (VSMC), because there is no suitable method to investigate the effect of endogenously produced vasoactive substances on growth of these cells. In this study, we transferred angiotensin-converting enzyme (ACE) and/or renin cDNAs into cultured VSMC using the efficient Sendai virus (hemagglutinating virus of Japan) liposome-mediated gene transfer method, to examine their relative roles in VSMC growth in vitro. Within 35 min or 6 h, the transfection of ACE cDNA into VSMC by hemagglutinating virus of Japan method resulted in a twofold higher ACE activity than control vector, whereas a cationic liposome (Lipofectin)-mediated method failed to show any effect. This in vitro system provided us with the opportunity to investigate the influence of endogenous vascular RAS on VSMC growth. Transfection of ACE or renin cDNA resulted in increased DNA and RNA synthesis, which was inhibited with the specific angiotensin II receptor antagonist (DuP 753: 10(-6) M). Angiotensin I added to ACE-transfected VSMC increased RNA synthesis in a dose-dependent manner. Cotransfection of renin and ACE cDNAs stimulated further RNA synthesis as compared to ACE or renin cDNA alone. These results showed that transfected components of RAS can modulate VSMC growth through the endogenous production of vascular angiotensin II, and that ACE as well as renin are rate limiting in determining the VSMC RAS activity. We conclude that the hemagglutinating virus of Japan liposome-mediated gene transfer technique provides a new and useful tool for study of endogenous vascular modulators such as vascular RAS.
Asunto(s)
Desarrollo de Músculos , Músculo Liso Vascular/crecimiento & desarrollo , Peptidil-Dipeptidasa A/genética , Renina/genética , Transfección/métodos , Animales , Aorta/citología , Células Cultivadas , Liposomas , Músculo Liso Vascular/citología , Virus de la Parainfluenza 1 Humana/genética , RatasRESUMEN
Glomerulosclerosis, a final common lesion of various glomerular diseases, is characterized by mesangial cell proliferation and extracellular matrix (ECM) expansion. TGF-beta and PDGF are known to play a critical role in the regulation of ECM metabolism and mesenchymal cell proliferation, respectively. However, there is little evidence to demonstrate the direct role of each of these growth factors in the pathogenesis of glomerulosclerosis. Using an in vivo transfection technique, we could realize the selective overexpression of single growth factor in the kidney. The introduction of either TGF-beta or PDGF-B gene alone into the kidney induced glomerulosclerosis, although the patterns of action of these growth factors were different; TGF-beta affected ECM accumulation rather than cell proliferation and PDGF affected the latter rather than the former.
Asunto(s)
Glomeruloesclerosis Focal y Segmentaria/metabolismo , Glomeruloesclerosis Focal y Segmentaria/patología , Glomérulos Renales/patología , Riñón/metabolismo , Factor de Crecimiento Derivado de Plaquetas/biosíntesis , Transfección , Factor de Crecimiento Transformador beta/biosíntesis , Animales , Colágeno/análisis , Colágeno/metabolismo , ADN/administración & dosificación , ADN/metabolismo , Portadores de Fármacos , Técnica del Anticuerpo Fluorescente , Glomeruloesclerosis Focal y Segmentaria/etiología , Humanos , Glomérulos Renales/metabolismo , Liposomas , Plásmidos , Factor de Crecimiento Derivado de Plaquetas/genética , Ratas , Ratas Sprague-Dawley , Factores de Tiempo , Factor de Crecimiento Transformador beta/genéticaRESUMEN
Heat shock protein 70 (HSP70) has been reported to be involved in the myocardial self-preservation system. To obtain the evidence that HSP70 plays a direct role in the protection from myocardial ischemia-reperfusion injury, rat hearts were transfected with human HSP70 gene by intracoronary infusion of hemagglutinating virus of Japan (HVJ)-liposome containing human HSP70 gene. The control hearts were infused with HVJ-liposome without the HSP70 gene. The hearts from whole-body heat-stressed or nontreated rats were also examined. Western blot and immunohistochemical analysis showed that apparent overexpression of HSP70 occurred in the gene transfected hearts and that gene transfection might be more effective for HSP70 induction than heat stress. In Langendorff perfusion, better functional recovery as well as less creatine phosphokinase leakage after ischemia were obtained in the gene transfected hearts with HSP70 than in the control or nontreated hearts. Furthermore, the gene transfected hearts showed better functional recovery than the heat-stressed hearts. These results indicated that overexpressed HSP70 plays a protective role in myocardial injury, suggesting the possibility that gene transfection with HSP70 may become a novel method for myocardial protection through enforcing the self-preservation systems.