RESUMEN
Energy metabolism reprogramming is becoming an increasingly important hallmark of cancer. Specifically, cancers tend to undergo metabolic reprogramming to upregulate a celldependent glutamine (Gln) metabolism. Notably, hepatocellular cell adhesion molecule (HepaCAM) has been previously reported to serve a key role as a tumour suppressor. However, the possible regulatory role of HepaCAM in Gln metabolism in prostate cancer (PCa) remains poorly understood. In the present study, bioinformatics analysis predicted a significant negative correlation among the expression of HepaCAM, phosphatidylinositol4,5bisphosphate 3kinase catalytic subunit α (PIK3CA), glutaminase (GLS) and solute carrier family 1 member 5 (SLC1A5), components of Gln metabolism, in clinical and genomic datasets. Immunohistochemistry results verified a negative correlation between HepaCAM and PIK3CA expression in PCa tissues. Subsequently, liquid chromatographytandem mass spectrometry (LCMS/MS) and gas chromatographymass spectrometry (GCMS) assays were performed, and the results revealed markedly reduced levels of Gln and metabolic flux in the blood samples of patients with PCa and in PCa cells. Mechanistically, overexpression of HepaCAM inhibited Gln metabolism and proliferation by regulating PIK3CA in PCa cells. In addition, Gln metabolism was discovered to be stressresistant in PCa cells, since the expression levels of GLS and SLC1A5 remained high for a period of time after Gln starvation. However, overexpression of HepaCAM reversed this resistance to some extent. Additionally, alpelisib, a specific inhibitor of PIK3CA, effectively potentiated the inhibitory effects of HepaCAM overexpression on Gln metabolism and cell proliferation through mass spectrometry and CCK8 experiments. In addition, the inhibitory effect of PIK3CA on the growth of tumor tissue in nude mice was also confirmed by immunohistochemistry in vivo. To conclude, the results from the present study revealed an abnormal Gln metabolic profile in the blood samples of patients with PCa, suggesting that it can be applied as a clinical diagnostic tool for PCa. Additionally, a key role of the HepaCAM/PIK3CA axis in regulating Gln metabolism, cell proliferation and tumour growth was identified. The combination of alpelisib treatment with the upregulation of HepaCAM expression may serve as a novel method for treating patients with PCa.
Asunto(s)
Proliferación Celular/genética , Glutamina/metabolismo , Transducción de Señal/genética , Animales , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral/metabolismo , Proliferación Celular/fisiología , Fosfatidilinositol 3-Quinasa Clase I/genética , Fosfatidilinositol 3-Quinasa Clase I/metabolismo , Modelos Animales de Enfermedad , Glutamina/genética , Masculino , Ratones , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/prevención & controlRESUMEN
Hematological and biochemical profile studies help to evaluate functional changes of animals used in experiments. The aim of this study was to determine the hematological and biochemical profile of immunosuppressed BALB/c nude and C57BL/6 SCID mice after bovine ovarian xenotransplantation. Therefore, a total of 74 female mice were divided into four groups: non-xenotransplanted animals, xenotransplanted animals, xenotransplanted animals treated with eCG and xenotransplanted animals treated with FSH + LH. After anesthesia, blood samples were collected and hematologic and biochemical values were evaluated. The results showed no significant differences (p ≤ 0.05) for hematological parameters between the control group and the treatment groups of both strains. However, considering the biochemical profile, it was observed an increase of AST concentrations (p ≤ 0.05) in both strains and a decrease of ALT concentrations (p ≤ 0.05) only in C57BL/6 SCID strain of the groups subjected to hormonal treatment compared with those non subjected. Additionally, the values of the renal enzymes, urea and creatinine, did not differ (p ≤ 0.05) between the groups. Our findings suggest that the xenotransplantation procedure as well as the hormonal dosages had no significant effect on the well-being of the animals considering the evaluated hematological and biochemical profile.
