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Objective @# To investigate the application of indocyanine green in near-infrared fluorescence imaging to determine the scope of necrotic bone resection in osteoradionecrosis of the jaw and to provide a reference for clinicians@*Methods @#Eight patients with osteoradionecrosis of the jaws were enrolled. Indocyanine green was intravenously injected through the elbow vein 10 minutes before osteotomy. After conservative resection of necrotic bone lesions based on imaging results, the scope of potential dead bone resection in the area of low fluorescence intensity was gradually expanded at an initial distance of 0.3 cm. Near-infrared fluorescence imaging and fluorescence intensity determination of bone cross-section were performed before and after extended resection. Statistical differences were analyzed. All patients with osteonecrosis underwent regular follow-up to evaluate the postoperative efficacy@*Results@#Indocyanine green was injected into all 8 patients with osteoradionecrosis for near-infrared fluorescence imaging and the scans were clear; the fluorescence intensity of fresh bone wounds with an expanded mandibular resection range of (0.95 ± 0.14) cm was (226.2 ± 15.8) au, which was higher than that based on intraoperative macroscopic observation and radiological results (108.8 ± 3.4) au, (t = 20.718, P<0.001). The postoperative follow-up improvement rate of 8 patients was 87.5%.@* Conclusion @#Near-infrared fluorescence imaging with indocyanine green can assist in the successful removal of necrotic bone until fresh bleeding of the jaw wound occurs, which has important clinical value in defining the resection range of osteoradionecrosis of the jaw.
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【Objective】 To explore the influence of anti-HLA-Ⅰ with different mean fluorescence intensity (MFI) on the efficacy of HLA-A and -B gene matching platelet transfusion, so as to provide scientific data for clinical platelet gene matching transfusion strategy. 【Methods】 A total of 81 PTR patients had applied for HLA-Ⅰgene matched platelets from the platelet gene database established by our laboratory, and 28 (MFI <5 000) of them needed further avoiding of partial donor-specific antibodies and they were enrolled as the research subjects. According to the platelet MFI value of HLA-Ⅰ antibody-targeting antigen, they were divided into negative transfusion group (MFI <500) (group A) and positive transfusion groups (MFI≥500) ; the latter were further divided into group B (500≤MFI <1 000), group C (1 000≤MFI <3 000) and group D (MFI≥3 000) according to MFI value. Corrected count increment (CCI) in platelet count was used to compare the platelet transfusion effect in 4 groups. 【Results】 Among 28 platelet recipients with MFI <5 000, 19(67.86%) patients successfully received 72 effective transfusions. The first CCI (×109/L) in groups A, B, C and D were 10.27±7.46, 7.58±4.75 (P>0.05), 17.36±7.63 (P>0.05) and -0.77±2.30 (P<0.05), respectively. There was no statistical difference among group A, B and C. 【Conclusion】 The application of HLA-Ⅰ gene matching platelets in PTR patients can adjust the MFI threshold(<2 000) appropriately according to the patient′s condition without compromising the platelet transfusion effect.
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Objective To study the effects of nano silver (Ag) and titanium dioxide (TiO2) on the content of nucleic acid in staphylococcus aureus in order to explore their antibacterial mechanisms.Methods After preparation of beef extract peptone liquid cultures,the effects of minimal inhibitory concentrations (MICs) of nano Ag and TiO2 on staphylococcus aureus strains were determined.With the 1/2 MICs nano Ag and TiO2,the contents of DNA and RNA macromolecules from staphylococcus aureus cultures were measured to determine the damage degree of staphylococcus aureus cell membranes by ultraviolet spectrophotometer,and then the fluorescence intensities of the staphylococcus aureus cells were observed under fluorescence microscope and the fluorescence values were tested by fluorescence spectrophotometer to determine the contents of nucleic acid DNA and RNA.Results The MICs of nano Ag and TiO2 were 1.6 mg/mL and 5.781 μg/mL.After treatment with the 1/2 MICs nano Ag and TiO2,nano Ag group and TiO2 group were compared with the control group (culture fluid without adding antibacterial agent),respectively,and there were no significant differences in the contents of DNA and RNA macromolecules from staphylococcus aureus cultures between n anoAg group and control group as well as between TiO2 group and control group were (P＞0.05),and there were significant decreases in fluorescence intensities and the contents of nucleic acid DNA and RNA (P＜0.01).Conclusions Nano Ag and TiO2 had obvious antibacterial effects on staphylococcus aureus and the antibacterial properties of nano Ag was stronger than that of TiO2.The antibacterial mechanisms of nano Ag and TiO2 against staphylococcus aureus may be associated with the inhibition of the synthesis of nucleic acid DNA and RNA,inhibiting protein synthesis and then bacterial growth.
