RESUMEN

Among young women, the incidence of uterine corpus cancer is increasing. Most young women can not preserve fertility because simple total hysterectomy with bilateral salpingo-oophorectomy is the standard method for early endometrial cancer so far. We present a case of early endometrial adenocarcinoma which succeeded in pregnancy and delivery after resectoscopic surgery. Following a circumferential resection of the lesion including the mucosa and muscle layer under resectoscopic guidance, the patient became pregnant by means of in vitro fertilization-embyo transfer with hormone replenishment. She underwent cesarean section at 33 weeks and five days of gestation and had a healthy baby. Resectscopic surgery can help to preserve fertility among young women who have early invasive endometrial cancer.

Asunto(s)

Adenocarcinoma/cirugía , Neoplasias Endometriales/cirugía , Histeroscopía/métodos , Adulto , Cesárea , Transferencia de Embrión , Femenino , Fertilización In Vitro , Humanos , Embarazo

RESUMEN

Various types of oculocutaneous albinism (OCA) are associated with reduced pigmentation in the skin, hair, and eyes that results from mutations in genes involved in melanin synthesis. Immortal mouse melanocyte lines (melan-a, melan-b, and melan-c) provide opportune models with which to investigate the etiology of two different types of OCA (types I and III), which arise from mutations in Tyr and Tyrp1, respectively. We compared intracellular processing, sorting, and degradation of tyrosinase and Tyrp1, and the effects on their catalytic function and melanin synthesis, in these wild-type and mutant melanocytes. A mutation in either Tyr or Tyrp1 increased the time of association of tyrosinase and Tyrp1 with calnexin and Bip, which in turn resulted in the retention of these mutant products in the ER. A mutation in either gene selectively enhanced the duration and efficiency of chaperone interactions (even with the wild-type protein in the mutant melanocytes) and markedly slowed their transport to melanosomes. These results show that OCA1 and OCA3 are (in some cases, at least) ER retention diseases wherein a mutation in one melanogenic protein affects the maturation and stability of the other in the melanogenic pathway.

Asunto(s)

Albinismo Oculocutáneo/etiología , Retículo Endoplásmico/fisiología , Proteínas de Choque Térmico , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Monofenol Monooxigenasa/genética , Monofenol Monooxigenasa/metabolismo , Oxidorreductasas , Albinismo Oculocutáneo/enzimología , Animales , Proteínas de Unión al Calcio/metabolismo , Calnexina , Proteínas Portadoras/metabolismo , Chaperón BiP del Retículo Endoplásmico , Hexosaminidasas/química , Oxidorreductasas Intramoleculares/metabolismo , Sustancias Macromoleculares , Melaninas/análisis , Melanocitos/enzimología , Melanocitos/metabolismo , Ratones , Chaperonas Moleculares/metabolismo , Mutación , Células Tumorales Cultivadas

RESUMEN

In response to agouti signal protein, melanocytes switch from producing eumelanin to pheomelanin concomitant with the down-regulation of melanogenic gene transcription. We previously reported that a ubiquitous basic helix-loop-helix transcription factor, known as ITF2, is up-regulated during this switch, and we now report that treatment of melanocytes with melanocyte-stimulating hormone down-regulates expression of ITF2. To more fully characterize the involvement of ITF2 in regulating melanogenic gene transcription, ITF2 sense or antisense constructs were introduced into melan-a melanocytes. Gene and protein expression analyses and luciferase reporter assays using promoters from melanogenic genes showed that up-regulation of ITF2 suppressed melanogenic gene expression as well as the expression of Mitf, a melanocyte-specific transcription factor. In addition, stable ITF2 sense transfectants had significant reductions in pigmentation and a less dendritic phenotype compared with mock transfectants. In contrast, ITF2 antisense-transfected melanocytes were more pigmented and more dendritic. These results demonstrate that up-regulation of ITF2 during the pheomelanin switch is functionally significant and reveal that differential expression of a ubiquitous basic helix-loop-helix transcription factor can modulate expression of melanogenic genes and the differentiation of melanocytes.