Asunto(s)
Recuento de Células Sanguíneas , Proteínas Sanguíneas/análisis , Ratones Endogámicos C57BL/metabolismo , Ratones Desnudos/metabolismo , Ratones SCID/metabolismo , Ovario/trasplante , Trasplante Heterólogo/métodos , Animales , Fenómenos Bioquímicos , Femenino , Hormona Folículo Estimulante/administración & dosificación , Riñón/metabolismo , Hígado/metabolismo , Hormona Luteinizante/administración & dosificación , Ratones , Ratones Endogámicos C57BL/sangre , Ratones Desnudos/sangre , Ratones SCID/sangre , Modelos AnimalesRESUMEN
WW domain-binding protein 2 (WBP2) has been demonstrated as oncogenic in breast cancer. Many studies have revealed the WBP2 gene as a high-risk gene for leukoariaosis and cerebral white matter lesions is important in the pathologic stage of glioma development. This study aimed to illustrate the underlying mechanism by which WBP2 regulates the process of glioma development. The expression pattern of WBP2 in several tumor cells was determined, clarifying the carcinogenic action of WBP2 in glioma cells. Overexpression of WBP2 in glioma cells promoted cell proliferation and migration, and the number of S-phase cells, whereas the depletion of WBP2 by RNAi-mediated knockdown restrained cell growth and cell cycle progression. Upregulation of WBP2 significantly enhanced the tumorigenic ability of U251 cells in vivo. MS/GST pulldown assay identified α-enolase (ENO1) and Homer protein homolog 3 (Homer3) as novel potent interaction partners of WBP2. Knockdown of ENO1 or Homer3 allowed cell growth and migration to return to normal levels. Furthermore, in vitro and in vivo experiments indicated that the oncogenic role of WBP2 in glioma was through modulating ENO1 and glycolysis activity via the ENO1-PI3K/Akt signaling pathway. Collectively, these results reveal that WBP2 plays a vital role in the occurrence and development of glioma, indicating a target gene for glioblastoma treatment.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Biomarcadores de Tumor/metabolismo , Neoplasias Encefálicas/metabolismo , Proteínas de Unión al ADN/metabolismo , Glioma/metabolismo , Ratones Desnudos/metabolismo , Fosfopiruvato Hidratasa/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Adulto , Anciano , Anciano de 80 o más Años , Animales , Biomarcadores de Tumor/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/fisiopatología , Ciclo Celular , Movimiento Celular , Proliferación Celular , Proteínas de Unión al ADN/genética , Femenino , Glioma/genética , Glioma/fisiopatología , Glucólisis , Proteínas de Andamiaje Homer/genética , Proteínas de Andamiaje Homer/metabolismo , Humanos , Masculino , Ratones , Ratones Desnudos/genética , Persona de Mediana Edad , Oncogenes , Fosfopiruvato Hidratasa/genética , Unión Proteica , Transactivadores , Proteínas Supresoras de Tumor/genéticaRESUMEN
Advances in tissue engineering led to the development of various tissue-engineered skin substitutes (TESS) for the treatment of skin injuries. The majority of the autologous TESS required lengthy and costly cell expansion process to fabricate. In this study, we determine the possibility of using a low density of human skin cells suspended in platelet-rich plasma (PRP)-enriched medium to promote the healing of full-thickness skin wounds. To achieve this, full-thickness wounds of size 1.767 cm2 were created at the dorsum part of nude mice and treated with keratinocytes (2 × 104 cells/cm2) and fibroblasts (3 × 104 cells/cm2) suspended in 10% PRP-enriched medium. Wound examination was conducted weekly and the animals were euthanized after 2 weeks. Gross examination showed that re-epithelialization was fastest in the PRP+cells group at both day 7 and 14, followed by the PRP group and NT group receiving no treatment. Only the PRP+cells group achieved complete wound closure by 2 weeks. Epidermal layer was presence in the central region of the wound of the PRP+cells and PRP groups but absence in the NT group. Comparison between the PRP+cells and PRP groups showed that the PRP+cells-treated wound was more mature as indicated by the presence of thinner epidermis with single cell layer thick basal keratinocytes and less cellular dermis. In summary, the combination of low cell density and diluted PRP creates a synergistic effect which expedites the healing of full-thickness wounds. This combination has the potential to be developed as a rapid wound therapy via the direct application of freshly harvested skin cells in diluted PRP.
Asunto(s)
Plasma Rico en Plaquetas/citología , Piel Artificial/normas , Cicatrización de Heridas , Animales , Fibroblastos/patología , Fibroblastos/fisiología , Queratinocitos/patología , Queratinocitos/fisiología , Ratones Desnudos/lesiones , Ratones Desnudos/metabolismo , Plasma Rico en Plaquetas/metabolismo , Piel/efectos de los fármacos , Piel/lesiones , Traumatismos de los Tejidos Blandos/terapiaRESUMEN
Diabetes is induced in mice by using streptozotocin (STZ), a compound that has a preferential toxicity toward pancreatic ß cells. We evaluated nude male mice from various sources for their sensitivity to a single high dose (160 to 240 mg/kg) of STZ. Diabetes was induced in male mice (age: median, 12 wk; interquartile range, 11 to 14 wk; body weight, about 30 g) from Taconic Farms (TAC), Jackson Laboratories (JAX), and Charles River Laboratories (CRL). Mice were monitored for 30 d for adverse side effects, blood glucose, and insulin requirements. In CRL mice given 240 mg/kg STZ, more than 95% developed diabetes within 4 to 5 d, and loss of body weight was relatively low (mean, 0.4 g). In comparison, both TAC and JAX mice were more sensitive to STZ, as evidenced by faster development of diabetes (even at a lower STZ dose), greater need for insulin after STZ, greater body weight loss (mean: TAC, 3.5 g; JAX, 3.7 g), and greater mortality. We recommend conducting exploratory safety assessments when selecting a nude mouse source, with the aim of limiting morbidity and mortality to less than 10%.
