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Objective The purpose of this study was to examine effects of Isopsoralen on the osteoblast proliferation and differentiation and find its possible molecular mechanisms for anti-osteoporosis.Methods OCT-1 cells were cultured with common methods.While growing well,cells were cultured with 3 doses(10 μg·mL-1,30u μg·mL-1 and 60 μg·mL-1)of Isopsoralen for 48 h,or with purified bone morphogenetic protein 2(BMP2)protein(50 ng·mL-1).We first determined the effect of Isopsoralen on cell proliferation by MTT assay.The real time RT-PCR was also used to quantify changes in the mRNA levels of several genes,such as BMP2,Runt-related transcription factor 2(Runx2),and Osterix(Osx).We also used the Western blot analysis to evaluate the expression of Runx2 and Osx proteins.At last we used the BMP2loxp/loxp mice to isolate the primary calvaria osteoblasts,cultured with Isopsoralen of the best dose for 48 h after the in vitro conditional gene knockout technology,and tested the gene expressions of Runx2 and Osx.And the alkaline Phosphatase(ALP)staining was also performed.Result Isopsoralen(10 μg·mL-1)can promote osteoblast proliferation obviously.From the real time RT-PCR analysis,Isopsoralen can enhance the BMP2 mRNA levels,the effect of 10 μg·mL-1 was the best,and 30 μg·mL-1 followed.In addition,we found that Isopsoralen(10 μg·mL-1)can enhance the Runx2 mRNA levels significantly.We also found that lower doses of Isopsoralen can enhance the Osx mRNA levels,the effect of 30 μg·mL-1 was the best,and 10 μg·mL-1 followed.From the Western blot analysis,low doses of Isopsoralen(10 μg·mL-1 and 30 μg·mL-1)can stimulate the expression of Runx2 protein.Besides,three doses of Isopsoralen can stimulate the expression of Osx protein,and the effect of 10 μg·mL-1 and 30 μg·mL-1 are better.Finally,the results of in vitro conditional gene knockout experiment showed that the overexpression of Runx2 and Osx genes in osteoblasts,as well as ALP staining,induced by Isopsoralen are BMP2 dependent.Conclusions In this study,we firstly demonstrate that Isopsoralen can stimulate osteoblast proliferation and differentiation by mediating BMP2/Runx2/Osx signaling pathway.
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The authors summarize Professor SHI Qi's clinical experience in diagnosing and treating chronic tendon and bone disease.The specific diagnosing and treating thinking and methods could be summarized as follows:1)Three stages,which means chronic tendon and bone disease could be treated according to early,medium and late stages.2) Three differentiations,which include differentiating disease,type and syndrome.3) Three examining,which include seeing patient clearly,reading the disease and getting the key point.In addition,Prof.SHI emphasizes threepoint syndrome differentiation which means the combination of the lesion's target,peri-target and whole syndrome characteristics differentiation.In the process of treatment,Prof.SHI emphasizes three methods combination of herb,technique and breathing technique.Both internal and external treatments should be used.Prof.SHI advocates that the control strategy should be the prevention,treatment and recuperation integration concept,including preventing disease,early treatment to prevent deterioration and preventing reoccurrence after cure.
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Objective To establish a HPLC method for determination of three active components in Shuanghua Caoshanhu Hanpian.Methods Agilent TC-C18 (4.6 mm × 250 mm,5 μm) column was used.The mobile phase was acetonitrile-water containing 0.2% phosphoric acid with gradient elution mode at a flow rate of 1.0 mL· min-1,the column temperature was 30 ℃,the detection wavelength was 327 mn (0-10 min) for chlorogenic acid,342 nm (10-35 min) for isofraxidin and msmarinic acid.Results Chlorogenic acid had a good linearity in the range of 0.052-0.939 μg (r =0.999 8,n =6),the average recovery was 99.23% with RSD of 1.4% (n =9).Isofraxidin had a good linearity in the range of 0.004-0.077 μg (r =0.999 8,n =6),the average recovery was 100.33 % with RSD of 1.8 % (n =9).Rosmarinic acid had a good linearity in the range of 0.012-0.213 μg (r =0.999 9,n =6),the average recovery was 99.42% with RSD of 1.4% (n =9).Conclusion The method is simple,accurate and reliable.It can be used to determine chlorogenic acid,isofraxidin and rosmarinic acid in Shuanghua Caoshanhu hanpian at the same chromatogram condition.
