A useful property of optical imaging is the potential to modulate the detectable signal to improve target/nontarget ratios. When administered as a dimer of a fluorophore- and a quencher-conjugated duplex arranged to inhibit fluorescence but designed to dissociate only in the presence of its target, the fluorescence signal should in principle appear only in the target. This laboratory has demonstrated the feasibility of this approach by using a duplex consisting of a linear oligomer conjugated with Cy5.5 (emitter) hybridized to another linear oligomer conjugated with Iowa Black (quencher) in a pretargeting optical study. Now eight duplexes consisting of combinations of 18 mer linear phosphodiester (PO) and phosphorothioate (PS) DNAs and phosphorodiamidate morpholinos (MORFs) conjugated with Cy5.5 (emitter) and Iowa Black (quencher) were variously screened for in vitro duplex stability. The MORF/PO duplex was selected for further study based on evidence of stability in 37 degrees C serum. Simultaneously, the kinetics of quenching were investigated in vitro and in vivo in mice. Thereafter, mice were implanted in one thigh with MORF/PO Cy 5.5 microspheres and the complementary PS Iowa Black administered iv to measure the extent and kinetics of duplex formation in the target. While all duplexes were stable in buffer, only the MORF/PO duplexes and possibly all PS containing duplexes were stable in 37 degrees C serum for at least 4 h. The kinetics of quenching were found to be rapid in vitro, with a 80-90% decrease in Cy5.5 fluorescence immediately following formation of a PS/PS homoduplex, and in vivo, with a 27 to 38% decrease in target thigh/nontarget ratio within 1 h following administration of the complementary PS Iowa Black complementary DNA but not the random control DNA to mice implanted with MORF/PO Cy5.5 microspheres. This investigation has provided additional evidence that Cy5.5 may be efficiently and rapidly quenched by Iowa Black when both are conjugated to complementary oligomers and that the resulting inhibition of fluorescence is sufficiently persistent for imaging. 相似文献
Aroma serves as one of the decisive factors influencing the value of banana commodities. Most of characteristic volatile organic components (VOCs) are formed during post-harvesting. However, the changing of VOCs of banana at different post-harvesting stages remain ambiguous. In this study, the VOCs of Cavendish banana for the four typical post-harvesting stages (green stage/half of yellow stage/yellow ripening stage/over ripening stage) are clarified using headspace solid phase micro-extraction (HS-SPME), combined with gas chromatography-mass spectrometry (GC–MS). The results inferred that the relative content of branched-chain esters such as acetate and butyrate, which form the main contributors of aroma in bananas, is higher in the T2 and T3 stages. Further, RNA-Seq technology was employed to clarify the formation mechanism of banana aroma in the post-harvesting stage. The MaTGL4 gene of the linoleic acid metabolism pathway and the MaBCAT3 and MaBCAT5 genes of the valine, leucine and isoleucine degradation pathway in banana suggest the expression is active late in the ripening stage, and the upregulated expression of these genes is analogous to the formation of aroma components such as branched-chain esters and hexenal. The above results not only provide baseline data on the differences in physical and chemical properties of VOCs in various post-harvesting stages of banana production, but also provide theoretical guidance facilitating the subsequent improvement of the commercial value of bananas through genetic improvement.
We investigated ATP-binding cassette transporters A1/G1 expression and function in mediating cholesterol efflux by examining the macrophages of cigarette-smoking patients with coronary artery disease (CAD) before and after smoking abstinence. Peripheral blood monocyte cells were collected from nonsmokers (n = 17), non-CAD (NCAD) smokers (n = 35), and CAD smokers (n = 32) before and after 3 months of smoking cessation. We found that the ABCA1 expression level was lower in macrophages from NCAD and CAD smokers than from nonsmokers at baseline. The ABCA1 function of mediating cholesterol efflux was reduced in NCAD and CAD smokers as compared with nonsmokers. After 3 months of smoking cessation, ABCA1 expression and function were improved in CAD smokers. However, ABCG1 expression and function did not change after smoking cessation. Furthermore, ABCA1 expression was inhibited by tar in human acute monocytic leukemia cell line THP-1-derived macrophages through the inhibition of liver X receptors. Nicotine and carbon monoxide did not inhibit ABCA1 expression. Our results indicate that chronic cigarette smoking impaired ABCA1-mediated cholesterol efflux in macrophages and that tobacco abstinence reversed the function and expression of ABCA1, especially in CAD patients. It was tobacco tar, rather than nicotine or carbon monoxide, that played a major role in the tobacco-induced disturbance of cellular cholesterol homeostasis. 相似文献
Claudins play an important role in tumor metastasis and in invasiveness of colorectal cancer (CRC). We have evaluated the relationship between CRC and expression of the claudin genes in Chinese patients with CRC. We measured CLDN1 and CLDN7 mRNA using quantitative PCR, and protein levels with immunohistochemistry in cancer tissues and adjacent normal tissue. Cancer tissues had significantly higher levels of CLDN1, and significantly lower levels of CLDN3, CLDN4, and CLDN7 than did normal tissue. CLDN3, CLDN4, and CLDN7 expression levels were higher in CRC of the protruded type than in CRC of the infiltrative type. Expression of CLDN7 correlated with lymph node metastasis. Stage N0 cancer tissues had higher levels of CLDN7 than did stages N1 and N2, suggesting that CLDN7 expression was closely related to the extent of lymph node metastasis. CLDN1 protein was upregulated, but CLDN7 protein was downregulated in cancer tissues when compared with expression in adjacent normal tissues. In conclusion, CLDN3, CLDN4, and CLDN7 were significantly downregulated, whereas CLDN1 was significantly upregulated in CRC. The altered expression of claudin genes may play a role in the initiation and development of CRC. 相似文献
The general strategy and rationale underlying the design of COPD therapeutics that possess protease inhibitory activity and are also capable of releasing a species that attenuates inflammation by inhibiting caspase-1, are described. The synthesis and in vitro biochemical evaluation of a dual function molecule that sequentially inhibits HNE and caspase-1 in a time-dependent manner is reported. 相似文献