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1.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 28-34, 2019.
Article in Chinese | WPRIM | ID: wpr-802229

ABSTRACT

Objective: To investigate the protective mechanism of Liuwei Dihuangwan on diabetes mellitus with liver injury in terms of inflammation. Method: The 18 db/db mice were selected from animal model of spontaneous type 2 diabetes, mice were randomly divided into model group, Liuwei Dihuangwan group(9.75 g·kg-1·d-1,once a day), resveratrol group(42.6 mg·kg-1·d-1,once a day)based on fasting blood-glucose (FBG). The 12 litter wild db/m mice were randomly divided into normal group, Liuwei Dihuangwan group(9.75 g·kg-1·d-1,once a day). The normal group and the model group were given the same amount of distilled water by gavage. The mice in each group were treated for 16 weeks. FBG, triglyceride (TG) and alanine aminotransferase (ALT) were examined. Histopathological changes were observed by liver biopsy hematoxylin-eosin (HE) staining. Silent information regulator 6 (SIRT6),nuclear factor-kappaB p65 (NF-κB p65),monocyte chemoattractant protein-1 (MCP-1), vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in the liver tissue were detected by Western blot. Result: Compared with normal group, FBG, TG and ALT in model group increased significantly(PκB p65,MCP-1,VCAM-1 and ICAM-1 in model group were significantly up-regulated(PPPPκB p65, MCP-1 and VCAM-1 were significantly decreased (PPConclusion: Liuwei Dihuangwan can protect the diabetes mice from liver injury, and its mechanism is related to the improvement of lipid metabolism and the inhibition of inflammatory response.

2.
China Journal of Orthopaedics and Traumatology ; (12): 761-763, 2016.
Article in Chinese | WPRIM | ID: wpr-230402

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of vacuum sealing drainage (VSD) combined with artificial dermis for treatment of the ankle and foot soft tissue defects.</p><p><b>METHODS</b>A total of 15 patients with skin and soft tissue defect of ankle and foot were treated from January 2011 to December 2013, including 10 males and 5 females, with an average age of 32.5 years old ranging from 3 to 55 years old, involving 8 cases by traffic accident, 2 cases by machine accident, 5 cases by crush injury;8 cases with soft tissue exposure, 2 cases with tendor exposure, 5 cases with bone exposure. VSD was used to cover the wounds and continuous negative pressure drainage after debridement, the wounds covered with artificial dermis after the second granulation tissue grew well, again VSD was used to cover the wounds and negative pressure suction was applied, after 7 to 14 days negative pressure closed drainage was removed, free skin graft was transplanted above the artificial dermis, sterile gauze was used to compression bandage.</p><p><b>RESULTS</b>All cases were followed up for 3 to 14 months with an average of 6.5 months. Skin graft of 15 of patients survived after transplantation, artificial dermis graft interval was 7 to 14 days with an average of 9.5 days. There was no obvious scar hyperplasia and contracture, no obvious pigmentation, appearance and functional recovery.</p><p><b>CONCLUSIONS</b>After the implantation of artificial dermis and traditional skin graft method need for 2 to 3 weeks, vacuum sealing drainage technique combined with artificial dermis in treatment of soft tissue defect of foot and ankle skin has advantages of simple operation, significantly shorten the time of the second phase of the skin, without flap to repair, little injury to donor skin area, wound healing quality high, clinical effect of satisfaction.</p>