Asunto(s)

Proteínas de Unión al ADN/fisiología , Melanocitos/metabolismo , Proteínas del Tejido Nervioso , Transactivadores/fisiología , Transcripción Genética , Animales , Antígenos de Neoplasias , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice , Northern Blotting , Diferenciación Celular , AMP Cíclico/metabolismo , Proteínas de Unión al ADN/química , Células Dendríticas/metabolismo , Regulación hacia Abajo , Genes Reporteros , Secuencias Hélice-Asa-Hélice , Luciferasas/metabolismo , Antígeno MART-1 , Melaninas/metabolismo , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Microscopía Electrónica , Modelos Biológicos , Proteínas de Neoplasias/metabolismo , Oligonucleótidos Antisentido/metabolismo , Fenotipo , Plásmidos/metabolismo , Pruebas de Precipitina , Regiones Promotoras Genéticas , Isoformas de Proteínas , ARN Mensajero/metabolismo , Ribonucleasas/metabolismo , Factores de Transcripción TCF , Transactivadores/química , Factor de Transcripción 4 , Factores de Transcripción/metabolismo , Transfección , Regulación hacia Arriba

RESUMEN

Oculocutaneous albinism type 1 (OCA1) is an autosomal recessive disease resulting from mutations of the tyrosinase gene (TYR). To elucidate the molecular basis of OCA1 phenotypes, we analysed the early processing and maturation of several different types of mutant tyrosinase with various degrees of structural abnormalities (i.e. two large deletion mutants, two missense mutants that completely destroy catalytic function and three missense mutants that have a temperature-sensitive phenotype). When expressed in COS7 cells, all mutant tyrosinases were sensitive to endoglycosidase H digestion, and immunostaining showed their localization in the endoplasmic reticulum (ER) and their failure to be sorted further to their target organelles. Pulse-chase experiments showed that all mutant tyrosinases were retained by calnexin in the ER and that they were degraded at similarly rapid rates, which coincided with their dissociation from calnexin. Temperature-sensitive mutant enzymes were sorted more efficiently at 31 degrees C than at 37 degrees C, and their degradation was accelerated at 37 degrees C compared with 31 degrees C. Thus in contrast to the current concept that mutant tyrosinases are transported to melanosomes but are functionally inactive there, our results suggest that mutant tyrosinases may not be transported to melanosomes in the first place. We conclude that a significant component of mutant tyrosinase malfunction in OCA1 results from their retention and degradation in the ER compartment. This quality-control process is highly sensitive to minimal changes in protein folding, and so even relatively minor mutations in peripheral sequences of the enzyme not involved with catalytic activity may result in a significant reduction of functional enzyme in melanosomes.

Asunto(s)

Albinismo Oculocutáneo/enzimología , Monofenol Monooxigenasa/metabolismo , Pigmentación/genética , Albinismo Oculocutáneo/genética , Animales , Células COS , Proteínas de Unión al Calcio/metabolismo , Calnexina , Cisteína Endopeptidasas/metabolismo , Inhibidores de Cisteína Proteinasa/farmacología , Retículo Endoplásmico/metabolismo , Humanos , Hidrólisis , Inmunohistoquímica , Monofenol Monooxigenasa/genética , Complejos Multienzimáticos/metabolismo , Complejo de la Endopetidasa Proteasomal , Transporte de Proteínas , Fracciones Subcelulares/enzimología , Temperatura

RESUMEN

This study deals with the cloning and characterization of monosaccharide transporter cDNAs in rice. OsMST1-3 (Oryza sativa monosaccharide transporters 1-3) have two sets of putative six transmembrane domains separated by a central long hydrophilic region. Heterologous expression of OsMST3 in the yeast Saccharomyces cerevisiae indicated that OsMST3 has transport activity for some monosaccharides in an energy-dependent H+ co-transport manner. Northern blot and in situ hybridization analyses showed that OsMST3 mRNA is detectable in leaf blades, leaf sheaths, calli and roots, especially the xylem as well as in sclerenchyma cells in the root. These results suggested that OsMST3 is involved in the accumulation of monosaccharides required for cell wall synthesis at the stage of cell thickening.

Asunto(s)

Hexosas/metabolismo , Proteínas de la Membrana/genética , Proteínas de Transporte de Monosacáridos/genética , Oryza/genética , Secuencia de Aminoácidos , Transporte Biológico , Clonación Molecular , Metabolismo Energético , Modelos Moleculares , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Distribución Tisular

