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Objective To evaluate the clinical effect of treatment for deep burn wound on finger by proper digital artery flap.Methods From March 2013 to October 2016, 24 patients with deep burn wound on finger were treated by proper digital artery flap.Postoperative observation included wound repair, flap survival, complications and functional recovery of fingers.Results All the 24 flaps survived and no necrosis happened.The marginal abnormal circulation of flap occurred in only 5 cases, which cured by dress changing.All flaps kept well in contour, skin color, temperature and texture.Movement function of donor and recipient fingers was nearly normal.Conclusion Proper digital artery flap avoided the deficiencies distant pedicled flap, so it is a favorite choice for digital soft tissue defect caused by deep burn injury.
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Objective Using M2-Gi1α fusion protein expressed by baculovirus-Sf9 cell system to find the specific ligand for M2 receptor and detect the interaction of the two parts of the fusion protein.Methods The fused M2-Gi1α cDNAs were generated in a two-step PCR and then expressed in Sf9 cells.[3H]QNB and[35S]GTPγS binding experiments were employed to study the function of M2-Gi1α fusion protein.Results The expression level of M2-Gi1α fusion protein was 8.44±0.39 nmol·g-1 protein.The affinity of GDP to the Gi1α part changed under the affection of different ligands.The IC50 value in the appearance of acetylcholine,oxotremorine,arecoline,atropine,fangchinoline,levitimide were 21.35 μmol·L-1,23.86 μmol·L-1,11.91 μmol·L-1,0.13 μmol·L-1,1.05 μmol·L-1,1.75 μmol·L-1,and 2.5 μmol·L-1 when there was no ligand.Conclusion The M2-Gi1α fusion protein expressed in baculovirus-Sf9 cell system has pharmacological specificity for M2 receptor and the efficient coupling function between the two parts.The M2-Gi1αfusion protein is a helpful tool for detecting the new specific ligands of the M2 receptor.
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Aim To generate m3AChR-G11 fusion protein in baculovirus-Sf9 cells and test the couping function,the interation and the influence factors be-tween m3AChR and G11 protein,as well as screen the specific ligands for m3AChR. Methods m3AChR-G11 fused DNA was generated through a two-step PCR and then expressed in Sf9 cells to produce fusion protein. The total concentration for membrane protein was de-tected by BCA method,[3H]QNB and [35 S]GTP?S binding experiment as perfomed to study the function of m3AChR-G11 fusion protein. Results The expression level of m3AChR-G11 was 7. 76 ? 10 -9 mol?g -1. The affinity of GDP to G11 partner changed in the presence of different muscarinic ligands. IC50 values of GDP in the presence of ACh,Pilo,CCh,MCN-A-343,Atro,4-Damp and Dafi were 82. 2,93. 70,12. 10,14. 30, 1. 93,1. 37,0. 72 ? 10 -6 mol ? L -1 respectively,and that in the absence of muscarinic was 1. 99 ? 10 -6 mol ?L-1.Concluslons The m3AChR-G11 fusion protein has the pharmacological specificity of m3 receptor and the efficient coupling interaction of the two partners. Affinity of GDP to ligand-bound fusion protein represents the species of muscarinic ligands. This is helpful in screening and detecting the new specific ligands to muscarinic receptors.
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Aim The m_5AChR-G_ 11? fusion protein was expressed by baculovirus-Sf9 cells system, then using it to identify the specific agonists and antagonists for m_5AChR via detecting the affinity of GDP and m_5AChR-G_ 11?. Methods The m_5AChR-G_ 11? fused cDNAs were generated via a two-step PCR protocol and inserted into pBacPAK9 virus vector. We expressed m_5AChR-G_ 11? fusion protein and m_5AChR protein using baculovirus-Sf9 cell system. [ 3H]QNB and [ 35S]GTP?S binding tests were performed to detect the expressional level of receptor proteins and determine the affinity of GDP and m_5AChR-G_ 11? fusion protein. Results The expression level of m_5AChR-G_ 11? was (47.6?3.2) nmol?g -1 protein. The affinity of GDP to G_ 11? partner changed in the presence of different muscarinic ligands. IC_ 50 values of GDP in the presence of ACh, YM796, Oxotremorine, Methixene, Dextimide and atropine were 128.0, 72.1, 68.5, 16.2, 14.9 and 9.7 ?mol?L -1 respectively, and that in the absence of muscarinic ligand was 20.8 ?mol?L -1. Conclusion The m_5AChR-G_ 11? fusion protein has the pharmacological specificity of M_5 receptor and the efficient coupling interaction of the two partner. Affinity of GDP to ligand bound m_5AChR-G_ 11? fusion protein represents the species of muscarinic ligands. ACh is a full agonist for m_5AChR-G_ 11? fusion protein, YM796 and oxotremorine are partial agonists, while methixene, dextimide and atropine are antagonists.
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AIM: To prepare m 1AChR-G 11 and m 4AChR-G 16 fusion protein in Baculovirus-Sf9 cell system and detect the effects of various muscarinic ligands on the interaction between m 1AChR and G 11 and m 4AChR and G 16 ,and screen different kinds of ligands specific for m 1 and m 4. METHODS: To prepare fused DNA of m 1AChR-G 11 and m 4AChR-G 16 in two PCR, then expressed in Sf9 cells and detect the pharmacological function of m 1AChR-G 11 fusion protein and m 4AChR-G 16 fusion protein by [ 3H]QNB and [ 35 S]GTP?S binding experiments; To expore the way of the activation of m 1AChR-G 11 and m 4AChR-G 16 fusion protein by various ligands includingcetylcholine (ACh), Pilocarpine (Pilo), 4-hydroxy-2-butynyl-1-trimethylammonium-m-chloro-carbanilatechloride (McN-A-343),tetrandrine, pirenzepine (PZ), alcuronium, atropine,R-(+)-hyoscyamine and gallamine by displacement by GDP on [ 35 S]GTP?S binding experiments. RESULTS: The expression levels of m 1AChR-G 11 and m 4AChR-G 16 fusion protein were (45 39?2 62) nmol?g -1 protein, (47 04?1 58) nmol?g -1 protein. The affinity of GDP to G 11 and G 16 partner changed in the presence of different muscarinic ligands. CONCLUSION: The m 1AChR-G 11 and m 4AChR-G 16 showed the pharmacological specificity to m 1 and m 4 receptor and the efficient signaling of the two partners. Ligands of m 1AChR and m 4AchR mediated different signal transduction by changing the affinity of G 11 /G 16 and GDP. So m1AChR-G 11 fusion protein and m 4AChR-G 16 fusion protein can be taken as a tool to screen ligands specific for m 1AChR and m 4AChR.