3.
China Journal of Orthopaedics and Traumatology ; (12): 891-895, 2014.
Article in Chinese | WPRIM | ID: wpr-249261

ABSTRACT

<p><b>OBJECTIVE</b>To evaluate the clinical effects by comparing three different fixation methods: tension band, hollow lag screw and anatomical plate.</p><p><b>METHODS</b>From January 2010 to January 2012, 82 patients with olecranon fractures who underwent surgical treatments were followed-up. All the patients were divided into three groups: tension band fixation group (group A), hollow lag screw fixation group (group B), anatomical plate fixation (group C). In group A, there were 35 patients, including 19 males and 16 females, ranging in age from 32 to 49 years old, with an average of (43.6 ± 8.7) years old, and the patients were treated with tension band fixation. According to Colton classification, there were 5 cases of type I, 3 cases of type II A,19 cases of type II B, and 8 cases of type II C in group A. Among 20 patients in group B, there were 13 males and 7 females, ranging in age from 27 to 50 years old, with an average of (41.5 ± 9.3) years old. The patients in group B were treated with hollow lag screw fixation. According to Colton classification, there were 4 cases of type I, 4 cases of type II A, and 12 cases of type II B in group B. In group C, there were 27 patients totally, including 15 males and 12 females, ranging in age from 30 to 55 years old, with an average of (38.2 ± 6.2) years old. The patients in group C were treated with anatomical plate fixation. According to Colton classification, there were 4 cases of type II B, 13 cases of type II C, and 10 cases of type II D in group C. The Fracture healing time, complications and functional recovery were retrospectively observed and recorded.</p><p><b>RESULTS</b>All the patients were followed up, and the duration ranged from 8 to 24 months, with an average of 15 months. The average healing time of patients in group C was the longest among three groups. The flexion-extension and rotation activities of elbow joint in group B and C were better than that in group C. According to Broberg & Morrey score system, the therapeutic effects of patients in group A and B were better than that of group C. In group C, 2 patients had incision infections, 6 patients complained of foreign body sensation, 1 patient got a delayed fracture healing, and 1 patient had the heterotopic ossification. There were no occurrences of incision infections in group A and B; internal fixation loosening occurred in 3 patients in group A and 2 patients in group B; delayed fracture healing occurred in 2 patients in group A and 2 patients in group B; and skin bursa formation occurred in 6 patients in group A and 1 patient in group B.</p><p><b>CONCLUSION</b>All the three ways are effective methods for the treatment of olecranon fractures. Fixation methods should be selected depending on the type of fracture.</p>


Subject(s)
Adult , Female , Humans , Male , Middle Aged , Bone Plates , Bone Screws , Case-Control Studies , Fracture Fixation, Internal , Methods , Fracture Healing , Olecranon Process , Wounds and Injuries , Ulna Fractures , General Surgery
4.
Chinese Journal of Experimental Ophthalmology ; (12): 581-585, 2012.
Article in Chinese | WPRIM | ID: wpr-635841