Asunto(s)
Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Experimental/patología , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Estreptozocina/toxicidad , Análisis de Varianza , Animales , Masculino , Ratones , Especificidad de la EspecieRESUMEN
Hallmarks of malignant melanoma are its propensity to metastasize and its resistance to treatment, giving patients with advanced disease a poor prognosis. The transition of melanoma from non-invasive radial growth phase (RGP) to invasive and metastatically competent vertical growth phase (VGP) is a major step in tumor progression, yet the mechanisms governing this transformation are unknown. Matrix metalloproteinase-1 (MMP-1) is highly expressed by VGP melanomas, and is thought to contribute to melanoma progression by degrading type I collagen within the skin to facilitate melanoma invasion. Protease activated receptor-1 (PAR-1) is activated by MMP-1, and is also expressed by VGP melanomas. However, the effects of MMP-1 signaling through PAR-1 have not been examined in melanoma. Here, we demonstrate that an MMP-1/PAR-1 signaling axis exists in VGP melanoma, and is necessary for melanoma invasion. Introduction of MMP-1 into RGP melanoma cells induced gene expression associated with tumor progression and promoted invasion in vitro, and enhanced tumor growth and conferred metastatic capability in vivo. This study demonstrates that both the type I collagenase and PAR-1 activating functions of MMP-1 are required for melanoma progression, and suggests that MMP-1 may be a major contributor to the transformation of melanoma from non-invasive to malignant disease.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Metaloproteinasa 1 de la Matriz/metabolismo , Melanoma/fisiopatología , Invasividad Neoplásica/fisiopatología , Receptor PAR-1/fisiología , Transducción de Señal/fisiología , Neoplasias Cutáneas/fisiopatología , Animales , Línea Celular Tumoral , Colagenasas/metabolismo , Progresión de la Enfermedad , Femenino , Expresión Génica , Humanos , Masculino , Metaloproteinasa 1 de la Matriz/fisiología , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Melanoma/patología , Ratones , Ratones Desnudos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Neovascularización Patológica , Osteonectina/metabolismo , Neoplasias Cutáneas/patologíaRESUMEN
In this study, we addressed the hypothesis that transcriptional suppression of erythroblastosis virus E26 oncogene homolog 1 (ETS-1) is an efficient therapeutic approach to pancreatic adenocarcinoma by investigating the effect of ETS-1 suppression in human pancreatic cancer cells. We accomplished this by using an adenoviral vector encoding only the DNA-binding domain of wild-type ETS-1 (ETS-1 dominant negative, ETS-1-DN). ETS-1-DN decreases ETS-1-binding by competing for its binding to DNA. Adenoviral-mediated transfer of ETS-1-DN (adenoviral ETS-1-DN construct, AdETS-1-DN) into pancreatic tumor cell lines did not affect their proliferation rate in vitro but did significantly inhibit their in vivo growth in nude mice. Furthermore, to test the efficacy of ETS-1-DN in vivo, we injected the AdETS-1-DN into established human pancreatic adenocarcinomas grown in nude mice. This treatment significantly reduced tumor size as compared to saline injection, without any detectable side effects. Microvessel density in mouse xenografts displayed significantly lower values in tumors in which ETS-1 was downregulated. In addition, expression of the ETS-1-DN in the pancreatic cancer cells resulted in downregulation of urokinase-type plasminogen activator (u-PA) and metalloproteinase-1 (MMP-1) expression. Taken together, these data suggest that transcriptional inactivation of ETS-1 is able to significantly affect angiogenesis and growth of pancreatic cancer. This effect may be due in part to downregulation of MMP-1 and u-PA expression. Our results suggest that ETS-1-DN is a promising candidate for antiangiogenic gene therapy in pancreatic cancer.
