OBJECTIVE To study the effect of antisense oligonucleotides (ASODN) of survivin, a member of a gene family of inhibitors of apoptosis, on the ability of cisplatin (DDP) to induce apoptosis in osteosarcoma cells.METHOD...OBJECTIVE To study the effect of antisense oligonucleotides (ASODN) of survivin, a member of a gene family of inhibitors of apoptosis, on the ability of cisplatin (DDP) to induce apoptosis in osteosarcoma cells.METHODS ASODN of survivin were synthesized and transfected into osteosarcoma OS-732 cells. The effects of ASODN alone, and with DDP as well as the effect of sense oligonucleotides (SODN) with and without DDP were examined. The reverse tanscriptase-polymerase chain reaction (RTPCR) was utilized to detect the expression of survivin mRNA in each group of OS-732 cells. The cells were examined by flow cytometry (FCM) and staining with acridine orange (AO) to determine the morphology of the cells and the level of apoptosis in each group. The MTT assay was used to estimate the condition of cell growth.RESULTS In comparing the cells transfected with ASODN with the control, SODN and DDP groups, we found that the expression of survivin mRNA was significantly decreased (P<0.01), and the level of apoptosis enhanced. ASODN-treated cells appeared atrophic with condensed chromatin, characteristic of typical apoptotic changes. Cell growth was relatively inhibited in the ASODN groups. Furthermore, the apoptotic index (AI) and cell growth-inhibition ratio (IR) were significantly higher in the ASODN+DDP group compared to each group treated with a single agent and compared to the SODN+DDP group.CONCLUSION ASODN can specifically inhibit the expression of the survivin gene in OS-732 cells, and correspondingly suppress survivin function, resulting in an increase in sensitivity to DDP and thus the ability of DDP to induce apoptosis in osteosarcoma cells.展开更多
: For the sake of providing some important information relevant to the study of the molecular mechanism of genic male sterility in plants, gene differential expression in flower buds at different developmental stages,...: For the sake of providing some important information relevant to the study of the molecular mechanism of genic male sterility in plants, gene differential expression in flower buds at different developmental stages, as well as in rosette leaves, florescence leaves, and scapes was analyzed using cDNA amplified fragment length polymorphism (cDNA-AFLP) in the genic male sterile A and fertile B line of Chinese cabbage pak-choi. Following amplification of 125 pairs of primer combinations, 11 differential fragments were obtained, of which eight were from the B line and the other three were from the A line. Of 11 differential fragments, four were verified by Northern hybridization that were expressed preferentially in fertile flower buds. Results of GenBank BLAST showed that one fragment was with unknown function, whereas the other fragments have strong nucleotide sequence similarities with the polygalacturonase (PG) gene, the pectinesterase (PE) gene, and the polygalacturonase inhibitory protein (PGIP4) gene. Only full-length cDNA from the differential fragment BcMF-A18T16-1 was amplified by rapid amplification of cDNA ends (RACE) and Northern analysis showed that this fragment was expressed only in medium and large-sized flower buds of the B line. The full-length cDNA, designated as BcMF2 (Brassica campestris Male Fertile 2), was 1 485 bp long and was composed of a 1 263-bp open reading frame, which had 83% nucleotide similarity to a PG gene from Arabidopsis encoding polygalacturonase. Analysis of the basic structure of the protein revealed that it had one polygalacturonase active site (RVTCGPGHGLSVGS) at 256th site of amino acids and was classified as being a member of family 28 of the glycosyl hydrolases. The role of the BcMF2 gene on microspore development is discussed in the present paper.展开更多
文摘OBJECTIVE To study the effect of antisense oligonucleotides (ASODN) of survivin, a member of a gene family of inhibitors of apoptosis, on the ability of cisplatin (DDP) to induce apoptosis in osteosarcoma cells.METHODS ASODN of survivin were synthesized and transfected into osteosarcoma OS-732 cells. The effects of ASODN alone, and with DDP as well as the effect of sense oligonucleotides (SODN) with and without DDP were examined. The reverse tanscriptase-polymerase chain reaction (RTPCR) was utilized to detect the expression of survivin mRNA in each group of OS-732 cells. The cells were examined by flow cytometry (FCM) and staining with acridine orange (AO) to determine the morphology of the cells and the level of apoptosis in each group. The MTT assay was used to estimate the condition of cell growth.RESULTS In comparing the cells transfected with ASODN with the control, SODN and DDP groups, we found that the expression of survivin mRNA was significantly decreased (P<0.01), and the level of apoptosis enhanced. ASODN-treated cells appeared atrophic with condensed chromatin, characteristic of typical apoptotic changes. Cell growth was relatively inhibited in the ASODN groups. Furthermore, the apoptotic index (AI) and cell growth-inhibition ratio (IR) were significantly higher in the ASODN+DDP group compared to each group treated with a single agent and compared to the SODN+DDP group.CONCLUSION ASODN can specifically inhibit the expression of the survivin gene in OS-732 cells, and correspondingly suppress survivin function, resulting in an increase in sensitivity to DDP and thus the ability of DDP to induce apoptosis in osteosarcoma cells.
基金国家自然科学基金，the Key Sci-Technology Project of Zhejiang Province,China
文摘: For the sake of providing some important information relevant to the study of the molecular mechanism of genic male sterility in plants, gene differential expression in flower buds at different developmental stages, as well as in rosette leaves, florescence leaves, and scapes was analyzed using cDNA amplified fragment length polymorphism (cDNA-AFLP) in the genic male sterile A and fertile B line of Chinese cabbage pak-choi. Following amplification of 125 pairs of primer combinations, 11 differential fragments were obtained, of which eight were from the B line and the other three were from the A line. Of 11 differential fragments, four were verified by Northern hybridization that were expressed preferentially in fertile flower buds. Results of GenBank BLAST showed that one fragment was with unknown function, whereas the other fragments have strong nucleotide sequence similarities with the polygalacturonase (PG) gene, the pectinesterase (PE) gene, and the polygalacturonase inhibitory protein (PGIP4) gene. Only full-length cDNA from the differential fragment BcMF-A18T16-1 was amplified by rapid amplification of cDNA ends (RACE) and Northern analysis showed that this fragment was expressed only in medium and large-sized flower buds of the B line. The full-length cDNA, designated as BcMF2 (Brassica campestris Male Fertile 2), was 1 485 bp long and was composed of a 1 263-bp open reading frame, which had 83% nucleotide similarity to a PG gene from Arabidopsis encoding polygalacturonase. Analysis of the basic structure of the protein revealed that it had one polygalacturonase active site (RVTCGPGHGLSVGS) at 256th site of amino acids and was classified as being a member of family 28 of the glycosyl hydrolases. The role of the BcMF2 gene on microspore development is discussed in the present paper.
