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OBJECTIVE:To optimize the extraction technology of Qingjie atomized liquid. METHODS:The extraction technol-ogy of Qingjie atomized liquid was optimized by orthogonal test with hyperoside content as index,using the amount of added wa-ter,extraction time and extraction times as factors,and the verification test was conducted. RESULTS:The optimal extraction tech-nology was as follows as 12-folds water,extracting for 2 times and 1 hour each time. The content of hyperoside was 4.129 μg/g in 3 validation tests(RSD=1.81%,n=3). CONCLUSIONS:The optimal extraction technology is stable and practical,and can pro-vide reference for the formulation of technology and quality standard for Qingjie atomized liquid.
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Objective To develop an RP-HPLC method for the content determination of heseridin in Hezhong Lipi Pills.Methods The Agilent HC-C18 column (250 mm×4.6 mm, 5μm) was used with methanol-0.1% phosphoric acid (35∶65) as the mobile phase at the flow rate of 1.0 mL/min. The detection wavelength was 283 nm, and the column temperarute was 40℃.Results The linear range of hesperidin was 0.15-3.64μg,r=1.000 (n=6). The average recovery was 98.97%, RSD=0.90% (n=6).Conclusion The method is accurate, simple, sensitive and with good repeatability, which can be applied to the quality control ofHezhong Lipi Pills.
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Objective To detect the effects of glucocorticoids on the expression of toll-like receptor 4 (TLR4) on CD14+ monocytes in peripheral blood from patients with chronic obstructive pulmonary disease (COPD) and explore the relationship between inherent immunity deficiency and the pathogenesis of COPD.Methods A total of 30 hospitalized acute exacerbations of COPD (AECOPD) patients from 2012 January to March were divided them into 2 groups of mild-moderate (A,n =15) and severe-critical (B,n =15) according to pulmonary functions.The same basic treatment (anti-infection,oxygen inhalation,bronchial smooth muscle relaxing and cough relief) was administered.And glucocorticoids were added to Group B.Two weeks later,AECOPD achieved a clinical remission and it was designated as a stable COPD group.Another 30 healthy cases over the same period were selected as control group.Flow cytometry was used to measure the relative mean fluorescence intensity (rmfi) and relative positive cell percent (rpcp) of TLR4 onr CD14 positive peripheral blood monocytes.Conventional pulmonary function test (FEV1 percentage of predicted value,FEV1/FVC value) was performed.Results The value of rmfi for TLR4 of AECOPD group was 5.14 ±0.26 and rpcp (4.24 ±0.17)％ and they were similar to those of COPD group [4.46 ±0.24 and (3.49 ± 0.24) ％ respectively,all P ＞ 0.05].AECOPD and COPD groups were both significantly lower than normal control group [13.75 ±0.28 and (10.25 ±0.13)％ respectively,all P ＜0.01].The rmfi of TLR4 of Group B was 3.12 ±0.15 and rpcp (3.17 ±0.10)％.And both were significantly lower than those of Group A [5.48 ±0.23 and (5.12 ±0.20)％ respectively,all P ＜0.01].The value of rmfi for TLR4 of Group B before glucocorticoid treatment was 3.12 ±0.15 and rpcp (3.17 ±0.10)％.And both were significantly lower than those of later glucocorticoid treatment [5.24 ± 0.25 and (5.44 ± 0.19) ％ respectively,all P ＜ 0.01].Both values were significantly lower than those of normal control group [13.75 ± 0.28 and (10.25 ± 0.13) ％ respectively,all P ＜ 0.01].The expression of TLR4 on CD14 + monocytes in peripheral blood of AECOPD patients was positively correlated with FEV1 percentage of predicted value(r1 =0.824,P＜0.001) and FEV1/FVC (r2 =0.756,P＜0.001).Conclusions Innate immunodeficiency plays an important role in the pathogenesis of COPD.And the expression of TLR4 is an important indicator for disease severity.Glucocorticoids treat COPD through inhibiting airway inflammation response and enhancing defensive function of innate immune system.
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Objective:To clone the fused gene of hVEGF_(165) and hirudin,construct a eukaryotic expression vector hVEGF_(165)-FH/pcDNA3.0 and express the vector in human endothelial cell strain. Methods: hVEGF_(165) cDNA fragment was amplified by PCR and fused hirudin peptide was synthesized by primer annealing. The fused hVEGF_(165 ) and hirudin gene was cloned into vector pcDNA3.0 to construct a new vector hVEGF_(165)-FH/pcDNA3.0 and the recombinant plasmid was confirmed by restriction enzyme and auto-sequencing. The correctly cloned plasmids were subjected to rapid in vitro transcription and translation identification. Liposome mediated, recombinant plasmids were used to transfect human endothelial cell strain and the products were identified. Results: hVEGF_(165)-FH was successfully fused and formed a 711 bp target fragment. The recombinant vector hVEGF_(165)-FH/pcDNA3.0 was successfully transformed into human endothelial cell strain which expressed the fusion protein. Conclusion: The recombinant vector hVEGF_(165)-FH/pcDNA3.0 can express fused protein in human endothelial cell strain, which paves a way for further study on the activity of the fusion protein and gene therapy.
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Objective: To investigate the effect of human vascular endothelial growth factor on restenosis after angioplasty. Methods: A rabbit model of injured carotid artery was established using percutaneous transluminal angioplasty. The pcDNA3/hVEGF165（500 μg,n=12） and pcDNA3 （500 μg,n=12） were separately transfected into injured arterial wall with 30 min incubation. The carotid artery was imaged by arotic angiography at the end of week 2 and week 4. Pathology analysis and Northern blot analysis were performed for harvested injured artery segment. Results: Arotic angiography showed carotid artery diameter narrowness were obviously lessened at week 2 and week 4 in experimental group than that in control group; H-E stains showed lumina narrow ratio were obviously reduced at week 2 and week 4 in experimental group than that in control group［（9.58±1.35）% vs (31.72±1.72)%；(18.09±2.93)% vs (44.05±3.28)%， P<0.01 ］; By Northern blot analysis, the expression of hVEGF165mRNA in experimental group were upregulated than in contol group. Conclusion: pcDNA3/hVEGF165 can be transfected into smooth muscle cell and continue to secret bioactivity protein at least for 4 weeks; it can accelerate reendothelialization and prevent restenosis.
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AIM: To examine the inhibition of matrix metalloproteinase (MMP) activity by doxycycline (Doxy) and its effect on vascular smooth muscle cells (SMCs) proliferation, neointimal hyperplasia and vascular remodeling. METHODS: The model of rat common carotid artery injury was established by balloon-dilatation. Doxy was administered to the animals of treatment group at dose of 30 mg?kg -1?d -1. The activity of MMPs in the tissue of injured carotid arteries was measured by gelatin zymography. The thickness and area of neointimal, lumen area and the proliferation of SMCs were measured by histological and morphometric analysis. RESULTS: 1. After Doxy treatment, the activity of MMP-9 in the carotid arteries was reduced by 26.3% and 34.5% compared to that in rats without Doxy treatment at 24 hours and 3 days after balloon injury, respectively (P