Asunto(s)
Adenocarcinoma/terapia , Silenciador del Gen , Neovascularización Patológica/terapia , Neoplasias Pancreáticas/terapia , Proteína Proto-Oncogénica c-ets-1/metabolismo , Transcripción Genética/fisiología , Adenocarcinoma/irrigación sanguínea , Adenoviridae/genética , Animales , Línea Celular Tumoral , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Regulación Neoplásica de la Expresión Génica , Terapia Genética , Humanos , Masculino , Ratones , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Ratones SCID , Neoplasias Pancreáticas/irrigación sanguínea , Trasplante HeterólogoRESUMEN
Four groups of 16 age-matched female Crl:SKH1-hrBR hairless mice were exposed to either control soil or polychlorinated biphenyl (PCB)-contaminated soil (retrieved from an electrical waste landfill in Southern Illinois) for 11 weeks. The mice were exposed in a study to determine interactions between environmental PCBs and ultraviolet radiation (UVR), but the UVR group did not differ and provided a replicate for the residue study. Ear biopsies were performed immediately after the termination of soil exposure. The mice were maintained in regular bedding for 37 weeks thereafter. The ear-skin, trunk-skin, fat-pad, and liver samples were collected and weighed at the end of the study (week 48) and analyzed for PCB residues. A total of 141 PCB congeners were target analytes. There were significant differences in body weights and food consumption from week 2 to 28. The liver weights of mice treated with PCB only were significantly greater than those of UVR-treated mice. The fat-pad weight did not differ among treated groups. PCB residues in the ear biopsies specimens of mice exposed to contaminated soil were 342.3 and 317.2 ppm in the PCB- and PCB + UVR-treated groups, respectively, and contained both persistent and episodic congeners. After 37 weeks of isolation from soil, the ear PCB residues decreased to 21.5 ppm (PCB group) and 14.5 ppm (PCB + UVR group), and only persistent congeners contributed to the total PCB residues. The accumulation of PCB residues was highest in the fat pad (fat pad > ear skin > trunk skin > liver) in both PCB +/- UVR groups at the end of the study. However, the percentage of individual congeners contributing to total PCBs in these different tissues did not differ.
Asunto(s)
Ratones Desnudos/metabolismo , Bifenilos Policlorados/farmacocinética , Contaminantes del Suelo/farmacocinética , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Tejido Adiposo/patología , Animales , Carga Corporal (Radioterapia) , Peso Corporal/efectos de los fármacos , Residuos de Medicamentos/metabolismo , Oído Externo/efectos de los fármacos , Oído Externo/metabolismo , Oído Externo/patología , Ingestión de Alimentos/efectos de los fármacos , Femenino , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Endogámicos , Tamaño de los Órganos/efectos de los fármacos , Bifenilos Policlorados/toxicidad , Piel/efectos de los fármacos , Piel/metabolismo , Piel/patología , Contaminantes del Suelo/toxicidad , Factores de Tiempo , Distribución TisularRESUMEN
The hypothesis that stem cells may seed cancer has emerged from the cancer stem cells concept. However, the experimental systems necessary to provide more direct evidence to support the hypothesis have been lacking. We have used fetal neural progenitor cells (hNPC) transduced with the telomerase hTERT gene to investigate the neoplastic potential of hNPCs. The hTERT-transduced line, hNPCs-G3 lost normal diploid karyotype, showed loss of contact inhibition, anchorage independence, and formed neuroblastoma-like tumours in all of 10 mice. These data suggest that hNPCs have the potential for neoplastic transformation. These data have implications for providing a novel tool to test the feasibility of new anticancer treatment strategies and raise the possibility of a risk for the use of hNPCs in cell transplantation.