ABSTRACT

Background Recently,there were many studies on corneal innervations during mammalian development.However,there were fewer studies on discussing corneal innervations before and after mouse eye openings.Objective The present study was to investigate the change in the regulation of corneal nerve fiber length and density before and after mouse eye openings to offer a basis for clinical research in human.Methods Thirty SPF C57BL/6 mice were divided into postnatal 1 day(P1 d),P7 d,P13 d(1 day before eye opening),P14 d(eye halfopened),P17 d(1 day after eye opening)and P23 d(7 day after eye opening)groups,with 5 mice and 10 eyes for each group.Entire corneal stretches were prepared and immunostaining with an anti-neuron-specific β-Ⅲ tubulin antibody was performed to label the corneal nerve fibers.Confocal microscopic pictures from the corneal dorsal-nasal region (DN),dorsal-temporal(DT),ventral-nasal region(VN)and ventral-temporal(VT)were taken using Delta Vision Core.From these pictures,the mouse corneal area,total length and density of nerve fibers in the 4 regions were calculated.The use of the animals complied with Statement of ARVO.Results Corneal areas of P1 d,P7d,P13 d,P14 d,P17 d and P23 d mice were(0.404±0.007),(1.362±0.154),(1.573±0.080),(1.603±0.046),(1.847±0.052),(2.445±0.798)mm2,respectively ; the total lengths of nerve fibers were(3.718±1.044),(19.065±3.350),(23.687±0.907),(27.309±2.477),(31.989±3.976),(41.214±1.573)mm,respectively ; the densities of nerve fibers were(9.592±1.138),(14.506±1.908),(15.088±1.241),(16.772±1.897),(16.821±2.102),(17.660±1.216)mm/mm2,respectively,all showing significant increases with age(F =22.906,P =0.000 ; F =0.424,P =0.000 ; F =2.375,P=0.000).A positive correlation of the increasing corneal areas and increasing lengths of nerve fibers was found(r=0.983,P<0.01).Nerve fiber densities in the four corneal regions significantly increased with age(DN region:F =0.159,P =0.000 ; DT region:F =2.1 72,P =0.001 ; VN region:F =1.998,P =0.000 ; VT region:F=2.352,P=0.000).From P13 d to P14 d,the corneal nerve fiber densities in the DN region decreased by 6.0% without significant difference(t =0.589,P =0.572); and the corneal nerve fiber densities in the DT region,VN region and VT region decreased by 4.6%,5.5% and 0.1%,respectively,without significant difference from P14 d to P17 d(t=0.549,P=0.596;t=0.701,P=0.501 ;t=-0.100,P=0.919).Conclusions The development of nerve fibers in the whole cornea or the four corneal regions is influenced by eye opening in mouse to various extents.From P13 d to P14 d,the corneal nerve fiber densities in the DN region decreased by 6.0% without significant difference.From P14 d to P17 d,the corneal nerve fiber densities in the DT region,VN region and VT region decrease by 4.6%,5.5% and 0.1%,respectively,without significant difference.Afterwards,the growth of nerve fibers increased in pace and the growth rate is recovered.

5.
Chinese Journal of Plastic Surgery ; (6): 35-39, 2011.
Article in Chinese | WPRIM | ID: wpr-268649

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of Hirudin on random skin flap survival in rats.</p><p><b>METHODS</b>24 SD rats were randomly divided into control group and experimental group. The "McFarlane flap (3 cm x 9 cm)" rat models were established on the rat dorsum. 3 ml Hirudin (30 ATU) was injected into the flap in the experimental group, while 3 ml saline in the control group. The injection was performed for 7 days. The flap survival area in the two groups was measured. The tissue samples were taken from proximal (I), middle (II) and distal (III) portions of flaps for histologic study. The VEGF and bFGF expression was also detected with immunohistochemistry method.</p><p><b>RESULTS</b>7 days after operation, the flap survival rate was (69.52 +/- 3.23)% in the experimental group, while (50.36 +/- 2.37)% in control group, showing a significant difference between the two groups (P < 0. 01). In the middle portion, tissue edema and infiltration of neutrophils in experimental group was markedly slighter than that in control group. The VEGF and bFGF expression and neovascularization was enhanced markedly in experimental group.</p><p><b>CONCLUSIONS</b>Hirudin can increase the survival of random pattern skin flaps. It may increase the VEGF, bFGF expression through a series of complex regulatory pathway. Then flap neovascularization is promoted and the flap blood supply is increased.</p>


Subject(s)
Animals , Male , Rats , Fibroblast Growth Factor 2 , Metabolism , Graft Survival , Hirudins , Pharmacology , Rats, Sprague-Dawley , Skin Transplantation , Surgical Flaps , Vascular Endothelial Growth Factor A , Metabolism
6.
Chinese Journal of Otorhinolaryngology Head and Neck Surgery ; (12): 100-104, 2010.
Article in Chinese | WPRIM | ID: wpr-318287

