ABSTRACT
cloud point extraction, hydrophilic interaction chromatography, anti-AIDS drugs, serum@#【Objective】 Cloud point extraction （CPE） combined with hydrophilic interaction （HILIC） was employed to simultaneously determine the anti-AIDS drugs in the serum of rats. 【Methods】Triton X-114 was used as extraction medium to extract four nucleoside antiviral drugs from rat serum. Response surface methodology was employed to further optimize CPE parameters. The content of four anti-AIDS drugs in rat serum was simultaneously determined by HILIC method.【Results】 The optimized conditions were as follows：5%（w/v）Triton X- 114，0.3 mol/L NaCl，PH 5.0， the water- bath equilibrium 20 min at 40 ℃ . Under optimized extraction conditions，the extraction rates of the four anti- AIDS drugs were all over 85.0%，which was pretty close to the predicted result. The extracted samples were analyzed under the optimal chromatographic conditions. The recovery rate was over 75.0% and RSD was less than 5.0%. The optimized chromatographic conditions were as follows：Ze month- CN（250 × 4.6 mm，5 micron）as stationary phase，mobile phase （methanol：acetonitrile：ammonium acetate buffer）（5∶5∶90），column temperature 35 ℃，detection wavelength 275 nm，flow rate 0.5 mL/min.【Conclusion】The developed method of CPE combined with HILIC can enrich four anti-AIDS drugs in rat serum ，which is simple，environment- friendly and can be used to accurately determine the blood concentration of four anti-AIDS drugs in rat serum，providing a new method for clinical blood concentration monitoring.
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression and the role of secreted frizzled-related protein 2 (SFRP2) in the earlobe keloid and find a valid way to treat the keloid with gene therapy.</p><p><b>METHODS</b>The expression of SFRP2 mRNA and protein was tested with in situ hybridization and Western Blot Analysis method in the different period of earlobe keloid.</p><p><b>RESULTS</b>The SFRP2 mRNA and protein expression at the keloid edge was significantly high in 12 month group than in 3 or 6 month groups (P < 0.01), but not than in 24 month group. The SFRP2 expression started to decrease in the keloid center 12 month later (P < 0.01). The SFRP2 expression was always higher in edge than in center during all the period (P < 0.05, P < 0.01).</p><p><b>CONCLUSIONS</b>The results suggest that SFRP2 may play an important role in the development of keloid, especially at the keloid edge. The high SFRP2 expression in endothelial cells and surrounding tissue is also important. It may be a new way for gene therapy of keloid by decreasing the SFRP2 expression.</p>
Subject(s)
Adolescent , Adult , Female , Humans , Male , Middle Aged , Young Adult , Ear , Keloid , Metabolism , Membrane Proteins , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To verify the interaction between secreted frizzled-related protein 2 (SFRP2) and osteoblast-specific factor 2 (OSF-2).</p><p><b>METHODS</b>HA-tagged OSF-2 fusion protein recombinant vector pCMV-HA-OSF-2, which could express in mammal cells was constructed, then identified by enzyme-cutting and transfected into human kidney 293 (HK293) cells with or without Myc-SFRP2 recombinant eukaryotic expression vector pCMV-HA-SFRP2. The interaction between SFRP2 and OSF-2 was detected through coimmunoprecipitation and Western blotting.</p><p><b>RESULTS</b>In electrophoresis bath, target fragment of SFRP2 coding gene with 800 bp and target gene OSF-2 with 2500 bp could be seen respectively after enzyme-cutting, which showed that pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 were constructed successfully. No HA-OSF-2 expression was detected after pCMV-Myc-SFRP2 or pCMV-HA-OSF-2 transfection. Whereas, HA-OSF-2 expressed by Myc antibody immunoprecipitation after pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 co-transfection.</p><p><b>CONCLUSIONS</b>HA-OSF-2 recombinant vector can express in mammal cells. Interaction exists between HA-OSF-2 and SFRP2.</p>
Subject(s)
Humans , Cell Line , Core Binding Factor Alpha 1 Subunit , Genetics , Metabolism , Genetic Vectors , Keloid , Metabolism , Membrane Proteins , Genetics , Metabolism , Recombinant Fusion Proteins , Metabolism , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To investigate the influence of substance P (SP) on the release of histamine in the human hypertrophic scar tissue, and to explore the prerequisite of their interaction.</p><p><b>METHODS</b>Tissue specimens of normal skin and hypertrophic scar from eight hospitalized patients were excised and cut into 0.5 to 1.0 mm3 pieces, and the histamine release by mast cell (MC) under the stimulation of different concentration of SP and calcium, as well as the different affect time of SP, were determined with fluorescence spectrometer. Then the histamine release rate was calculated.</p><p><b>RESULTS</b>There was obvious release of histamine when SP concentration was 1 x 10(-6) mol/L , and the release rate was (50.0 +/- 3.6) %, which was significantly higher than that by SP in the concentration of 0 mol/L [(44.0 +/- 3.2) %, P < 0.01]. Therefore it seemed to be dose-dependent. About 90% of histamine was released within 15 minutes of 5 x 10(-1) mol/L Substance P stimulation, and it was also time-dependent. The histamine release reached the peak when calcium concentration was 5 x 10(-3) mol/L, which seemed to be dose-dependent, but it decreased transiently when calcium concentration was 1 x 10(-3) mol/L. In all occasions, the influence of SP on the histamine release by MC in hypertrophic scar (HS) was markedly higher than that in normal skin (NS) (P < 0.01). Conclusion The influence of SP on the histamine release by MC in HS was markedly higher than that in NS, and it might be closely related to itching sensation and the formation of hypertrophic scar.</p>
Subject(s)
Child , Child, Preschool , Female , Humans , Male , Cicatrix, Hypertrophic , Metabolism , Histamine , Metabolism , Skin , Metabolism , Substance P , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To establish an ideal model of human hyperplastic scar (HS) in nude mice, so as to provide us a new model to carry out further studies on the mechanism of HS development.</p><p><b>METHODS</b>Full skin defect sized 2.0 cm x 1.5 cm was created on the back of 100 nude mice. The defect was thereafter covered with full thickness human skin. After the grafted skin survived, the nude mice were subjected to deep partial thickness burn of the grafted skin with heated copper rod. The development of the hyperplasia of the scar after wound healing was observed histologically and grossly.</p><p><b>RESULTS</b>Grafted full-thickness human skin took and survived well in 86 out of 100 nude mice. There was obvious and continuous hyperplasia of scar in 67 mice (78%). The external appearance and histological features of the HS appeared similar to those in human HS. The average thickness of the scar was 0.34 cm, with the thickest part measuring 0.6 cm. In addition, the time of hyperplastic change lasted for 63 - 217 days in average of 128 days.</p><p><b>CONCLUSION</b>Obvious and continuous scar hyperplasia could be found in this model, and the whole process beginning from wound healing to the formation of HS could be easily observed. The model was therefore suitable and ideal for the study of HS.</p>