Asunto(s)
Transformación Celular Neoplásica , Feto/citología , Regulación Neoplásica de la Expresión Génica/fisiología , Neuronas/fisiología , Células Madre/fisiología , Animales , Southern Blotting , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Recuento de Células/métodos , Diferenciación Celular/fisiología , Línea Celular Tumoral , ADN/metabolismo , Proteínas de Unión al GTP , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Inmunohistoquímica/métodos , Proteínas de Filamentos Intermediarios/genética , Cariotipificación/métodos , Ratones , Ratones Desnudos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas del Tejido Nervioso/genética , Nestina , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Complejo Represivo Polycomb 1 , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , ARN Mensajero/biosíntesis , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Coloración y Etiquetado , Telómero/metabolismo , Factores de Tiempo , Transducción Genética/métodos , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismoRESUMEN
Inhibitors of topoisomerases are widely used in the treatment of cancer, including inhibitors of topoisomerase I (camptothecin analogs such as irinotecan and topotecan) and topoisomerase II (etoposide and doxorubicin). The novel bis-phenazine, XR5944, is a joint inhibitor of topoisomerase I and II as shown by the stabilization of topoisomerase-dependent cleavable complexes. XR5944 demonstrated exceptional activity against human and murine tumor cells in vitro and in vivo. In a range of cell lines XR5944 (IC50 0.04-0.4 nM) was significantly more potent than TAS-103, originally proposed as a joint topoisomerase I and II inhibitor, as well as agents specific for topoisomerase I or II (topotecan, doxorubicin and etoposide). In addition, XR5944 was unaffected by atypical drug resistance and retained significant activity in cells overexpressing P-glycoprotein or multidrug resistance-associated protein. Antitumor efficacy of XR5944 was demonstrated in human carcinoma xenograft models (H69 small cell lung cancer and HT29 colon). In the HT29 model, which is relatively unresponsive to chemotherapy, XR5944 (15 mg/kg i.v., q4dx3) induced tumor regression in the majority of animals (six of eight), whereas TAS-103, dosed at its maximum tolerated dose (45 mg/kg i.v., q7dx3), only induced a delay in tumor growth compared with control animals. In the H69 model, low doses of XR5944 (5 mg/kg i.v., qdx5/week for 2 weeks or 10-15 mg/kg i.v., q4dx3), induced complete tumor regression in the majority of animals. In contrast, topotecan (20 mg/kg i.v., q4dx3) or etoposide (30 mg/kg i.v., q5dx5) only slowed the tumor growth rate. These studies show that XR5944 is a highly active novel anticancer agent that is well tolerated at efficacious doses.
Asunto(s)
Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias del Colon/tratamiento farmacológico , ADN-Topoisomerasas de Tipo II , Isoenzimas/antagonistas & inhibidores , Neoplasias Pulmonares/tratamiento farmacológico , Fenazinas/farmacología , Inhibidores de Topoisomerasa I , Inhibidores de Topoisomerasa II , Aminoquinolinas/metabolismo , Aminoquinolinas/farmacología , Animales , Antígenos de Neoplasias , Antineoplásicos/metabolismo , Antineoplásicos/toxicidad , ADN/química , ADN/metabolismo , ADN-Topoisomerasas de Tipo I/metabolismo , ADN-Topoisomerasas de Tipo II/metabolismo , Proteínas de Unión al ADN , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo , Doxorrubicina/metabolismo , Doxorrubicina/farmacología , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Etopósido/metabolismo , Etopósido/farmacología , Femenino , Humanos , Indenos/metabolismo , Indenos/farmacología , Concentración 50 Inhibidora , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Isoenzimas/metabolismo , Ratones , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Fenazinas/metabolismo , Fenazinas/toxicidad , Inducción de Remisión , Topotecan/metabolismo , Topotecan/farmacología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Ewing's sarcoma shows a strong tendency to metastasize to the lungs or the skeleton, or both. A peculiar feature of the secondary involvement of bone with this tumor is that it may also appear in the absence of clinically evident lung metastases, both at clinical presentation and during the course of the disease. Although osseous metastases are critically relevant for prognosis, the pathogenesis of this peculiar feature of Ewing's sarcoma is poorly understood, partly due to the lack of appropriate experimental in vivo models. We show that the intravenous injection of TC-71 Ewing's sarcoma cells into athymic 4-5-week-old Crl/nu/nu (CD1) BR mice reproducibly colonizes specific sites of the skeleton in addition to the lungs and lymph nodes. The distribution and the morphologic appearance of these experimental bone metastases mimic the pattern of skeletal involvement observed in humans. This experimental model of bone metastasis of Ewing's sarcoma may be the basis for future studies aimed at understanding the pathophysiology and treatment of Ewing's sarcoma.