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Objective To investigate the role of miR-10a in CD4+CD25+Treg-mediated immunosuppression during sepsis and its potential role in immunotherapy for sepsis.Methods Sepsis mouse model was established by cecal ligation and puncture(CLP).Balb/c mice of clean grade were sacrificed 1,3,5,and 7 days after operation.Blood as well as spleen samples were harvested at given intervals.The splenic CD4+CD25+Treg cells and CD4+T cells were isolated by MACS microbeads.Cells were cultured,and phenotypes were analyzed by flow cytometry.The miR-10a expressed in Treg cells were detected by Real-time PCR.After administration of LV-mmu-miR-10a-5p-inhibition,the immunosuppressive function have been detected.Statistical analyses were performed using one-way analysis of variance (SPSS 19.0,Chicago,USA) test followed by Dunnett-t test to compare among three or more groups or by Student's t-test to compare between two groups.Results The percentages of splenic Tregs (CD4+CD25+/CD4+T) was (7.34±1.2)％ in normal group,and the increase in percentage of Tregs in spleen has been observed in septic mice (P＜0.05).The mean fluorescence intensity (MFI) of Foxp3+Treg was increased in septic mice compared with sham group (P＜0.05).The expression of miR-10a was significantly elevated on CLP 1-7 day (P＜0.05).After down-regulation of miR-10a in septic mice,the percentages of Tregs (CD4+CD25+/CD4+T) was significantly increased in septic mice (P＜0.05),the MFI of Foxp3+Treg was increased in septic mice compared with control group (P＜0.05).The CD4+T cell proliferative activity in CLP-induced mice was significantly suppressed on CLP 3 day compared with sham group (P＜0.05).After down-regulation of miR-10a in septic mice,the CD4+T cell proliferative activity was significantly suppressed compared with control group (P＜0.05).Conclusions Treg plays a critical role in immunosuppression in septic mice.Inhibition of miR-10a in vivo could enhence immunesuppression of CD4+CD25+Treg.Therefore miR-10a may participate in the regulation of CD4+CD25+Treg immunosuppression in sepsis and become the target for immunotherapy.
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OBJECTIVE To develop a method to evaluate the compatibility of compounds in the fluorescence resonance energy transfer (FRET) model for β-secretase (BACE1) inhibitor screening.METHODS Two commercially available BACE1 inhibitor screening systems based on FRET were selected to evaluate the BACE1 inhibitory activities of (-)-epigallocatechin-3-gallate (EGCG) and Compound 1 according to the supplier's protocol.The inhibitory rates and slopes of the catalytic curves of the inhibitors were calculated.The effect of inhibitors on the fluorescence intensity of the systems were quantitatively calculated and the comparatively evaluated.RESULTS EGCG,a reported non-competitive inhibitor of BACE1,directly induced the reduction of fluorescence intensity of one of the systems.The slope of the line with the addition of EGCG (10.8±2.6) conformed to that of the line of EGCG inhibition (10.2±3.4),which indicated that EGCG was a pseudo-positive inhibitor of BACE1.Compound 1 had little effect on the fluorescence intensity of the systems,so the inhibitory activity of Compound 1 was confirmed.The compounds which showed inhibitory activity in preliminary screening should be checked in the blank control without BACE1 to calibrate the effect of compound on the system fluorescence intensity.The applicability of the tested compounds in the screening system could thus be evaluated to prevent pseudo-positive results.CONCLUSION This fluorescence calibration method with compound control can be universally used for assays based on FRET theory to evaluate the applicability of tested BACE1 inhibitors.
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Objective:To improve the immobilization efficiency of antibody molecules on immune microarray,the method of es-tablishment and optimization of agarose self-assembled membrane carrier with three-dimensional hydrogel structure was established. Methods: The agarose self-assembled membrane carrier was prepared by using glass slide as the carrier,using agarose and sodium periodate modification on glass surface. The agarose self-assembled membrane carrier was characterized by TEM, AFM and FTIR. The optimum preparation conditions were obtained. The carrier for two different species of fixed source antibody efficiency were studied. Antibody loading capacity of agarose self-assembled membrane carrier and ordinary aldehyde carrier were investigated and compared by fluorescence microscopy imaging and Image J software. Results: The agarose nano-membrane carrier had uniform and compact surface. This structure could increase the specific surface area and improve the probe fixed rate. The optimal concentration of agarose for preparation of carrier was 1. 0% . When the concentration of IgG was 0. 3-0. 4 mg/ml,the oxidized self-assembled chitosan film substrate had highest antibody loading capacity. And it had a 3. 94 fold higher antibody loading capacity than the ordinary aldehyde carrier. Conclusion: The agarose nano-membrane carrier is an ideal method for surface modification of immobilized antibody molecules, which is more suitable for preparation of immune microarray carrier.
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The tumor targeted fluorescent magnetic IR780-Fe3 O4 nanoparticles were prepared for separation and detection of circulating tumor cells ( CTCs ) . These IR780-Fe3 O4 nanoparticles were characterized by electron microscopy, fluorescence spectrometer, and superconducting quantum interferometer. The targeting effect of IR780-Fe3 O4 nanoparticles was analyzed on the tumor and normal cells by confocal microscope and flow cytometry, and the confocal microscope was used to target the location of IR780-Fe3 O4 nanoparticles in MCF-7 cells. The standard curve was drawn and evaluated accorded to the IR780-Fe3 O4 nanoparticles fluorescence intensity of tumor cells after incubation. The results showed that IR780-Fe3 O4 nanoparticles could target a variety of CTCs. Furthermore, cellular localization experiment proved that IR780-Fe3 O4 nanoparticles could target the mitochondria of tumor cells. With the method of coupling magnetic Fe3 O4 nanoparticles, IR780 could well distinguish the tumor and normal cells, which could be used for separating and detecting the CTCs in simulated blood.
