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Objective:To establish high performance liquid chromatography （HPLC） fingerprint and multi-component determination for the substance benchmark of Yiweitang， and to evaluate its quality in combination with chemical pattern recognition method. Method:Fifteen batches of substance benchmark of Yiweitang were prepared， the "Chinese medicine chromatographic fingerprint similarity evaluation system" （2012 edition） was used to calculate similarity. Cluster analysis， principal component analysis and orthogonal partial least squares-discriminant analysis were employed to handle the common peaks for evaluating the quality difference among 15 batches of the substance benchmark. The contents of catalpol， verbascoside and methylophiopogonanone A were determined with mobile phase system of acetonitrile-phosphoric acid solution at detection wavelengths of 210 nm and 334 nm. Result:There were 22 common peaks in HPLC fingerprint of the substance benchmark， among them， peaks 1， 9， 12， 14-17， 19 and 20 belonged to Rehmanniae Radix， peaks 3， 4， 6， 7 and 21 belonged to Glehniae Radix， peaks 5 and 22 belonged to Ophiopogonis Radix， peaks 2 and 18 belonged to Polygonati Odorati Rhiaoma， peak 8 was the common peak of Ophiopogonis Radix and Rehmanniae Radix， peak 10 was shared by Ophiopogonis Radix， Polygonati Odorati Rhiaoma<italic> </italic>and<italic> </italic>Rehmanniae Radix， peak 11 was the common peak of these four herbs， and peak 13 was shared by Polygonati Odorati Rhiaoma and Rehmanniae Radix. The similarities between HPLC fingerprints of 15 batches of the substance benchmark and the control fingerprint were all >0.90， the samples could be divided into four categories by three chemical pattern recognition methods. Quantitative analysis showed that the contents of catalpol， verbascoside and methylophiopogonanone A among 15 batches of samples ranged from 0.37% to 1.14%， 0.002% to 0.054% and 0.016% to 0.079%， respectively. Conclusion:The established fingerprint and determination for the substance benchmark of Yiweitang have good separation and high accuracy， which reflect the overall chemical composition characteristics of Yiweitang， and can provide experimental basis for the further development and quality control of the compound preparations of this famous classical formula.
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@#AIM: To analyze the efficacy of bevel-up phaco chop versus bevel-down phaco drill in the treatment of hard nuclear cataract and the influence on corneal endothelium.<p>METHODS: Between August 2018 and April 2020, 94 patients(104 eyes)with hard(grade Ⅳ-Ⅴ)nuclear cataract treated with phacoemulsification were enrolled in this retrospective study. Among them, 50 eyes treated by bevel-up phaco chop were included in the control group, while 54 eyes treated by phaco drill technique were included in the observation group. The time of using ultrasound, ultrasonic energy and accumulated energy complex parameter in the two groups were recorded. The best corrected visual acuity(BCVA)in 7d, corneal endothelial cell count in 2mo, the proportions of normal hexagonal cells and the degree of corneal edema in 7d were compared between the two groups.<p>RESULTS: The time of using ultrasound, ultrasonic energy and accumulated energy complex parameter of the observation group were shorter and lower than those of the control group(<i>P</i><0.05). The BCVA of both groups increased after operation, and the observation group had higher BCVA than the control group at the same time(<i>P</i><0.05). In 1 and 2mo, the corneal endothelial cell counts and the proportions of normal hexagonal cells in both groups were lower than those before operation. Meanwhile, the corneal endothelial cell count and the proportions of normal hexagonal cells in the observation group were higher than those in the control group(<i>P</i><0.05). Besides, corneal edema was milder in the observation group than in the control group at 24h and 7d after operation(<i>P</i><0.05).<p>CONCLUSION: Compared with bevel-up phaco chop, phaco drill can significantly shorten the time of using ultrasound, reduce ultrasound energy, promote visual recovery, and reduce corneal endothelial cell damage.