ABSTRACT

<p><b>OBJECTIVE</b>To introduce the treatment of severe obstructive sleep apnea-hypopnea syndrome (OSAHS) with Repose bone screw system and to evaluate the treating effect.</p><p><b>METHODS</b>Thirty-two patients with severe OSAHS (Friedman classification II, III) were treated with tongue ablation + uvulopalatopharyngoplasty + hard palate shortening + Repose suspension. All patients were followed-up for 6 months. Apnea hypopnea index (AHI), lowest oxygen percent saturation (LSaO₂), body mass index (BMI), vallecula epiglottic-lateral pharyngeal wall (V-LPW) and pharyngeal airway space (PAS) were used for the diagnosis and evaluation.</p><p><b>RESULTS</b>The AHI in 32 patients reduced from (78.3 +/- 11.6)/h to (18.4 +/- 12.5)/h (x(-) +/- s), t = 13.5, P = 0.000. The LSaO₂ increased from (0.632 +/- 0.007) to (0.794 +/- 0.006), t = 4.1, P = 0.000. The BMI was statistical indiscriminate. The V-LPW increased from (12.1 +/- 3.2) mm to (16.9 +/- 2.6) mm (t = 2.5, P = 0.014). PAS increased from (9.2 +/- 3.3) mm to (15.6 +/- 2.4) mm (t = 7.6, P = 0.000). The total effective rate was 93.8%. Three genioglossus stabilization patients complained about foreign body sensation in the mouth floor 3 - 7 days after procedures, which disappeared in 6 months. One genioglossus stabilization patients complained about mandibular osteomyelitis 3 months after procedures, which recovered after anti-inflammatory treatment for 5 days. One genioglossus stabilization patients appeared hypoglossia bleeding, which stopped after compression. One hyoid suspension patients showed haematoma of submandibular region and cured after incision and drainage.</p><p><b>CONCLUSION</b>Repose system was an effective method, safe, simple, with few complications.</p>


Subject(s)
Adult , Female , Humans , Male , Middle Aged , Bone Screws , Sleep Apnea, Obstructive , General Surgery , Tongue , General Surgery
7.
China Journal of Orthopaedics and Traumatology ; (12): 445-447, 2010.
Article in Chinese | WPRIM | ID: wpr-297819

ABSTRACT

<p><b>OBJECTIVE</b>To study the clinical significance of MESS scoring system in the treatment of fractures of lower limb combined with vascular injuries, and to evaluate its reliance.</p><p><b>METHODS</b>From March 2006 to March 2008, 28 patients with fractures of lower limb combined with vascular injuries were graded by MESS scoring system. There were 17 patients were male and 11 patients were female, ranging in age from 23 to 53 years, averaged 38 years. Seventeen patients had fractures at the superior segment of tibia and fibia, 7 patients had fractures at the inferior segment of femur, and other 4 patients had dislocation of knee joint. Among the patients, 18 patients had MESS scores more than 7.0 point, in which 13 patients were treated with one-stage amputation, 5 patients were treated with two-stage amputation; the other 10 patients had the MESS scores less than 7.0 point, and were treated with open reduction and internal fixation, in which 8 patients were treated with transplantation of great saphenous vein to repair blood vessles, and 2 patients were treated with vascular end to end anastomosis.</p><p><b>RESULTS</b>Among the patients, including 18 patients whose MESS scores more than 7.0 point were treated with one-stage or two-stage amputation, and 10 patients whose MESS scores less than 7.0 point were treated with limb salvage operations, all the limbs survived. During the follow-up period (ranged from 0.5 to 1 year, the movement and sensory function of the limbs recovered well.</p><p><b>CONCLUSION</b>MESS is a simple and reliable tool to determine the proper strategy for the patients suffering from vascular injuries with fractures.</p>


Subject(s)
Adult , Female , Humans , Male , Middle Aged , Amputation, Surgical , Blood Vessels , Wounds and Injuries , Fracture Fixation , Methods , Fractures, Bone , General Surgery , Lower Extremity , Wounds and Injuries , Trauma Severity Indices , Vascular Surgical Procedures
8.
Journal of Southern Medical University ; (12): 1517-1520, 2009.
Article in Chinese | WPRIM | ID: wpr-282663