RESUMEN

Assembly, target-signaling and transport of tyrosinase gene family proteins at the initial stage of melanosome biogenesis are reviewed based on our own discoveries. Melanosome biogenesis involves four stages of maturation with distinct morphological and biochemical characteristics that reflect distinct processes of the biosynthesis of structural and enzymatic proteins, subsequent structural organization and melanin deposition occurring in these particular cellular compartments. The melanosomes share many common biological properties with the lysosomes. The stage I melanosomes appear to be linked to the late endosomes. Most of melanosomal proteins are glycoproteins that should be folded or assembled correctly in the ER through interaction with calnexin, a chaperone associated with melanogenesis. These melanosomal glycoproteins are then accumulated in the trans Golgi network (TGN) and transported to the melanosomal compartment. During the formation of transport vesicles, coat proteins assemble on the cytoplasmic face of TGN to select their cargos by interacting directly or indirectly with melanosomal glycoproteins to be transported. Adapter protein-3 (AP-3) is important for intracellular transport of tyrosinase gene family proteins from TGN to melanosomes. Tyrosinase gene family proteins possess a di-leucine motif in their cytoplasmic tail, to which AP-3 appears to bind. Thus, the initial cascade of melanosome biogenesis is regulated by several factors including: 1) glycosylation of tyrosinase gene family proteins and their correct folding and assembly within ER and Golgi, and 2) supply of specific signals necessary for intracellular transport of these glycoproteins by vesicles from Golgi to melanosomes.

Asunto(s)

Glicoproteínas/metabolismo , Melanosomas/metabolismo , Monofenol Monooxigenasa/metabolismo , Secuencia de Aminoácidos , Animales , Proteínas de Unión al Calcio/metabolismo , Calnexina , Retículo Endoplásmico/metabolismo , Glicoproteínas/química , Humanos , Melanosomas/ultraestructura , Modelos Biológicos , Monofenol Monooxigenasa/química , Monofenol Monooxigenasa/genética , Pliegue de Proteína , Señales de Clasificación de Proteína , Transporte de Proteínas , Vesículas Transportadoras/metabolismo , Red trans-Golgi/metabolismo

RESUMEN

Glucose effects on the expression of the abscisic acid-inducible Rab16A gene were examined in rice and barley embryos. Glucose feeding to rice embryos negatively affects the endogenous abscisic acid content and represses the promoter activity of the Rab16A gene. Glucose repression of the Rab16A gene takes place both at a transcriptional and a post-transcriptional level. Modulation of the abscisic acid content in rice embryos triggered by glucose did not directly influence the expression of the rice alpha-amylase gene RAmy3D, which is known to be under glucose control. The possible interaction between the glucose and abscisic acid signaling pathway is discussed.

Asunto(s)

Ácido Abscísico/fisiología , Grano Comestible/efectos de los fármacos , Glucosa/farmacología , Proteínas de Choque Térmico/genética , Semillas/efectos de los fármacos , Ácido Abscísico/metabolismo , Técnicas de Cultivo , Grano Comestible/embriología , Grano Comestible/genética , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Hordeum/efectos de los fármacos , Hordeum/embriología , Hordeum/genética , Oryza/efectos de los fármacos , Oryza/embriología , Oryza/genética , Proteínas de Plantas/genética , Estabilidad del ARN/efectos de los fármacos , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Semillas/genética , Semillas/metabolismo , Transcripción Genética/efectos de los fármacos , alfa-Amilasas/genética

RESUMEN

1. To investigate the underlying mechanism for the angiotensin II-induced desensitization of the contractile response during the prolonged stimulation of the vascular smooth muscle, we determined the effects of angiotensin-II on (1) cytosolic Ca2+ concentration ([Ca2+]i) and tension using fura-2-loaded medial strips of the rabbit femoral artery, (2) 45Ca2+ influx in ring preparations, and (3) Ca(2+)-sensitivity of the contractile apparatus in alpha-toxin permeabilized preparations. 2. In the presence of extracellular Ca2+, high concentrations of angiotensin-II elicited biphasic increases in [Ca2+]i and tension, which consisted of initial transient and subsequent lower and sustained phases. 3. The 45Ca2+ influx initially increased after the application of 10(-6) M angiotensin-II, and thereafter gradually decreased. At 20 min after the application, there was a discrepancy between the level of [Ca2+]i and the extent of 45Ca2+ influx. 4. The relationships between [Ca2+]i and tension suggested that the angiotensin-II-induced increase in the Ca(2+)-sensitivity of the contractile apparatus was maintained during the desensitization of smooth muscle contraction. 5. When 10(-6) M angiotensin-II was applied during the sustained phase of contraction induced by 118 mm K(+)-depolarization, at 10 min after the application, the [Ca2+]i levels were significantly lower and the tension levels were significantly higher than those prior to the application of angiotensin-II. 6. In conclusion, the decrease in [Ca2+]i, which is partially due to the inhibition of the Ca2+ influx, is mainly responsible for the desensitization evoked by high concentrations of angiotensin-II, and angiotensin-II seems to activate additional mechanisms which inhibit Ca2+ signaling during prolonged stimulation.

