RESUMEN
In order to study the biological significance of alpha-mannosidase Man2c1, hMan2c1 transgenic mice were developed. In 113 F0 mice, eight were found to be genomic PCR positive for hMan2c1; 9/20 (45%) F1 mice, 16/21 (76.2%) F2 mice, and 12/14 (85.7%) F3 mice were genomic PCR positive for hMan2c1. RT-PCR demonstrated hMan2c1 mRNA transcription in four of eight transgenic lines. Enzymatic activity on p-nitrophenyl-alpha-D: -mannopyranoside was enhanced in 35# and 54# transgenic mice and real-time RT-PCR showed hMan2c1 mRNA expression in these mice. Reduced Con A binding to splenocytes implied N-glycosylation modification of host proteins by hMan2c1 transgene. hMan2c1 transgene promoted growth, invasion, and metastasis to lung of implanted hepatoma H22 and sarcoma S180. The average weights of H22 and S180 tumors were 3.98 +/- 1.62, 3.29 +/- 0.76, 1.69 +/- 1.09, and 3.19 +/- 0.44, 2.72 +/- 1.38, 0.97 +/- 0.41 g for 35#, 54# transgenic mice and wild type mice (W), respectively, (35# or 54# versus W, paired t-test, P < 0.05). In 35# and 54# mice 5/10 and 3/10 showed lung metastasis of H22 tumor in contrast with 1/10 in W mice. In 35# and 54# mice 1/6 and 2/6 showed lung metastasis of S180 tumor in contrast with 0/6 in W mice. The possible mechanism of the promotion was explored on both humoral and cellular immunity. Reduced antibody response to BSA was observed in transgenic mice, suggesting that specific antibody response to tumor antigens might be suppressed by hMan2c1 transgene. However, NK cytotoxicity in splenocytes was not affected by the transgene.
Asunto(s)
Manosidasas/genética , Ratones Transgénicos , Neoplasias/genética , Transgenes/fisiología , Animales , Formación de Anticuerpos/genética , Células COS , Transformación Celular Neoplásica/genética , Chlorocebus aethiops , Citotoxicidad Inmunológica/genética , Progresión de la Enfermedad , Humanos , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/patología , Manosidasas/metabolismo , Ratones , Ratones Endogámicos ICR , Neoplasias/inmunología , Neoplasias/patología , Bazo/metabolismo , Bazo/patología , Células Tumorales Cultivadas , Escape del Tumor/genética , alfa-ManosidasaRESUMEN
Since CD44 was found to be significantly inhibited in the human nasopharyngeal carcinoma cell line CNE-2L2 with profound reduction of malignant activities caused by inhibition of alpha-mannosidase Man2c1 gene expression and CD44 has been observed to be involved in many tumor-supporting functions, we studied the association of CD44 expression with the malignant activities of CNE-2L2 cells. Suppression of CD44 gene expression by RNA silencing technique resulted in profound reduction of malignant potential of the cells, including growth in vitro, colony formation, tumorigenesis and metastasis of tumors in nude mice. Direct injection of the adenoviruses harboring and producing siRNA to CD44 into the tumor inoculated with CNE-2L2 cells in nude mice caused inhibition of tumor growth. The data indicate a positive association of CD44 expression with the malignant activities of CNE-2L2 cells and suggest a possible therapeutic effect of direct introduction of siRNA to CD44 into tumors in some human solid tumors with high expression of CD44 gene.