ABSTRACT

<p><b>OBJECTIVE</b>To examine the synergistic effect of recombinant human high mobility group box 1 (HMGB1) protein and lipopolysaccharides (LPS) on the release of interleukin-8 (IL-8) and monocyte chemotactic protein 1 (MCP-1) in human umbilic vein endothelial cells (HUVECs), and explore the role of mitogen-activated protein kinases (MAPK) signal transduction in cytokine release.</p><p><b>METHODS</b>HUVECs were incubated with recombinant HMGB1 (0-75 ng/ml) for 24 h and the culture medium supernatant was harvested for detection of IL-8 and MCP-1 with LiquiChip system. At 0, 1, 3, 6, 12 and 24 h after stimulation with 15 ng/ml HMGB1 or 15 ng/ml HMGB1 plus 10 ng/ml LPS, the levels of IL-8 and MCP-1 in the HUVECs were examined. To test the effect of MAPK inhibitors, HUVCs were pretreated with the inhibitors SB203580 (20 mol/L), PD98059 (20 mol/L), and JNK inhibitor II (50 nmol/L) 1 h before HMGB1 and LPS stimulation.</p><p><b>RESULTS</b>The levels of IL-8 and MCP-1 were significantly increased in the HUVECs stimulated with HMGB1 protein at the concentrations of 3, 15 and 75 ng/ml in comparison with the control levels (P<0.01). Since 3-6 h after the stimulation with HMGB1, the levels of IL-8 and MCP-1 began to increase gradually, and steadily increased at 12 and 24 h, all significantly higher than those of the control group (P<0.01). Stimulation of the HUVECs with LPS (10 ng<ml) or HMGB1 (15 ng/ml) alone resulted in significantly increased levels of IL-8 and MCP-1 (P<0.01), which were further increased after costimulation with LPS and HMGB1, suggesting a synergistic effect between HMGB1 and LPS (P<0.01). This synergistic effect was significantly inhibited by pretreatment with MAPK signaling kinases inhibitors, especially the p38 MAP kinase inhibitor SB203580, and the cocktail of MAP kinase inhibitors almost totally blocked the expression of these chemokines in HUVECs treated with HMGB1 and LPS.</p><p><b>CONCLUSION</b>HMGB1 protein can activate HUVECs to produce the chemokines IL-8 and MCP-1 in a dose- and time-dependent manner. HMGB1 also acts synergistically with LPS to induce IL-8 and MCP-1 release, which might play an important role in the development of sepsis. MAPK signal transduction plays an important role in HMGB1 and LPS-induced IL-8 and MCP-1 release.</p>


Subject(s)
Humans , Cell Line , Chemokine CCL2 , Blood , Metabolism , Dose-Response Relationship, Drug , Endothelial Cells , Metabolism , HMGB1 Protein , Pharmacology , Interleukin-8 , Blood , Metabolism , Mitogen-Activated Protein Kinases , Metabolism , Protein Kinase Inhibitors , Pharmacology , Time Factors
9.
Chinese Journal of Microsurgery ; (6)2006.
Article in Chinese | WPRIM | ID: wpr-676221

ABSTRACT

Objective To observe the expression and distribution the low affinity receptor p75 of nerve growth factor in the traumatic neuroma and investigate its significance and relationship with the growth of peripheral nerve and the formation of the traumatic neuroma.Methods Fifty-one cases of traumatic neuro- ma were collected and were divided into four groups according to its course,i.e,groupⅠ(less than one month),groupⅡ(1~3 months),groupⅢ(3~6 months) and groupⅣ(more than six months).Fifteen cases of normal nerve samples were harvested as the control group(n=15).Immunohistochemical studies were performed to observe the expression of p75.the expression level was detected by the computer graph analysis system.Results There was no significant expression of p75 in the normal nerve while in the four studied groups,significant expression of p75 waspositively observed.The level of p75 was weak in the early time and achieved its peak in three months,but the high level state was maintained even after six months.The one-way ANOVA statistic analysis manifests that there were no significant differences of the expression of p75 among the groupⅡ,ⅢandⅣ(P>0.05),but the differences among the groupⅠand the latter three groups were signifi- cant (P<0.05).p75 was mainly distributed in the Schwans cell.Conclusion p75 is expressed in the traumatic neuroma and may play an important role in the modulation progress of the formation of traumatic neu- roma with an unknown mechanism.