Asunto(s)

Angiotensina II/farmacología , Calcio/metabolismo , Calcio/farmacología , Citosol/metabolismo , Arteria Femoral/efectos de los fármacos , Músculo Liso Vascular/efectos de los fármacos , Animales , Radioisótopos de Calcio , Fura-2 , Técnicas In Vitro , Masculino , Contracción Muscular/efectos de los fármacos , Potasio/farmacología , Conejos , Fosfolipasas de Tipo C/farmacología , Vasoconstricción/efectos de los fármacos

RESUMEN

Sulfur-containing tyrosine analogs such as 4-S-cysteaminylphenol (4-S-CAP) and its N-acetyl derivative, N-acetyl-4-S-CAP, are tyrosinase substrates and can cause selective cytotoxicity or cell death of melanocytes and melanoma cells. It is not clear, however, if the cytotoxicity derives from a cytostatic or cytocidal effect. The latter can also be either apoptotic or necrotic. This paper summarizes our attempt to clarify the nature of melanocytotoxicity and cell death by using a new derivative of 4-S-CAP, N-propionyl-4-S-CAP (NPr-CAP). The i.p. administration of NPr-CAP caused marked depigmentation of black hair follicles in C57 mice. At 12 h postadministration of NPr-CAP, follicular melanocytes showed histochemical and morphologic features indicative of apoptosis by TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining and electron microscopy. The agarose gel electrophoresis of DNA from drug-treated melan a2 cells, an immortal melanocyte line of C57 black mice, showed the nucleosomal DNA ladder pattern. NPr-CAP caused irreversible cytotoxicity in melan a2 and the effect was inhibited by a tyrosinase inhibitor, phenylthiocarbamide. The tyrosinase-mediated cytotoxicity of NPr-CAP was further confirmed by the decreased viability of COS 7 monkey-kidney cells, which expressed a level of high tyrosinase activity through transfection of human tyrosinase cDNA. NPr-CAP, however, also transiently inhibited the proliferation of melan c cells, a control tyrosinase-negative albino melanocyte line, and vector-transfected COS 7 cells. Thus, the major process of NPr-CAP-mediated melanocytotoxicity involves cytocidal apoptosis associated with active tyrosinase. In addition, there is transient, nontyrosinase-mediated cytostatic cytotoxicity.

Asunto(s)

Antineoplásicos/farmacología , Apoptosis , Cistamina/análogos & derivados , Cisteamina/análogos & derivados , Melanocitos/efectos de los fármacos , Monofenol Monooxigenasa/metabolismo , Fenoles/farmacología , Animales , Células COS , Línea Celular , Cistamina/farmacología , Cisteamina/farmacología , Femenino , Color del Cabello/efectos de los fármacos , Folículo Piloso/efectos de los fármacos , Humanos , Cinética , Ratones , Ratones Endogámicos C57BL , Microscopía Electrónica , Estrés Oxidativo , Embarazo , Transfección

RESUMEN

To understand the process of expression of tyrosinase, a key enzyme of melanogenesis, we examined its maturation in the endoplasmic reticulum (ER) by using a heterogeneous expression system. When human tyrosinase cDNA was introduced into COS 7 cells, tyrosinase activity was minimally detected. Immunofluorescence study revealed that tyrosinase was immunolocalized in the nuclear rim, the reticular network, and the punctuated structures. Because a cytoplasmic tail of tyrosinase-gene family protein functions as a lysosomal targeting signal in non-melanocytic cells, and immature and/or misfolded molecules are selectively retained in the ER, the observed localization suggested the inefficient maturation in the COS 7 cells. We thus examined if supplementation of calnexin, a membrane-bound chaperone with affinity for oligosaccharide-processing intermediates containing monoglucose, could improve the process. As expected, the activity was enhanced approximately 2-fold by co-transfection of cDNA encoding calnexin. In contrast, co-transfection of the cytosolic tail-free calnexin, which inhibits calnexin function by allowing premature egress of its ligands from the ER, suppressed expression of this enhanced tyrosinase activity. When alpha-glucosidase activity, which is required for calnexin function, was inhibited by castanospermine (CST) treatment, expression of tyrosinase activity was completely abolished. To confirm the direct involvement of calnexin in tyrosinase maturation, the interaction of calnexin with tyrosinase was examined. Immunoprecipitation of calnexin from extracts of [35S]methionine labeled cells with anti-calnexin antibody revealed that the association is highest immediately after the pulse and that nascent tyrosinase is gradually dissociated upon chase. The association was completely inhibited when CST was included in the medium. Hence, we suggest that the proper folding of tyrosinase is largely dependent on its direct interaction with calnexin for the determined duration in the ER.