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Nourishing kidney-yin (NKY) granules and warming kidney-yang (WKY) granules represent one of the prescriptions that prescribed in treating primary osteoporosis (POP) in light of tonifying kidney and nourishing essence principle as well as the theory of "treating both the disease and traditional Chinese medicine (TCM) syndrome".Both granules were created through the systematic analysis of clinic prescriptions by Professor Shi Qi.Consequently clinical investigations have well established that NKY granules significant improved bone mineral density (BMD) as well as relieved the kidney-yin deficiency syndromes in POP patients.Meanwhile,WKY granules relieve kidney-yang deficiency syndrome and the quality of life (QOL).What is more,pharmacological study established the application of common cnidium fruit,and fructus ligustri lucidi alleviated bone loss in OVX-induced mice.In addition,investigation with effective components identified that both NKY and WKY granules play systematic pharmacological effects on bone remodeling by regulating the expression of BMP/Smad,Wnt/β-catenin,RANKL/RANK/OPG axis,and Notch.The drug discovery was performed by the lead of traditional Chinese medicine (TCM) theory.It is one successful transformation investigation based on pharmacological effects,clinical intervention,animal model,cell culture and molecular investigation.
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Objective:To establish a wavelength switching HPLC method for the simultaneous determination of chlorogenic acid and mangiferin in Yinhua Mangguo tablets. Methods:An Agilent ZORBAX Eclipse XDB-C18 (150 mm × 4. 6 mm, 5 μm) column was used. The mobile phase was methanol-water containing 0. 2% phosphoric acid with gradient elution at the flow rate of 1. 0 ml· min-1;the detection wavelength was 327 nm(0-8 min) for chlorogenic acid and 258 nm(8-20 min) for mangiferin. Results:Chlorogenic acid had good linearity within the range of 0. 140-2. 516 μg, and the average recovery was 99. 4% with RSD of 1. 2% (n=9). Mangiferin had good linearity within the range of 0. 042-0. 761 μg, and the average recovery was 99. 9% with RSD of 1. 4% (n=9). Conclu-sion:The method is simple, accurate and reproducible. It can be used to determine chlorogenic acid and mangiferin in Yinhua Mang-guo tablets with the same chromatogram conditions.
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Objective To establish an HPLC method of wavelength switching simultaneous determination for the contents of mangiferin and hesperidin inYinhua Mangguo Tablets.Methods Agilent ZORBAX Eclipse XDB-C18 (4.6 mm×150 mm, 5 μm) column was used. The mobile phase was methanol-water containing 0.2% phosphoric acid with gradient elution mode at the flow rate of 1.0 mL/min; the detection wavelength was 258 nm (0–20 min) for mangiferin, 283 nm (20–50 min) for hesperidin.Results Mangiferin had a good linearity in the range of 0.042–0.761 μg, and the average recovery was 99.86%, with RSD of 1.5% (n=9). Hesperidin had a good linearity in the range of 0.076–1.361 μg, and the average recovery was 99.91%, with RSD of 0.7% (n=9).Conclusion The method is simple, accurate and reproducible, which can be used to the quality control ofYinhua Mangguo Tablets.
