RESUMEN
An improved PCR-based subtractive hybridization strategy was used to clone apoptosis-related genes induced by all-trans retinoic acid (ATRA) from human promyelocytic leukemia cell line HL-60 cells. The protocol used the cap-finder method, long-distance PCR, streptavidin magnetic bead-mediated subtraction and spin column chromatography. Twenty-seven clones related to apoptosis were identified by reverse dot blot assay. Seventeen were known genes, of which seven have been reported to be apoptosis related. The remaining 10 were unknown genes, five of which were sequenced and named apr-1 to apr-5. apr-1, apr-2, apr-3 and TNF were reidentified by reverse dot blot, and it is suggested that they might be related to apoptosis. The results suggest that this strategy might be efficient for large-scale cloning of differentially expressed genes in target cells.
Asunto(s)
Apoptosis/genética , Clonación Molecular/métodos , Perfilación de la Expresión Génica/métodos , Regulación Leucémica de la Expresión Génica , Separación Inmunomagnética , Proteínas de Neoplasias/genética , Reacción en Cadena de la Polimerasa/métodos , Técnica de Sustracción , Apoptosis/efectos de los fármacos , Biotinilación , Células Clonales/química , ADN Complementario/genética , Reacciones Falso Positivas , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Células HL-60/química , Células HL-60/efectos de los fármacos , Humanos , Immunoblotting , Masculino , Estreptavidina , Tretinoina/farmacologíaRESUMEN
Giant-cell tumor of bone seems to occur more frequently in Chinese people than in those residing in Western countries. The estimated incidence is about 20 per cent of all primary tumors of bone. Of 208 surgically treated and pathologically proved giant-cell tumors, 194 were benign. We excluded patients with primary or secondary amputation unrelated to recurrence and those followed for less than two years or lost to follow-up. Of the remaining 111 patients who were followed for more than two years, twenty-nine had a recurrence, giving a recurrence rate of 26.1 per cent. The rate of recurrence was highest following curettage and bone-grafting (41.2 per cent) and was much lower in patients who were treated by resection and fusion (7.1 per cent). Since resection of this tumor with reconstructive procedures, either by massive homogenous bone-grafting or artificial joint replacement, is complicated and might cripple the patient if it fails, we propose excision and curettage with bone-grafting as the most suitable method of treatment in the majority of patients with giant-cell tumor of bone.
Asunto(s)
Neoplasias Óseas/cirugía , Tumores de Células Gigantes/cirugía , Adolescente , Adulto , Amputación Quirúrgica , Neoplasias Óseas/diagnóstico por imagen , Neoplasias Óseas/epidemiología , Trasplante Óseo , Niño , China , Legrado , Femenino , Tumores de Células Gigantes/diagnóstico por imagen , Tumores de Células Gigantes/epidemiología , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia , RadiografíaRESUMEN
AIM: To establish the role of vascular endothelial growth factor (VEGF) in the oncogenesis of human gastric carcinoma more directly. METHODS: The expression of VEGF and its receptor kinase-domain insert containing receptor (KDR) in human gastric cancer tissue were observed by immunohistochemical staining. VEGF levels were manipulated in human gastric cancer cell using eukaryotic expression constructs designed to express the complete VEGF(165) complimentary DNA in either the sense or antisense orientation. The biological changes of the cells were observed in which VEGF was up-regulated or down-regulated. RESULTS: VEGF-positive rate was 50%, and VEGF was mainly localized in the cytoplasm and membrane of the tumor cells, while KDR was mainly located in the membrane of vascular endothelial cells in gastric cancer tissues and peri-cancerous tissue. In 2 cases of 50 specimens, the gastric cancer cells expressed KDR, localized in both the cytoplasm and membrane. Introduction of VEGF(165) antisense into human gastric cancer cells (SGC-7901, immunofluorescence intensity, 31.6%)) resulted in a significant reduction in VEGF-specific messenger RNA and total and cell surface VEGF protein (immunofluorescence intensity, 8.9%) (P<0.05). Conversely, stable integration of VEGF(165) in the sense orientation resulted in an increase in cellular and cell surface VEGF (immunofluorescence intensity, 75.4%) (P<0.05). Lowered VEGF levels were associated with a marked decrease in the growth of nude mouse xenografted tumor (at 33 days postimplantation, tumor volume: 345.40 +/- 136.31 mm3)(P<0.05 vs control SGC-7901 group: 1534.40 +/- 362.88 mm3), whereas up-regulation of VEGF resulted in increased xenografted tumor size (at 33 days postimplantation, tumor volume: 2350.50 +/- 637.70 mm3) (P<0.05 vs control SGC-7901 group). CONCLUSION: This study provides direct evidence that VEGF plays an important role in the oncogenesis of human gastric cancer.