Asunto(s)
Neoplasias Óseas/secundario , Modelos Animales de Enfermedad , Metástasis de la Neoplasia/fisiopatología , Sarcoma de Ewing/secundario , Antígeno 12E7 , Animales , Antígenos CD/metabolismo , Neoplasias Óseas/metabolismo , Moléculas de Adhesión Celular/metabolismo , Femenino , Glucógeno/metabolismo , Integrinas/metabolismo , Neoplasias Pulmonares/secundario , Metaloproteinasas de la Matriz/metabolismo , Ratones , Ratones Desnudos/metabolismo , Receptores de Factores de Crecimiento/metabolismo , Sarcoma de Ewing/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Células Tumorales Cultivadas/metabolismo , Células Tumorales Cultivadas/patologíaRESUMEN
Mutations in the winged-helix nude (whn) gene result in the nude mouse and rat phenotypes. The pleiotropic nude phenotype which affects the hair, skin, and thymus suggests that whn plays a pivotal role in the development and/or maintenance of these organs. However, little is known about whn function in these organs. We show here that in skin whn is specifically expressed in epithelial cells and not the mesenchymal cells, and using a hair reconstitution assay, we demonstrate that the abnormal nude mouse hair development is attributable to a functional defect of the epithelial cells. Examination of nude mouse primary keratinocytes in culture revealed that these cells have an increased propensity to differentiate in an abnormal fashion, even under conditions that promote proliferation. Furthermore, nude mouse keratinocytes displayed a 100-fold increased sensitivity to the growth-inhibitory/differentiation effects of the phorbol ester TPA. In parallel with these findings, we directly show that whn functions as a transcription factor that can specifically suppress expression of differentiation/TPA-responsive genes. The region of Whn responsible for these effects was mapped to the carboxy-terminal transactivating domain. These results establish whn as a key regulatory factor involved in maintaining the balance between keratinocyte growth and differentiation. The general implications of these findings for an epithelial self-renewal model will be discussed.
Asunto(s)
Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Piel/crecimiento & desarrollo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Animales , Northern Blotting , Células Cultivadas , Proteínas de Unión al ADN/fisiología , Epitelio/crecimiento & desarrollo , Factores de Transcripción Forkhead , Regulación del Desarrollo de la Expresión Génica , Cabello/metabolismo , Immunoblotting , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Ratones , Ésteres del Forbol/efectos adversos , Plásmidos , Regiones Promotoras Genéticas/fisiología , Piel/citología , Factores de Transcripción/fisiología , Transcripción Genética , TransfecciónRESUMEN
A partial cDNA clone, RLC34, was isolated from a rat brain cDNA library. Its sequence exhibits high identity with BAT3 (88.4% and 94.9% for DNA and the deduced amino acid sequence, respectively), a gene located within the region of human major histocompatibility complex III (MCHIII region). RLC34 detected a transcript the same size in human and rat, similar to that reported for BAT3. Southern blot analysis of RLC34 showed similar restriction patterns as those of the human BAT3 gene. A panel of rodent tissue samples were examined and the RLC34 was found to be predominantly expressed in the germ cells of rodent testes. The expression is developmentally regulated with increased transcripts seen at 17-20 days after birth. Its testicular expression, its association with spermatogenesis, and its location in MCHIII suggest a correlation of RLC34 with the growth-reproduction complex (grc). This finding may also provide a clue to study the function of other genes localized in this area of the MCHIII region.
Asunto(s)
Biosíntesis de Proteínas , Ratas/genética , Testículo/metabolismo , Adulto , Animales , Secuencia de Bases , Proteínas Portadoras , Cricetinae , ADN Complementario/genética , Expresión Génica , Humanos , Masculino , Mesocricetus/genética , Mesocricetus/metabolismo , Ratones , Ratones Desnudos/genética , Ratones Desnudos/metabolismo , Chaperonas Moleculares , Datos de Secuencia Molecular , Proteínas Nucleares , Sistemas de Lectura Abierta , Especificidad de Órganos , Proteínas/genética , Ratas/metabolismo , Ratas Sprague-Dawley , Alineación de Secuencia , Homología de Secuencia , Especificidad de la EspecieRESUMEN
The ultra-immunocytochemical technique applied in the present study revealed the occurrence of methionine-enkephalin (met-enkephalin)-like substance in the dense-core granules of Merkel cells of nude mice sinus hair. Incubation of ultra-thin sections of sinus hair with met-enkephalin antisera conjugated with gold particles showed specific association of gold particles on the dense-core granules of the Merkel cells. Gold particles were heavily and specifically located on the dense-core granules as well as in the adjacent cytoplasm. Dense-core granules of degenerating Merkel cells also exhibit met-enkephalin immunoreactivity. The nerve terminals associated with the Merkel cell did not show met-enkephalin immunoreactivity. Therefore, it is concluded that a met-enkephalin-like substance is present and stored in nude mice Merkel cell dense-core granules and it might act as a neurotransmitter or neuromodulator which could be involved in the functioning of the cell. Non-osmicated tissue should be used to locate this substance because of the possibility of cross-linkage of the amino acid sequence with osmium tetroxide.
