A novel matrix metalloproteinase (MMP) was identified from leukocytes and found to be specifically expressedby peripheral blood leukocytes among 29 different tissuesexamined. Named leukolysin, it encodes for 562 resid...A novel matrix metalloproteinase (MMP) was identified from leukocytes and found to be specifically expressedby peripheral blood leukocytes among 29 different tissuesexamined. Named leukolysin, it encodes for 562 residueswith a conserved MMP structure, i.e., pre-, pro-, catalytic , hinge- and hemopexin-like domains, but also a RXK/RRmotif, known for its role in MMP zymogen activation, anda C-terminal hydrophobic segment. Overall, leukolysindisplays the strongest homology to the newly identifiedMT-MMP subgroup with 45% and 39% identities to MT4and MT1-MMPs vs 30% and 31.5% to MMP1 and 3 respectively. Unlike MT4-MMP whose proteolytic activityremains undefined, a C-terminally truncated leukolysin isexpressed as a strong gelatinolytic species at 28 kDa whichis derived from a cell-associated 34 kDa proenzyme, presumably by furin or proprotein convertase mediated removal of the propeptide (-6 kDa). By green fluorescentprotein (GFP) tagging, the intracellular proenzyme is localized to granules throughout the cell, suggesting thatactivation occur immediately prior to secretion. Taken together, leukolysin may be part of the proteolytic arsenaldeployed by leukocytes during inflammatory responses.展开更多
Objective： To investigate the specific blockage effect of individual antisense RNA on mutant p53 gene in vitro. Methods： The single strand antisense transcription system containing mt-p53 exon 8 sequence （pGEM3zf（...Objective： To investigate the specific blockage effect of individual antisense RNA on mutant p53 gene in vitro. Methods： The single strand antisense transcription system containing mt-p53 exon 8 sequence （pGEM3zf（＋/-）p53exon8） was constructed. The ligation of antisense RNAwith mt-p53 gene was confirmed by in situ hybridization; MDA-MB-231 human breast cancer cells were transfected with ASp53exon8＇RNA cotionic liposome-mediated. Expression of mt-p53 protein was examined by immunocytochemical staining and Western blot. Cell proliferation was evaluated by MTT assay; Cell cycle distribution was determined by flow cytometry （FCM）; Apoptosis was observed by TUNEL. Results： In transfected MDA-MB-231 cells, hybridization signals were observed in cytoplasm. ASp53exon8＇RNA transfection induced inhibition of cell proliferation, G2/M phase arrest and increasing apoptotic rates. In addition, expression of p53 protein was down-regulated. Conclusion： pGEM3zf（＋/-）p53exon8 was well constructed and ASp53exon8＇RNA can block mt-p53 gene expression specifically and then inhibit MDA-MB-231 cell proliferation in vitro, which may serve as therapeutic means for human malignancy.展开更多
Baicalein had been proved to have anti-cancer activity in vitro and in vivo, including the inhibition of malignant proliferation, migration, adhesion and invasion of many kinds of cancer cells. The special AT-rich seq...Baicalein had been proved to have anti-cancer activity in vitro and in vivo, including the inhibition of malignant proliferation, migration, adhesion and invasion of many kinds of cancer cells. The special AT-rich sequence binding protein 1 （SATB1） is a tissue-specific expression of nuclear matrix-binding protein and is reported to be a breast cancer ＂gene group organizer＂. Previous studies have shown that SATB1 is involved in the growth, metastasis and prognosis of breast cancer. The present study was aimed to investigate whether baicalein inhibits the proliferation and migration of MDA-MB-231 human breast cancer cells through down-regulation of the SATB1 expression. Methods： MDA-MB-231 cells were treated for 24 h, 48 h and 72 h with various concentrations of baicalein （0, 5, 10, 20, 40 and 80 pM） respectively. Then, the proliferation and migration of MDA-MB-231 cells following treatment with baicalein were determined using colorimetric 3-（4, 5-dimethylthia- zol-2-yl） 2, 5-diphenyltetrazolium bromide （MTT） and wound healing assays. Thereafter, western blot analysis was performed to detect the changes of SATB1 protein expression in MDA-MB-231 cells. Results： Along with the prolongation of time and increase of drug concentration, inhibitory effect of baicalein on proliferation and migration of MDA-MB-231 cells gradually in- creased, in a time.