10.
Journal of Southern Medical University ; (12): 1101-1105, 2006.
Article in Chinese | WPRIM | ID: wpr-334986

ABSTRACT

<p><b>OBJECTIVE</b>To study the interaction between Toll-like receptor (TLR) 4 and myeloid differentiation protein-2 (MD-2) in living cells using fluorescence resonance energy transfer (FRET) technology.</p><p><b>METHODS</b>The coding sequences of TLR4 and MD-2 (without the signal peptide sequence) were amplified by PCR and cloned into enhanced cyan fluorescence protein (CFP) and enhanced yellow fluorescence protein (YFP) expression vectors carrying TLR4 signal peptides (pECFP-C1-SP and pEYFP-C1-SP). HEK293 cells were transfected respectively or together with the reconstructed plasmids verified by enzyme digestion and sequence analysis, and the expression and sublocalization of these fluorescence proteins in the cells were observed using fluorescence microscope. FRET in the cells coexpressing CFP-TLR4 and YFP-MD-2 was detected using routine and acceptor photobleaching method.</p><p><b>RESULTS</b>The reconstructed plasmids were expressed in HEK293 cells. The cyan or yellow fluorescence was located in the cytoplasm, mainly around the nucleus in the cells transfected with pECFP/TLR4 or pEYFP/MD-2, and both the cyan and yellow fluorescence located mainly in the membrane and occasional in the cytoplasm of cells cotransfected with pECFP/TLR4 and pEYFP/MD-2. Routine or acceptor photobleaching detected FRET phenomena in cells coexpressing CFP-TLR4 and YFP-MD-2, suggesting direct interaction between TLR4 and MD-2.</p><p><b>CONCLUSION</b>This study provides direct evidence of the interaction between TLR4 and MD-2 in living cells.</p>


Subject(s)
Humans , Cell Line , Fluorescence Resonance Energy Transfer , Methods , Luminescent Proteins , Genetics , Metabolism , Lymphocyte Antigen 96 , Genetics , Metabolism , Plasmids , Genetics , Protein Binding , Recombinant Fusion Proteins , Genetics , Metabolism , Toll-Like Receptor 4 , Genetics , Metabolism , Transfection
11.
Journal of Southern Medical University ; (12): 553-557, 2006.
Article in Chinese | WPRIM | ID: wpr-255256

ABSTRACT

<p><b>OBJECTIVE</b>To construct a p38 MAPK protein delivery system based on TAT protein and study its functions in eukaryotic cells.</p><p><b>METHODS</b>Recombinant vectors pHis-TAT-p38 and pHis-TAT-p38(AF) were constructed, and two recombinant proteins, His-TAT-p38 and His-TAT-p38(AF), were expressed and purified in E. coli. The two fusion proteins were then incubated with ECV304 cells, respectively. The phosphorylation of ATF2 was detected to assay the effect of His-TAT-p38 on endogeneious p38 activity after the cells were stimulated by sorbitol.</p><p><b>RESULTS</b>The results of restriction enzyme digestion and DNA sequencing showed that the two recombinant vectors were correctly constructed. The recombinant proteins of His-TAT-p38 and His-TAT-p38(AF) were isolated and purified by SDS-PAGE, and Western blotting suggested that His-TAT-p38 and its mutant with dual phosphorylation sites could enter the cells efficiently in a time- and concentration-dependent manner. His-TAT-p38 was found capable of increasing the activity of endogenous p38 in ECV304 cells, but His-TAT-p38(AF) inhibited the phosphorylation of ATF2 so as to block the transduction of p38 signal pathway when the cells were stimulated with sorbitol.</p><p><b>CONCLUSION</b>p38 MAPK protein delivery system based on TAT protein has been constructed successfully. It is confirmed that TAT can transfer the proteins into the cells in a time- and dose-depended manner. TAT-p38 and its dominant negative form possess high biological activity after transduction into ECV304 cells by TAT protein delivery system, and the former can increase the activity of endogenous ATF2, but the latter inhibits the transduction of endogeneious p38 signal pathway in ECV304 cells with high osmotic stress.</p>