Asunto(s)
Células APUD/ultraestructura , Gránulos Citoplasmáticos/química , Encefalina Metionina/análisis , Ratones Desnudos/metabolismo , Vibrisas/citología , Células APUD/química , Animales , Inmunohistoquímica , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos/anatomía & histología , Microscopía Electrónica/métodosRESUMEN
The pharmacokinetics of a disulfide linked conjugate of a murine monoclonal antibody A7 with neocarzinostatin (A7-NCS) was studied following its intravenous administration to nude mice. Disappearance of the conjugate from the circulation was biphasic: an early rapid phase was followed by a much slower phase. The conjugate was removed from the blood circulation with a half-life of 12 hr, showing nearly the same kinetics as the free antibody. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that the disulfide linkage in A7-NCS was stable at least for 48 hr after administration of the conjugate to nude mice. The conjugate concentration in a human colon cancer SW1116 derived tumor reached maximum at 24 hr after injection and remained high for an additional 24 hr. The passive hemagglutination inhibition assay revealed that NCS in the conjugated form can be efficiently delivered to the target tissue. The present report indicates that A7-NCS was sufficiently stable in circulation to reach the target tumor without releasing NCS.
Asunto(s)
Anticuerpos Monoclonales/metabolismo , Cinostatina/farmacocinética , Animales , Anticuerpos Monoclonales/administración & dosificación , Neoplasias del Colon/metabolismo , Portadores de Fármacos , Estabilidad de Medicamentos , Semivida , Humanos , Inyecciones Intravenosas , Radioisótopos de Yodo/farmacocinética , Ratones , Ratones Desnudos/metabolismo , Trasplante de Neoplasias , Neoplasias Experimentales/diagnóstico por imagen , Radioinmunodetección , Trasplante Heterólogo , Cinostatina/administración & dosificaciónRESUMEN
We characterized the tumorigenic and metastatic potential of a poorly differentiated, non-CEA-producing colon cancer cell line, MIP-101, after injection at different sites in athymic mice. After subcutaneous and intrasplenic injection tumor grew locally in 100 and 50%, respectively, but no metastases were found, even after intravenous injection. Intraperitoneal implantation, however, resulted in a high tumor take (10/10) and subsequent liver colonization (8/10 mice). Exogenous CEA prior to intrasplenic injection induced metastasis in 7/8 mice (in 2 mice to the liver and in 5 mice to the lung). Intrasplenic injection of CX-1, a good CEA producer, resulted in hepatic metastases in 100% of the animals. These data suggest a direct or indirect role of CEA in the metastatic process. We conclude that MIP-101 has a high tumorigenic and invasive potential but a low metastatic proclivity, except when grown in the peritoneum, and that pretreatment of tumor-bearing animals with CEA affects the metastatic proclivity.
Asunto(s)
Adenocarcinoma/patología , Neoplasias del Colon/patología , Metástasis de la Neoplasia/patología , Animales , Antígeno Carcinoembrionario/sangre , Antígeno Carcinoembrionario/farmacocinética , Antígeno Carcinoembrionario/farmacología , Humanos , Inyecciones Intravenosas , Neoplasias Hepáticas Experimentales/secundario , Ratones , Ratones Desnudos/metabolismo , Trasplante de Neoplasias , Neoplasias Peritoneales/patología , Bazo/patología , Células Tumorales CultivadasRESUMEN
The biodistribution of yttrium- and indium-labeled monoclonal antibody (MAb) B72.3 IgG using three different chelate conjugates (SCN-Bz-EDTA, CA-DTPA, and SCN-Bz-DTPA) was compared in athymic mice bearing LS-174T tumors. The 88Y-SCN-Bz-DTPA-B72.3 yielded 40% ID/g at 5-7 days in the tumors, while the 88Y-SCN-Bz-EDTA-B72.3 and 88Y-CA-DTPA-B72.3 showed only 6-8% ID/g. The yttrium uptake in the bone with the SCN-Bz-EDTA and CA-DTPA conjugated IgG was over 14 and 11% ID/g, respectively, while 88Y-SCN-Bz-DTPA-B72.3 showed only 3% ID/g. In contrast, the 111In-labeled B72.3 uptake in the bone with all three chelate conjugates was 2-3% ID/g. The differences in yttrium- versus indium-labeled MAb biodistributions demonstrate the difficulty in using 111In-labeled MAbs to predict the biodistribution and dosimetry of 90Y-labeled MAbs unless chelate conjugates such as SCN-Bz-DTPA are used.