Asunto(s)

Proteínas de Unión al Calcio/metabolismo , Retículo Endoplásmico/química , Melaninas/análisis , Monofenol Monooxigenasa/química , Monofenol Monooxigenasa/metabolismo , Animales , Células COS/metabolismo , Proteínas de Unión al Calcio/genética , Calnexina , Secuencia de Carbohidratos , ADN Complementario/metabolismo , Glicosilación , Humanos , Melaninas/biosíntesis , Microscopía Confocal , Datos de Secuencia Molecular , Monofenol Monooxigenasa/genética , Oligosacáridos/metabolismo , Pruebas de Precipitina , Pliegue de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transfección

RESUMEN

There is increasing evidence showing that cereal alpha-amylase gene expression is controlled not only by the classical hormonal regulation, but also by feed-back sugar repression. We demonstrated by in situ hybridization that the sugar repression of rice alpha-amylase gene RAmy3D takes place in scutellar epithelium cells of callus-forming rice embryos. We also used a transient expression system to study the cis-acting elements involved in the sugar repression of the RAmy3D promoter activity. Site-directed mutagenesis of the 50-bp nucleotide sequence from -172 to -123 revealed that consensus sequences of G motif (TACGTA) and TATCCA T/C motif (GATA motif as its antisense sequence) are responsible for sugar repression. The promoter sequences required for sugar repression are reported and discussed.

Asunto(s)

Oryza/enzimología , Oryza/genética , Regiones Promotoras Genéticas , alfa-Amilasas/biosíntesis , alfa-Amilasas/genética , Secuencia de Bases , Secuencia de Consenso , Represión Enzimática , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Guanina , Datos de Secuencia Molecular , Secuencias Reguladoras de Ácidos Nucleicos , Alineación de Secuencia , Eliminación de Secuencia , Homología de Secuencia de Ácido Nucleico

RESUMEN

Tyrosinase-related protein (TRP)-1 is one of the most abundant melanosomal glycoproteins involved in melanogenesis. This report summarizes our recent research efforts related to the biological role and biosynthesis of TRP-1 and its transport from TGN (trans-Golgi network) to the stage I melanosome. Our UV irradiation and tyrosinase and TRP-1 cDNA co-transfection studies indicated that human TRP-1 is involved in not only melanogenesis but also prevention of melanocyte death, which may occur during biosynthesis of melanin pigment in the presence of tyrosinase. Furthermore, a coordinated gene interaction was indicated between tyrosinase and TRP-1, resulting in upregulation of mRNA and protein expression of LAMP (lysosome-associated membrane protein)-1 that would directly prevent the tyrosinase-mediated programmed cell death of melanocytes. Similar to tyrosinase, however, TRP-1 appears to require a molecular chaperone, calnexin, which we have cloned recently. Our cDNA transfection study of tyrosinase with calnexin showed clearly the necessity of calnexin in order to have efficient, functional activity of melanosomal glycoprotein, especially tyrosinase. Once glycosylation is completed, TRP-1 will be transported from TGN to the stage I melanosome. At this stage, TRP-1 will have its own target signal, in particular, tyrosine-rich leucine residues in cytoplasmic tail. Our TRP-1 cDNA transfection and immunoelectron microscopy study shows that TRP-1 will be transported through small vesicles, probably non-clathrin-coated type, to large vacuoles, identical to the MPR (mannose-6-phosphate receptor)-positive, late endosomes. In this transport process a low molecular weight G-protein, rab-7, was isolated from the purified melanosomal protein on 2D-PAGE and identified by subsequent sequencing and PCR amplification. Confocal microscopy with double immunostaining and immunoelectron microscopy confirmed the co-localization of rab-7 and TRP-1 in the melanosomes with early stages of maturation (I-HI). Furthermore, this process will also be regulated by phosphatidylinositol 3-kinase (PI-3 kinase).