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AIM:To explore the effect of comprehensive therapy in the treatment of fungal corneal ulcer and clinical outcome. METHODS:A total of 159 cases 159 eyes fungal corneal ulcer patients took comprehensive therapy of 10 measures, followed up more than 3mo to observe the treatment effect and the final visual acuity in our hospital for treatment. RESULTS: The average hospitalization time of fungal corneal ulcer was(12. 11±5. 13) d. A total of 148 cases of 148 eyes were cured, and the cure rate was 93. 1%;improvement in 8 cases, accounted for 5. 0%; 3 cases were ineffective, accounted for 1. 9%. The final visual acuity of 89 eyes ≥0. 3, accounted for 56. 0%(89/159). The final visual acuity of 136 eyes ≥0. 1, accounting for 85. 5%(136/159), and accounting for 91. 9% in 148 cured patients. CONCLUSION: The comprehensive therapy in the treatment of fungal corneal ulcer has higher cure rate and visual acuity.
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To construct the adenoviral vector with co-expressing keratinocyte growth factor (KGF) and enhanced green fluorescent protein (EGFP) for transfection into the mesenchymal stem cells (MSC), the target gene KGF was cloned into the shuttle plasmid with the report gene EGFP, then the recombinant shuttle plasmid was transformed into DH5a bacteria to recombine with backbone vector pAdxsi. Next, the plasmid pAd-EGFP-mKGF was amplified in H293 cells and the viral titer was determined. The MSC were separated and enriched by using bone marrow adherent culture and identified in vitro to observe the efficiency of transfection. The results indicated that the recombinant shuttle plasmid pShuttle-EGFP-mKGF digested with restriction endonucleases was confirmed by two products which length was about 0.6 kb and 5.1 kb, respectively; the recombinant plasmid pAdxsi-EGFP-mKGF digested with restriction endonucleases was confirmed by 7 products; recombinant adenoviral vector Ad-EGFP-mKGF was amplified to titer of 1.6 × 10(10) pfu/ml. At 10 h after transfecting MSC began to express fluorescence at 6 to 8 days later, the fluorescence reached to the peak with infection rate of 92.3, at 28 days the expression of fluorescence was still observed. It is concluded that the recombinant adenoviral vector Ad-EGFP-mKGF is successfully constructed and can transfect MSC effectively and safely.
Subject(s)
Animals , Male , Mice , Adenoviridae , Genetics , Bone Marrow Cells , Cell Biology , Fibroblast Growth Factor 7 , Genetics , Genetic Vectors , Green Fluorescent Proteins , Genetics , Mesenchymal Stem Cells , Cell Biology , Mice, Inbred C57BL , Plasmids , TransfectionABSTRACT
<p><b>OBJECTIVE</b>To explore the effects of total flavonoids of Hippophae rhamnoides L. (TFH), quercetin (Que) and isorhamnetin (Isor) on the intracellular free calcium ([Ca(2+)](i)) in vascular smooth muscle cells (VSMC) of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY).</p><p><b>METHODS</b>Fluo 3-acetoxymethylester (Fluo-3/AM) was used to observe the effects of TFH (100 mg/L) and its essential monomers, namely Que (10(-4) mol/L) and Isor (10(-4) mol/L) on changes of [Ca(2+)](i) in cultured SHR and WKY VSMC (abbr. to Ca-SHR & Ca-WKY) following exposure to high K(+), norepinephrine (NE) and angiotensin II (Ang II), and to compare with the effects of verapamil (Ver).</p><p><b>RESULTS</b>(1) TFH, Que and Isor had inhibitory effects on resting Ca-SHR (P < 0.05), but had no significant effects on Ca-WKY (P > 0.05). (2) High K(+) could increase Ca-SHR more significantly than Ca-WKY (P < 0.05); TFH, Que and Isor could inhibit the elevation of [Ca(2+)](i) induced by high K(+)-depolarization, with the effects similar to that of Ver, and the effect on Ca-SHR was more significant than that on Ca-WKY (P < 0.05). (3) NE and Ang II could increase Ca-SHR more significantly than Ca-WKY (P < 0.05), TFH, Que and Isor had remarkably inhibitory effect on the elevation of Ca-SHR and Ca-WKY induced by NE or Ang II. (4) In the absence of extracellular Ca(2+), TFH, Que and Isor also had certain inhibitory effect on Ca-SHR and Ca-WKY induced by NE, and the effect on the former was more significant than that on the latter (P < 0.05).</p><p><b>CONCLUSION</b>TFH, Que and Isor might decrease the levels of [Ca(2+)](i) in VSMCs by blocking both voltage-dependent calcium channels (VDC) and receptor-operated calcium channels (ROC) in physiological or pathological state, which may be one of the important mechanisms of their hypotensive and protective effects on target organs in patients with hypertension.</p>