- and dose- dependent manner （P 〈 0.05）. Meanwhile, after treated with baicalein in different concentrations for 48 h, the level of SATB1 protein expression of MDA-MB-231 cells decreased obviously, in a dose-dependent manner （P 〈 0.05）. Conclusion： Baicalein inhibits breast cancer cell proliferation and suppresses its invasion and metastasis by reducing cell migration possibly by down-regulation of the SATB1 protein expression, indicating that baicalein is a potential therapeutic agent for human breast cancer.展开更多
Mutation of mevalonate kinase （MVK） is thought to account for most cases of hyperimmunoglobulinemia D syndrome （HIDS） with recurrent fever. However, its mechanism and the relationship between elevated serum immuno...Mutation of mevalonate kinase （MVK） is thought to account for most cases of hyperimmunoglobulinemia D syndrome （HIDS） with recurrent fever. However, its mechanism and the relationship between elevated serum immunoglobulin D （IgD） and the clinical features of HIDS are unclear. In this study, we generated by fusion PCR a vector to express high levels of chimeric secretory IgD （cslgD） specifically in the liver. We then generated seven founder lines of transgenic mice by co-microinjection, and verified them using genomic PCR and Southern blotting. We detected the expression of csIgD by reverse transcription PCR, quantitative PCR, western blotting, and enzyme-linked immunosorbent assays. We demonstrated that csIgD could be specifically and stably expressed in the liver. We used flow cytometry to show that overexpression of csIgD in the bone marrow and spleen cells had no effect on B cell development. Morphologic and anatomical observation of the transgenic mice revealed skin damage, hepatosplenomegaly, and nephromegaly in some transgenic mice; in these mice, pathological sections showed high levels of cell necrosis and protein-like sediments in the liver, spleen, and kidney. We demonstrated that the genomic insertion sites of the transgeues did not disrupt the MVK gene on mouse chromosome 5. This transgenic mouse will be useful to explore the pathogenesis of HIDS.展开更多
文摘A novel matrix metalloproteinase (MMP) was identified from leukocytes and found to be specifically expressedby peripheral blood leukocytes among 29 different tissuesexamined. Named leukolysin, it encodes for 562 residueswith a conserved MMP structure, i.e., pre-, pro-, catalytic , hinge- and hemopexin-like domains, but also a RXK/RRmotif, known for its role in MMP zymogen activation, anda C-terminal hydrophobic segment. Overall, leukolysindisplays the strongest homology to the newly identifiedMT-MMP subgroup with 45% and 39% identities to MT4and MT1-MMPs vs 30% and 31.5% to MMP1 and 3 respectively. Unlike MT4-MMP whose proteolytic activityremains undefined, a C-terminally truncated leukolysin isexpressed as a strong gelatinolytic species at 28 kDa whichis derived from a cell-associated 34 kDa proenzyme, presumably by furin or proprotein convertase mediated removal of the propeptide (-6 kDa). By green fluorescentprotein (GFP) tagging, the intracellular proenzyme is localized to granules throughout the cell, suggesting thatactivation occur immediately prior to secretion. Taken together, leukolysin may be part of the proteolytic arsenaldeployed by leukocytes during inflammatory responses.
基金Key Project Fund of Basic Research, Tianjin Municipal Science and Technology Commission (No. 033801511), and the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry (2004–2005).