Asunto(s)

Endosomas/metabolismo , Aparato de Golgi/metabolismo , Melanocitos/metabolismo , Oxidorreductasas , Proteínas/fisiología , Secuencia de Aminoácidos , Animales , Antígenos CD/metabolismo , Transporte Biológico , Proteínas de Unión al Calcio/metabolismo , Calnexina , Humanos , Proteínas de Membrana de los Lisosomas , Melaninas/biosíntesis , Glicoproteínas de Membrana/metabolismo , Chaperonas Moleculares/metabolismo , Datos de Secuencia Molecular , Pigmentación , Biosíntesis de Proteínas , Señales de Clasificación de Proteína/metabolismo , Proteínas/genética , Piel , Transfección

RESUMEN

1. The mechanisms underlying the relaxation of the porcine renal artery induced by atrial natriuretic peptide (ANP) were investigated, using front-surface fluorimetry with fura-2 and receptor-coupled permeabilization by alpha-toxin. 2. ANP decreased the cytosolic Ca2+ concentration ([Ca2+]i) and tension during the contraction induced by a high external K+ solution, in a concentration-dependent manner. This ANP-induced decrease in [Ca2+]i during the contraction induced by high K+ solution was composed of two phases, an initial rapid phase, followed by a maintenance phase. The initial rapid decrease in [Ca2+]i, but not the maintained decrease in [Ca2+]i, was inhibited when the tissue was treated with thapsigargin, a selective Ca2+ pump inhibitor of the sarcoplasmic reticulum. When the tissues were treated with thapsigargin and external Ca2+ was replaced by Ba2+, which cannot be transported by the Ca2+ pump, ANP did not induce a decrease in [Ba2+]i, even though the elevation of tension induced by Ba2+ was strongly inhibited. 3. In the absence of extracellular Ca2+, ANP inhibited the release of Ca2+ from the intracellular store induced by noradrenaline (NA). 4. The [Ca2+]i (abscissa scale)-tension (ordinate scale) relationship observed during the contraction induced by various concentrations of high external K+ solution was shifted downwards by the addition of 10(-8) M ANP, indicating that, at any given [Ca2+]i, the tension generated by high K+ solution was considerably inhibited by the addition of 10(-8) M ANP. The [Ca2+]i-tension curve of the contraction obtained by the cumulative application of external Ca2+ (0-3.75 mM) during depolarization with 118 mM K+ solution was shifted to the left by 3 x 10(-7) M NA. This NA-induced [Ca2+]i-tension relationship was shifted to the right by 10(-8) M ANP, indicating that the ANP-induced reduction of Ca(2+)-sensitivity operates during the contraction induced by NA. 5. In alpha-toxin-permeabilized preparations, ANP induced relaxation of tissues precontracted with a mixture of 3 x 10(-7) M Ca2+, 10(-5) M guanosine 5'-triphosphate (GTP) and 10(-6) M NA. Thus a component of ANP-induced relaxation took place by way of a reduction in the Ca2+ sensitivity of the myofilaments, independent of changes in [Ca2+]i. 6. These results indicate that ANP induces relaxation of the porcine renal artery by: (1) reducing [Ca2+]i mainly via the activation of the Ca2+ pumps located on the sarcoplasmic reticulum and sarcolemma, as well as via inhibition of agoinist-induced release of Ca2+ from the intracellular store; and (2) decreasing the Ca(2+)-sensitivity of the contractile elements.

Asunto(s)

Factor Natriurético Atrial/farmacología , Arteria Renal/efectos de los fármacos , Animales , Calcio/metabolismo , Técnicas In Vitro , Relajación Muscular/efectos de los fármacos , Potasio/química , Arteria Renal/metabolismo , Arteria Renal/fisiología , Soluciones , Porcinos , Fosfolipasas de Tipo C/farmacología

RESUMEN

Percutaneous injection of absolute ethanol was found to cause rapid destruction of murine and human melanoma nodules. It took only 24-48 hr after the ethanol injection to form ulcers or scars on the lesions. Massive necrosis of murine and human melanoma nests and interstitials and infiltration of mononuclear cells into the demaged tissue were observed histopathologically after ethanol injection. The melanomas recurred approximately 1 week after the destruction of the melanoma nodules by ethanol. Biological response modifiers such as levamisole or interleukin (IL)-2 were effective in preventing the recurrence of murine melanoma after ethanol injection. IL-2 injection caused massive infiltration of NK cells, T cells, and macrophages into the damaged melanoma tissue. The therapeutic modality of a combination of ethanol and IL-2 injections could definitely be useful for the local treatment of human metastatic melanoma, but other modalities such as local hyperthermia or radiation should be considered in combination to attain a permanent cure.