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Objective:To set up a method for simultaneously determining the content of notoginsenoside R1 , ginsenoside Rg1 and ginsenoside Rb1 in compound Huanggen granules ( sugar free) . Methods:The HPLC method was carried out on an Agilent ZORBAX SB-C18(250 mm ×4.6 mm,5 μm)column. The mobile phase was acetonitrile-water with gradient elution. The flow rate was 1.0 ml· min-1 . The detection wavelength was at 203 nm. The column temperature was set at 25℃ and the sample size was 10 μl. Results:The linearity of notoginsenoside R1, ginsenoside Rg1 and ginsenoside Rb1 was within the range of 1. 6-10. 0 μg·ml-1(r=0. 999 6), 6. 3-39. 3 μg·ml-1(r=0. 999 8) and 6. 3-39. 7 μg·ml-1(r=0. 999 7), respectively. The average recovery was 98. 81%(RSD=1. 20%),99. 93%(RSD=0. 93%) and 99. 22%(RSD=0. 87%)(n=6) , respectively. Conclusion: The method is simple, repro-ducible and specific, which can be used in the quality control of the preparation.
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<p><b>BACKGROUND</b>Bone grafting is commonly used to repair bone defects. As the porosity of the graft scaffold increases, bone formation increases, but the strength decreases. Early attempts to engineer materials were not able to resolve this problem. In recent years, nanomaterials have demonstrated the unique ability to improve the material strength and toughness while stimulating new bone formation. In our previous studies, we synthesized a nano-scale material by reinforcing a porous β-tricalcium phosphate (β-TCP) ceramic scaffold with Na2O-MgO-P2O5-CaO bioglass (β-TCP/BG). However, the in vivo effects of the β-TCP/BG scaffold on bone repair remain unknown.</p><p><b>METHODS</b>We investigated the efficacy of β-TCP/BG scaffolds compared to autografts in a canine tibiofibula defect model. The tibiofibula defects were created in the right legs of 12 dogs, which were randomly assigned to either the scaffold group or the autograft group (six dogs per group). Radiographic evaluation was performed at 0, 4, 8, and 12 weeks post-surgery. The involved tibias were extracted at 12 weeks and were tested to failure via a three-point bending. After the biomechanical analysis, specimens were subsequently processed for scanning electron microscopy analysis and histological evaluations.</p><p><b>RESULTS</b>Radiographic evaluation at 12 weeks post-operation revealed many newly formed osseous calluses and bony unions in both groups. Both the maximum force and break force in the scaffold group (n = 6) were comparable to those in the autograft group (n = 6, P > 0.05), suggesting that the tissue-engineered bone repair achieved similar biomechanical properties to autograft bone repair. At 12 weeks post-operation, obvious new bone and blood vessel formations were observed in the artificial bone of the experimental group.</p><p><b>CONCLUSIONS</b>The results demonstrated that new bone formation and high bone strength were achieved in the β-TCP/BG scaffold group, and suggested that the β-TCP/BG scaffold could be used as a synthetic alternative to autografts for the repair of bone defects.</p>
Subject(s)
Animals , Dogs , Female , Autografts , Bone Substitutes , Chemistry , Ceramics , Fibula , Wounds and Injuries , General Surgery , Hydroxyapatites , Chemistry , Tibia , Wounds and Injuries , General Surgery , Tissue Engineering , Methods , Tissue Scaffolds , ChemistryABSTRACT
Objective To establish a method for detecting chlorogenic acid in compound oral liquid of Kangbingdu with HPLC.Methods The chromatographic column of Shim-pack VP-ODS C18(4.6 mm? 250 mm,5 ?m) was used,the mobile phase was consisted of acetonitrile-0.2% phosphoric acid aqueous solution(12∶88).The flow rate was 1.0 mL/min,the detective wavelength was 327 nm and column temperature was 30 ℃.Results The linear range of the method was 0.051~0.922 ?g(r=0.999 9,n=6),the average recovery was 99.04%,RSD was 1.31%(n=6).Conclusion The method is accurate,reproducible,simple and easy to do,which can be used in quality control of compound oral liquid of Kangbingdu.