Asunto(s)
Anticuerpos Monoclonales/metabolismo , Carcinoma/metabolismo , Neoplasias del Colon/metabolismo , Inmunotoxinas/metabolismo , Radioisótopos de Indio/metabolismo , Ratones Desnudos/metabolismo , Radioisótopos de Itrio/metabolismo , Animales , Anticuerpos Monoclonales/uso terapéutico , Carcinoma/radioterapia , Quelantes/farmacocinética , Quelantes/uso terapéutico , Neoplasias del Colon/radioterapia , Femenino , Humanos , Inmunoglobulina G/metabolismo , Inmunoglobulina G/uso terapéutico , Inmunotoxinas/uso terapéutico , Radioisótopos de Indio/uso terapéutico , Ratones , Trasplante de Neoplasias , Distribución Tisular , Trasplante Heterólogo , Radioisótopos de Itrio/uso terapéuticoRESUMEN
Using excised human skin and tissue grafted to athymic mice, the in vitro and in vivo delivery and metabolism of a salicylate diester were compared. Concentration profiles of this drug and its metabolites were obtained for the outer several hundred microns of the skin. These results show significant differences in the extent of enzymatic cleavage and distribution of metabolites between in vitro and in vivo studies. Furthermore, these data suggest that in vitro results may overestimate metabolism because of increased enzymatic activity and/or decreased capillary removal.
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Administración Cutánea , Ácidos Carboxílicos/administración & dosificación , Ésteres/administración & dosificación , Salicilatos/administración & dosificación , Animales , Ésteres/metabolismo , Ésteres/farmacocinética , Humanos , Técnicas In Vitro , Ratones , Ratones Desnudos/metabolismo , Salicilatos/metabolismo , Salicilatos/farmacocinética , Ácido SalicílicoRESUMEN
Nude mice each attached to a respirator to avoid pulmonary uptake were exposed in a glass exposure chamber to 200, 1000 or 3000 ppm of benzene, toluene or tetrachloroethylene (perclene) for 2, 4 or 6 h. The animals were killed at the end of the study and the amount of each solvent retained in the whole body was determined by gas chromatography. Skin absorption rates were calculated from the amount retained in the whole body using the single compartment model (elimination rate constant) obtained in a previous experiment. There was a linear relationship between the amount of skin absorption and exposure time, and also a linear relationship between the skin adsorption rate and concentration of exposed vapors. Skin absorption of solvent vapors occurs by passive diffusion as defined by Fick's law. The skin absorption coefficient (cm/h) of each solvent vapor was calculated by dividing the skin absorption rate by exposure concentration; the values were 1.24 for toluene, 1.00 for perclene and 0.619 for benzene. The coefficient may be useful for evaluating the amount of skin absorption (ng) was calculated by multiplying the skin absorption coefficient (cm/h), concentration of solvent vapor (ng/cm3), exposure time (h) and exposed skin area (cm2).
Asunto(s)
Benceno/metabolismo , Ratones Desnudos/metabolismo , Absorción Cutánea , Tetracloroetileno/metabolismo , Tolueno/metabolismo , Animales , Cámaras de Exposición Atmosférica , Ratones , Factores de TiempoRESUMEN
Bone turnover in T-cell deficient mice was investigated by comparing parameters of bone physiology in athymic (nude) and euthymic mice. Static and dynamic bone histomorphometry, serum biochemical assays, body weight and tibia length measurements, and bone ash determination were completed in 6- and 12-wk-old athymic (nude) mice (NIH: Swiss nu/nu) and euthymic mice (nu/+) (10 mice/group). In vitro bone resorbing activity stimulated by parathyroid hormone (PTH) or prostaglandin E2 (PGE2) was measured in calvaria of neonatal athymic and euthymic mice. Athymic mice had smaller vertebral tissue area (p less than 0.01), tibia length (p less than 0.001), and less body weight (p less than 0.01) than euthymic mice. The percent double labeled surface (p less than 0.05) and mineralizing perimeter (p less than 0.01) were reduced in athymic as compared to age-matched euthymic mice. Osteoclast number was reduced in the 6-wk athymic mice as compared to 6-wk euthymic mice. Osteoclastic perimeter was reduced in the 12-wk-old mice (athymic and euthymic) as compared to the 6-wk-old mice. Serum calcium was lower at both ages in athymic mice (p less than 0.01) as compared to euthymic mice. Serum alkaline phosphatase levels were reduced (p less than 0.01) in 12-wk-old athymic mice as compared to age-matched euthymic mice, and were greater in 6-wk-old mice than 12-wk-old mice. Athymic mice had greater femur density than euthymic mice (p less than 0.01), and lower (p less than 0.001) percent ash weight of dry bone compared to euthymic mice.(ABSTRACT TRUNCATED AT 250 WORDS)