Asunto(s)

Etanol/uso terapéutico , Factores Inmunológicos/uso terapéutico , Melanoma/terapia , Neoplasias Cutáneas/terapia , Anciano , Animales , Supervivencia Celular , Quimioterapia Combinada , Etanol/administración & dosificación , Femenino , Humanos , Factores Inmunológicos/administración & dosificación , Inyecciones Intradérmicas , Melanoma/tratamiento farmacológico , Melanoma/patología , Melanoma Experimental , Ratones , Recurrencia Local de Neoplasia , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/patología , Resultado del Tratamiento

RESUMEN

OBJECTIVE: The aim was to examine the effects of U46619, a thromboxane A2 analog, on intracellular Ca++ concentration and tension in human umbilical artery. STUDY DESIGN: By means of front-surface fluorometry and human umbilical arterial strips loaded with fura-2 we simultaneously measured intracellular Ca++ concentration and tension. RESULTS: In the presence of extracellular Ca++ (1.25 mmol/L), U46619 elevated intracellular Ca++ concentration and tension in a concentration-dependent manner. Oscillations of both intracellular Ca++ concentration and tension were evident in many cases. In the absence of extracellular Ca++ U46619 induced a transient elevation of intracellular Ca++ concentration that was associated with a transient increase and a sustained tension. At any given intracellular Ca++ concentration level the tension induced by the cumulative application of extracellular Ca++ during depolarization with high external K+ in the presence of U46619 was greater than that in the absence of this drug; thus the intracellular Ca++ concentration-tension relationship in the presence of U46619 shifted to the left. CONCLUSION: The tension developed by U46619 depends both on an influx of Ca++ from the extracellular space and on release of intracellular Ca++, and U46619 increased Ca++ sensitivity of the contractile apparatus.

Asunto(s)

Calcio/metabolismo , Endoperóxidos de Prostaglandinas Sintéticos/farmacología , Tromboxano A2/análogos & derivados , Arterias Umbilicales/efectos de los fármacos , Vasoconstricción/efectos de los fármacos , Vasoconstrictores/farmacología , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico , Análisis de Varianza , Femenino , Fluorometría/métodos , Humanos , Técnicas In Vitro , Contracción Muscular/efectos de los fármacos , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiología , Embarazo , Tromboxano A2/farmacología , Arterias Umbilicales/metabolismo , Arterias Umbilicales/fisiología

RESUMEN

We observed transient, erythematous skin eruptions in 6 patients during the intravenous administration of interferon (INF)-alpha for chronic active hepatitis C. The eruptions appeared 5 to 14 days (mean: 6.8 days) after initiating its administration. They were localized or disseminated and consisted of edematous, erythematous, and/or papular changes. Vesicles and petechiae also appeared in some cases. The eruptions disappeared in 10 to 14 days, despite the continuance of INF-alpha and without specific treatment. Histological examination obtained from skin eruptions revealed perivascular infiltration of lymphoid cells, the majority which were CD4-positive, of the upper dermis. Edematous changes were present in the papillary dermis. Vascular endothelial cells in the upper dermis expressed both intercellular adhesion molecule-1 (ICAM-1) and endothelial leukocyte adhesion molecule-1 (ELAM-1), while the epidermal keratinocytes produced neither. These findings suggested a nonallergic mechanism for such eruptions.

Asunto(s)

Erupciones por Medicamentos/etiología , Erupciones por Medicamentos/patología , Interferón-alfa/efectos adversos , Adulto , Vesícula/etiología , Vesícula/patología , Linfocitos T CD4-Positivos/patología , Adhesión Celular , Moléculas de Adhesión Celular/metabolismo , Selectina E , Edema/etiología , Edema/patología , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Eritema/etiología , Eritema/patología , Femenino , Hepatitis C/terapia , Hepatitis Crónica/terapia , Humanos , Inmunohistoquímica , Inyecciones Intravenosas , Molécula 1 de Adhesión Intercelular/metabolismo , Interferón-alfa/administración & dosificación , Queratinocitos/metabolismo , Masculino , Persona de Mediana Edad , Púrpura/etiología , Púrpura/patología , Receptores Inmunológicos/metabolismo , Piel/patología

RESUMEN

The murine melanoma cell line BL-6-beta m, which is a stable cell line transfected with a gene coding a unique actin subspecies called beta m to the BL-6 cell line, has low metastatic potentials as compared with those of the parent cell line. BL-6-beta m melanomas were found to be sensitive to in vivo local injection of IL-2, while BL-6 melanomas showed almost no response. Ganglioside analysis of BL-6 and BL-6-beta melanomas revealed that the main ganglioside of both melanomas was GM3, which suggested that different sensitivities between BL-6 and BL-6-beta m melanomas to the injection of IL-2 did not relate to the different compositions of main gangliosides. However, minor components of the gangliosides such as GM2 and GM1 emerged only in BL-6-beta m melanomas after treatment with IL-2. Local injection of IL-2 caused considerable infiltration of anti-asialo GM1-positive cells into the nests as well as the interstitials of BL-6-beta m melanomas. In contrast, in the BL-6 melanomas treated with IL-2, infiltration of the anti-asialo GM1-positive cells was hardly seen, although anti-Thy1,2 and anti-macrophage-positive cells were found to more or less the same extent as observed in BL-6-beta m melanomas. These results suggest that the murine metastatic variant melanoma cell lines BL-6 and BL-6-beta m have different properties in terms of sensitivity to in vivo IL-2 treatment, and a slight enhancement of the ganglioside components GM2 and GM1 expression only in BL-6-beta m after IL-2 treatment may play a role in the IL-2-mediated attraction of immune cells or may explain the different sensitivities of the two lines to treatment with IL-2.