Subject(s)
Humans , Blotting, Western , Cell Line , Cell Membrane , Metabolism , Eukaryotic Cells , Cell Biology , Metabolism , Gene Products, tat , Genetics , Metabolism , Peptides , Genetics , Metabolism , Phosphorylation , Protein Transport , Recombinant Fusion Proteins , Genetics , Metabolism , Transfection , p38 Mitogen-Activated Protein Kinases , Genetics , Metabolism
12.
Journal of Southern Medical University ; (12): 150-153, 2006.
Article in Chinese | WPRIM | ID: wpr-234174

ABSTRACT

<p><b>OBJECTIVE</b>To assess the value of two-color infrared fluorescence imaging system in detecting protein phosphorylation in comparison with chemiluminescent detection.</p><p><b>METHOD</b>The lung tissue homogenate of mice treated with lipopolysaccharide (LPS) for different time lengths were prepared to separate the proteins by SDS-polyacrylamide gel electrophoresis followed by transfer of the proteins onto PVDF membranes. The membranes were incubated with the antibodies against total p42/44 MAPK/phospho-p42/44 MAPK, and then with goat anti-mouse or anti-rabbit secondary antibodies conjugated to Alexa Fluor 680, IRDye 800 or horseradish peroxidase. The blotted proteins were detected and quantified using Odyssey infrared imaging system or chemiluminescent detection.</p><p><b>RESULTS</b>LPS treatment rapidly induced p42/44 MAPK phosphorylation, which reached the peak level 1 h after the treatment and resumed the baseline level at 12 h. Consistent results were obtained by the two detection methods, but two-color infrared fluorescence imaging system showed better sensitivity in detecting the target protein, and was easy to manipulate with good efficiency and capable of analyzing two proteins simultaneously.</p><p><b>CONCLUSION</b>Two-color infrared fluorescence system is a powerful system for detecting phosphorylation of proteins.</p>


Subject(s)
Animals , Mice , Blotting, Western , Methods , Fluorescence , Fluorescent Dyes , Chemistry , Lipopolysaccharides , Luminescent Measurements , Methods , Lung , Pathology , Mice, Inbred BALB C , Mitogen-Activated Protein Kinase 1 , Chemistry , Metabolism , Mitogen-Activated Protein Kinase 3 , Chemistry , Metabolism , Phosphorylation , Reproducibility of Results , Shock, Septic
13.
Journal of Southern Medical University ; (12): 1394-1407, 2006.
Article in Chinese | WPRIM | ID: wpr-232878

ABSTRACT

<p><b>OBJECTIVE</b>To construct an cell penetrating peptide-based expression vector capable of targeted delivery of proteins into the cell nuclei and study its function of protein transduction.</p><p><b>METHODS</b>The fusion protein expression vector pET14b-HC(L)NE (pET14-b-His-CPP-Linker-NLS-EGFP) incorporating cell penetrating peptide (CPP), nuclear localization signal(NLS), linker and enhanced green fluorescent protein (EGFP) was constructed based on His-tagged pET14b-HE (pET14b-His-EGFP) by site-directed mutagenesis PCR method. After identification by enzyme digestion and DNA sequencing, the recombinant plasmid was transformed into BL21(DE(3)) strain. The HC(L)NE fusion protein was expressed following IPTG induction and purified with Ni(2+)-NTA affinity chromatography. After dialysis and filtration, the HC(L)NE fusion protein was added into cultured eukaryotic cells. The protein transduction in the living cells was observed under fluorescence microscope and analyzed by Western blotting.</p><p><b>RESULTS</b>Enzyme digestion and DNA sequencing confirmed successful construction of the pET14b-HC(L)NE vector, and the fusion protein efficiently expressed in E. coli. Protein transduction experiments in eukaryotic cells revealed that the fusion protein could rapidly penetrate the cell membrane and reach the cell nucleus, and this internalization was time- and concentration-dependent.</p><p><b>CONCLUSION</b>The cell penetrating peptide-based expression vector for targeted protein delivery to the cell nucleus has been successfully constructed, and a transport system that can delivery exogenous proteins or polypeptides into the cytoplasm and cell nucleus is established, which provides an economical and efficient means for functional study of the proteins and polypeptide in cells and targeted drug delivery.</p>