文摘Objective： To investigate the specific blockage effect of individual antisense RNA on mutant p53 gene in vitro. Methods： The single strand antisense transcription system containing mt-p53 exon 8 sequence （pGEM3zf（＋/-）p53exon8） was constructed. The ligation of antisense RNAwith mt-p53 gene was confirmed by in situ hybridization; MDA-MB-231 human breast cancer cells were transfected with ASp53exon8＇RNA cotionic liposome-mediated. Expression of mt-p53 protein was examined by immunocytochemical staining and Western blot. Cell proliferation was evaluated by MTT assay; Cell cycle distribution was determined by flow cytometry （FCM）; Apoptosis was observed by TUNEL. Results： In transfected MDA-MB-231 cells, hybridization signals were observed in cytoplasm. ASp53exon8＇RNA transfection induced inhibition of cell proliferation, G2/M phase arrest and increasing apoptotic rates. In addition, expression of p53 protein was down-regulated. Conclusion： pGEM3zf（＋/-）p53exon8 was well constructed and ASp53exon8＇RNA can block mt-p53 gene expression specifically and then inhibit MDA-MB-231 cell proliferation in vitro, which may serve as therapeutic means for human malignancy.
基金Supported by grants from the National Natural Science Foundation of China(No.81274136)Xi’an Jiaotong University’s Cross Project Funds(No.Xjj2012141)the Talent Funds of the Second Affiliated Hospitalof Xi’an Jiaotong University(No.RCCGG201105)
文摘Baicalein had been proved to have anti-cancer activity in vitro and in vivo, including the inhibition of malignant proliferation, migration, adhesion and invasion of many kinds of cancer cells. The special AT-rich sequence binding protein 1 （SATB1） is a tissue-specific expression of nuclear matrix-binding protein and is reported to be a breast cancer ＂gene group organizer＂. Previous studies have shown that SATB1 is involved in the growth, metastasis and prognosis of breast cancer. The present study was aimed to investigate whether baicalein inhibits the proliferation and migration of MDA-MB-231 human breast cancer cells through down-regulation of the SATB1 expression. Methods： MDA-MB-231 cells were treated for 24 h, 48 h and 72 h with various concentrations of baicalein （0, 5, 10, 20, 40 and 80 pM） respectively. Then, the proliferation and migration of MDA-MB-231 cells following treatment with baicalein were determined using colorimetric 3-（4, 5-dimethylthia- zol-2-yl） 2, 5-diphenyltetrazolium bromide （MTT） and wound healing assays. Thereafter, western blot analysis was performed to detect the changes of SATB1 protein expression in MDA-MB-231 cells. Results： Along with the prolongation of time and increase of drug concentration, inhibitory effect of baicalein on proliferation and migration of MDA-MB-231 cells gradually in- creased, in a time.- and dose- dependent manner （P 〈 0.05）. Meanwhile, after treated with baicalein in different concentrations for 48 h, the level of SATB1 protein expression of MDA-MB-231 cells decreased obviously, in a dose-dependent manner （P 〈 0.05）. Conclusion： Baicalein inhibits breast cancer cell proliferation and suppresses its invasion and metastasis by reducing cell migration possibly by down-regulation of the SATB1 protein expression, indicating that baicalein is a potential therapeutic agent for human breast cancer.
基金supported by the National Basic Research Program of China(Grant No.2010CB945300)
文摘Mutation of mevalonate kinase （MVK） is thought to account for most cases of hyperimmunoglobulinemia D syndrome （HIDS） with recurrent fever. However, its mechanism and the relationship between elevated serum immunoglobulin D （IgD） and the clinical features of HIDS are unclear. In this study, we generated by fusion PCR a vector to express high levels of chimeric secretory IgD （cslgD） specifically in the liver. We then generated seven founder lines of transgenic mice by co-microinjection, and verified them using genomic PCR and Southern blotting. We detected the expression of csIgD by reverse transcription PCR, quantitative PCR, western blotting, and enzyme-linked immunosorbent assays. We demonstrated that csIgD could be specifically and stably expressed in the liver. We used flow cytometry to show that overexpression of csIgD in the bone marrow and spleen cells had no effect on B cell development. Morphologic and anatomical observation of the transgenic mice revealed skin damage, hepatosplenomegaly, and nephromegaly in some transgenic mice; in these mice, pathological sections showed high levels of cell necrosis and protein-like sediments in the liver, spleen, and kidney. We demonstrated that the genomic insertion sites of the transgeues did not disrupt the MVK gene on mouse chromosome 5. This transgenic mouse will be useful to explore the pathogenesis of HIDS.