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The tumor targeted fluorescent magnetic IR780-Fe3 O4 nanoparticles were prepared for separation and detection of circulating tumor cells ( CTCs ) . These IR780-Fe3 O4 nanoparticles were characterized by electron microscopy, fluorescence spectrometer, and superconducting quantum interferometer. The targeting effect of IR780-Fe3 O4 nanoparticles was analyzed on the tumor and normal cells by confocal microscope and flow cytometry, and the confocal microscope was used to target the location of IR780-Fe3 O4 nanoparticles in MCF-7 cells. The standard curve was drawn and evaluated accorded to the IR780-Fe3 O4 nanoparticles fluorescence intensity of tumor cells after incubation. The results showed that IR780-Fe3 O4 nanoparticles could target a variety of CTCs. Furthermore, cellular localization experiment proved that IR780-Fe3 O4 nanoparticles could target the mitochondria of tumor cells. With the method of coupling magnetic Fe3 O4 nanoparticles, IR780 could well distinguish the tumor and normal cells, which could be used for separating and detecting the CTCs in simulated blood.
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OBJECTIVE: To study the endocytosis of albumin nanoparticles by L02 cells. METHODS: Calcein human serum album nanoparticles (calcein HSA-NPs) were prepared by a desolvation technique with calcein as a fluorescent probe. The particle size distribution, in vitro release, cell uptake, intracellular drug distribution, and cytotoxicity were determined. RESULTS: The prepared calcein HSA-NPs with an average diameters of 203.3 nm showed sustained release profile in vitro. The L02 cell uptake of nanoparticles showed time, concentration, and temperature dependency. HSA-NPs could significantly improve the transmembrane transport capacity of calcein as a water-soluble small molecule (P < 0.01). Clathrin-mediated endocytosis and macropinocytosis were the main routes by which calcein HSA-NPs entered into cells. After entering into cells, nanoparticles were located in the intracellular space outside the nucleus, mainly in the lysosomes. HSA-NPs had no cytotoxicity basically. CONCLUSION: Calcein HSA-NPs can be used as an effective vehicle to study the mechanism of endocytosis. Copyright 2012 by the Chinese Pharmaceutical Association.
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ObjectiveTo study the feasibility of analyzing photosensitizers absorbing process by cells based on image processing technology.MethodsFluorescence images of Laryngeal cells were taken by inverted fluorescence microscope after adding photosensitizers for a certain time.Parameter L that reflects the fluorescence intensity of cells in different times was obtained and the calculation results were compared.ResultsThe fluorescence intensity of cells increased over time.Sobel operater and Otsu algorithm can both reflect the fluorescence intensity of images.ConclusionImage processing technology can effectively analyzes the process of photosensitizers absorbed by cells.
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Objective To obtain the near-infrared fluorescence image in vivo and find the relationship between the detecting depth and fluorescence probe concentration. Methods Signals of fluorescence probe in various depths of vivo and tissue phantom with cooled CCD camera were acquired. Results The fluorescence intensity informations with different fluorescence probe concentrations and depths in vivo and liquid phantom were obtained. Conclusion Relationship between fluorescence intensity,location and depth of detecting probe in vivo is found. The linear relation of fluorescence probe concentration and detecting intensity is simulated,which will be used as a reference for the experiment system.
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Aim To observe the effects of theaflavin on intracellular calcium concentration(i) level in rat ventricular myocytes and discuss the possible mechanisms.Methods The effects of theaflavin oni were investigated in rat ventricular myocytes.i was detected by laser confocal microscopy and represented by relative fluorescent intensity(FI-FI0)/FI0,%;FI0:control;FI:administration of drugs).Results ① Theaflavin(10,20,40 ?mol?L-1) had no effect on the i of ventricular myocytes in normal Tyrode′s solution.However,it reduced the i of ventricular myocytes in simulated ischemia solution in a concentration-dependent manner.② Pretreatment with Bay k8644(0.5 ?mol?L-1) mostly abolished the effects of theaflavin(20 ?mol?L-1) in simulated ischemia solution.③ Theaflavin(20 ?mol?L-1) markedly inhibited the low concentration of ryanodine-induced i increase in Ca2+-free Tyrode′s solution.④ When the propagating waves of elevated i(Ca2+ waves) were produced by increasing extracellular Ca2+ concentration from 1 mmol?L-1to 10 mmol?L-1,theaflavin(20 ?mol?L-1) could block the propagating waves of elevated i(Ca2+ waves),reduce the frequency and duration of propagating waves,and reduce i as well.Conclusion Theaflavin may reduce the i in isolated rat ventricular myocytes via inhibiting Ca2+ influx by voltage-dependent Ca2+ channel and alleviating Ca2+ release from sarcoplasmic reticulum(SR).