Subject(s)
Animals , Humans , Mice , Blotting, Western , COS Cells , Cell Line , Cell Membrane , Metabolism , Cell Nucleus , Metabolism , Chlorocebus aethiops , Genetic Vectors , Genetics , Green Fluorescent Proteins , Genetics , Metabolism , HeLa Cells , Microscopy, Fluorescence , NIH 3T3 Cells , Nuclear Localization Signals , Genetics , Peptides , Genetics , Metabolism , Protein Transport , Recombinant Fusion Proteins , Genetics , Metabolism , Transfection
14.
Chinese Journal of Oncology ; (12): 621-625, 2004.
Article in Chinese | WPRIM | ID: wpr-254270

ABSTRACT

<p><b>OBJECTIVE</b>To explore high effective and low toxic chemotherapeutic regimens in the treatment of non-small-cell lung cancer (NSCLC).</p><p><b>METHODS</b>A total of 126 patients with advanced NSCLC (Stage III, IV) were randomly divided into two groups: high dose impulsion chemotherapy group (HDIC group) and low dose density chemotherapy group (LDDC group) with 54 patients in HDIC group who received paclitaxel 135-175 mg/m2 on day 1, DDP 80-100 mg/m2 on day 1 and BCNU 125 mg given for brain metastasis on days 1-3 in a 4-6 weeks cycle. Seventy-two patients in LDDC group were given paclitaxel 60-80 mg/m2 on day 1, DDP 40-80 mg/m2 on day 1 repeated weekly and BCNU 125 mg given for brain metastasis with an interval of 2 weeks, in a 4-6 weeks cycle. Antiemetic agent and fluid were administered routinely in HDIC group whereas LDDC group was given antiemetic agent only.</p><p><b>RESULTS</b>Of 157 courses in HDIC group, an average of 2.9 courses per patient, CR 3, PR 23, SD 17 and PD 11 were observed. The effective remission rate was 48.1%, the median effective remission period was 4.5 months and the 1-year survival rate was 46.3%. Of 184 courses in LDDC group, an average of 2.6 courses per patient, CR 9, PR 30, SD 24 and PD 9 were observed. The effective remission rate was 54.2%, the median effective remission period was 6 months and the 1-year survival rate was 56.9%. The effective remission rate and the 1-year survival rate were higher in HDIC group than those in LDDC group, but there was no statistical difference between the two groups (P > 0.05). Severe toxicity was higher in HDIC group than in LDDC group. Two patients in HDIC group died of treatment-related complications (3.7%). Quality of life was better in LDDC group (70.8%) than in HDIC group (51.9%).</p><p><b>CONCLUSION</b>When comparing with high dose impulsion, low dose density regimen of paclitaxel plus cisplatin is more effective and better tolerated with improvement of quality of patients' life in the treatment of NSCLC due to its low dose and short interval duration.</p>


Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Antineoplastic Combined Chemotherapy Protocols , Therapeutic Uses , Brain Neoplasms , Carcinoma, Non-Small-Cell Lung , Drug Therapy , Carmustine , Cisplatin , Dose-Response Relationship, Drug , Drug Administration Schedule , Leukopenia , Lung Neoplasms , Drug Therapy , Pathology , Paclitaxel , Quality of Life , Remission Induction , Survival Rate , Thrombocytopenia
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