Asunto(s)

Actinas/fisiología , Gangliósidos/análisis , Factores Inmunológicos/uso terapéutico , Interleucina-2/uso terapéutico , Melanoma Experimental/terapia , Transfección , Actinas/genética , Animales , Antígenos de Neoplasias/análisis , Movimiento Celular , Femenino , Gangliósido G(M1)/análisis , Factores Inmunológicos/farmacología , Vigilancia Inmunológica , Inyecciones Intralesiones , Interleucina-2/farmacología , Células Asesinas Activadas por Linfocinas/inmunología , Melanoma Experimental/química , Melanoma Experimental/inmunología , Melanoma Experimental/patología , Ratones , Ratones Endogámicos C57BL , Metástasis de la Neoplasia , Trasplante de Neoplasias , Proteínas Recombinantes de Fusión , Antígenos Thy-1/análisis

RESUMEN

Local microwave hyperthermia in combination with local injections of interleukin 2 (IL-2) was found to exert a remarkable suppressive effect on murine melanoma growth. Both of these therapeutic modalities caused marked tumour infiltration with natural killer cells. After microwave hyperthermia or IL-2 injection alone there was minimal T-cell infiltration, but T cells were more in evidence following combination therapy.

Asunto(s)

Calor/uso terapéutico , Interleucina-2/administración & dosificación , Melanoma Experimental/terapia , Microondas , Neoplasias Cutáneas/terapia , Animales , Terapia Combinada , Femenino , Técnica del Anticuerpo Fluorescente , Inyecciones Intralesiones , Células Asesinas Naturales/inmunología , Melanoma Experimental/inmunología , Ratones , Ratones Endogámicos C57BL , Neoplasias Cutáneas/inmunología

RESUMEN

Murine B16-F10 melanoma was treated with local microwave hyperthermia, local injection of murine recombinant interferon-beta (rIFN-beta) or a combination of both in order to investigate the augmentation of anti-proliferative effects with this combination treatment. Concerning the local modulation of immunological reactions of the host, local hyperthermia at 43 degrees C for 15 min on murine melanoma caused remarkable infiltration of natural killer cells and local injection of rIFN-beta led to considerable infiltration of T cells. When these two modalities were combined, the infiltration of NK cells completely disappeared and, instead, remarkable augmentation of T cell infiltration occurred. Synergistic suppressive effects on melanoma growth with occasional scar formation were seen with this combined modality. These results indicate that local hyperthermia with a combination of rIFN-beta modulates local immune reactions of the host, and probably this immune reaction is partly involved in the course of the suppression of tumor growth.

Asunto(s)

Hipertermia Inducida , Interferón beta/uso terapéutico , Melanoma Experimental/patología , Melanoma Experimental/terapia , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/terapia , Animales , División Celular/fisiología , Movimiento Celular , Terapia Combinada , Interferón beta/farmacología , Ratones , Ratones Endogámicos C57BL , Microondas , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Linfocitos T/patología , Linfocitos T/fisiología , Células Tumorales Cultivadas

RESUMEN

We report a case of transitional cell carcinoma of the ureter and bladder that produced carbohydrate antigen 19-9 and carcinoembryonic antigen. The serum levels of these antigens were elevated in this patient and an immunohistochemical examination revealed that the carcinoma cells stained positively for both antigens.

Asunto(s)

Antígenos de Carbohidratos Asociados a Tumores/biosíntesis , Antígeno Carcinoembrionario/biosíntesis , Carcinoma de Células Transicionales/inmunología , Neoplasias Primarias Múltiples/inmunología , Neoplasias Ureterales/inmunología , Neoplasias de la Vejiga Urinaria/inmunología , Carcinoma de Células Transicionales/patología , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Primarias Múltiples/patología , Neoplasias Ureterales/patología , Neoplasias de la Vejiga Urinaria/patología