Establishment of Specific Chromatogram and Thin-layer Identification Method for Kaixinsan / 中国实验方剂学杂志

ObjectiveTo establish the specific chromatogram and thin layer chromatography（TLC） identification method of Kaixinsan（KXS） samples， in order to clarify the key quality attributes and provide reference for the quality evaluation of KXS. MethodHigh performance liquid chromatography（HPLC） specific chromatogram of KXS was developed with YMC Hydrosphere C18 column（4.6 mm×250 mm， 5 μm）， the mobile phase was acetonitrile（A）-0.2% formic acid aqueous solution（B） for gradient elution（0-15 min， 2%-20%A； 15-25 min， 20%-25%A； 25-30 min， 25%-30%A； 30-45 min， 30%-31%A； 45-50 min， 31%-44%A； 50-65 min， 44%-45%A； 65-73 min， 45%-75%A； 73-95 min， 75%-100%A； 95-105 min， 100%A； 105-105.1 min， 100%-2%A； 105.1-120 min， 2%A）， the detection wavelength was 320 nm. Ultra high performance liquid chromatography-linear ion trap-electrostatic field orbitrap mass spectrometry（UHPLC-LTQ-Orbitrap MS） was used to identify the chemical components of KXS with electrospray ionization（ESI）， negative ion mode and scanning range of m/z 50-2 000. TLC identification methods for Poria and Ginseng Radix et Rhizoma in KXS were established. ResultThere were 11 common peaks in the specific chromatogram of KXS， attributed to Polygalae Radix， Poria and Acori Tatarinowii Rhizoma. Taking peak 9（α-asarone） as the reference peak， the relative standard deviations of the retention times of 15 batches of KXS samples were<0.2%. A total of 34 compounds were identified by UHPLC-LTQ-Orbitrap MS， including terpenoids， phenylpropanoids， oligosaccharides and ketones. The established TLC had good separation and was rapid， reliable， simple， feasible， suitable for the identification of Poria and Ginseng Radix et Rhizoma in KXS. ConclusionThe specific chromatogram and TLC of KXS are stable and reproducible. The material basis of KXS is basically clarified by MS， which can provide a reference for the development and quality control of KXS.

Effect of lumbrokinase on the synthesis of Src and FAK induced by TGF-β1 in human renal tubular epithelial cells / 国际中医中药杂志

Objective To observe the regulation of lumbrukinase on the expression of α-smooth muscle actin,fibronectin,focal adhesion kinase and Src kinase induced by transforming growth factor β1 in human renal tubular epithelial cells.Methods According to random number table method,the human renal tubular epithelial cells were divided into the normal group,TGF-β1 model group,benazepril group,lumbrukinase low,medium and high dose group.Except the normal group,the other groups were treated with TGF-β1 10 μg/ml.After 30 minutes,the benazepril group added benazepril 10 μmol/L,and the low,medium,high dose groups of lumbrukinase were respectively treated with lumbrokinase 30,60,120 U/ml to intervene.After 24 hours of cultivation.Western blotting and real-time PCR were used to detect the expression of α-SMA,FN,FAK and Src.Results Compared with the model group,the expressions of α-SMA (0.84 ± 0.14,0.72 ± 0.08,0.69 ± 0.05 vs.1.24 ± 0.03) and FN (0.59 ± 0.09,0.55 ± 0.11,0.44 ± 0.08 vs.0.83 ± 0.18) and FAK (0.94 ± 0.04,0.79 ± 0.05,0.70 ± 0.02 vs.1.29 ± 0.07) and Src (0.87 ± 0.20,0.78 ± 0.15,0.71 ± 0.11 vs.1.23 ± 0.01) proteins in the high doses of lumbrical kinase group were significantly lower than those in the model group (P＜0.05),the expressions of α-SMA (3.13 ± 0.62,2.76 ± 0.14,2.15 ± 0.33 vs.4.12 ± 0.32) and FN (3.08 ± 0.34,2.78 ± 0.17,2.49 ± 0.11 vs.4.34 ± 0.06) and FAK (1.73 ± 0.23,1.63 ± 0.36,1.57 ± 0.27 vs.2.61 ± 0.59) and Src (2.11 ± 0.17,1.78 ± 0.25,1.71 ± 0.22 vs.2.78 ± 0.47) mRNA in the high doses of lumbrical kinase group decreased significantly (P＜0.05).Conclusions Lumbrokinase may prevent the development of renal fibrosis by regulating the expression ofFN,FAK and and reducing the production of α-SMA,FN.

Effect of drug-containing serum of Fushen Jiangzhuo formula on anti-fi-brosis factors HGF and BMP-7 in rat renal interstitial fibroblasts with re-nal tubulointerstitial lesion / 中国病理生理杂志

AIM:To study the protective effects and mechanism of Fushen Jiangzhuo formula (FSJZ) on the rat renal interstitial fibroblasts with renal tubulointerstitial lesion .METHODS:The serum containing FSJZ and blank con-trol serum were prepared .The rat model of mesangial proliferative glomerulonephritis ( MsPGN) was established and ex-tended the time of modeling to 20th weeks for developing tubulointerstitial lesion naturally .The rat renal interstitial fibro-blasts at the end of 12, 16, 20 week of modeling were isolated and cultured .The effects of FSJZ on the expression of hepa-tocyte growth factor (HGF) and bone morphogenetic protein-7 (BMP-7) at mRNA and protein levels in the pathological re-nal interstitial fibroblasts were determined .RESULTS:The anti-fibrosis factors HGF and BMP-7 were changed in patho-logical renal interstitial fibroblasts .Renal tubulointerstitial lesion significantly down-regulated the expression of HGF and BMP-7, while the drug containing serum of FSJZ significantly up-regulated the expression of HGF and BMP-7.CONCLU-SION:Drug containing serum of FSJZ has protective effect on pathological renal interstitial fibroblasts by regulating the mRNA and protein expression of HGF and BMP-7.

Establishment and application of a high-throughput screening assay for premature activation of HIV-1 precursors / 药学学报

Strict regulation of HIV-1 PR function is critical for efficient production of mature viral particles. During viral protein expression and viral assembly, HIV-1 PR located within Gag-Pol precursor must be inactive to prevent premature cytoplasmic processing of the viral Gag and Gag-Pol precursors. Premature activation of HIV-1 precursors leads to major defects in viral assembly and production of viral particles. A cell-level premature activation of HIV-1 precursors assay using bioluminescence resonance energy transfer (BRET) was established. Three thousand compounds were screened to evaluate this assay. The results showed that the assay is sensitive, specific and stable (Z' factor is 0.905).

Establishment and application of a screening anti-HIV-1 drug model targeted nuclear trafficking of virus RNA / 药学学报

The HIV-1 Rev protein facilitates nuclear export of unspliced and singly spliced viral transcripts containing RRE RNA through the CRM1 export pathway. Inhibition of Rev-mediated RNA nuclear export can arrest HIV-1 transcriptional process, which clearly, reveals a target for anti-HIV drug development. In this work, a cell-based assay has been established for screening anti-HIV compounds targeting the Rev-mediated RNA nuclear export. This assay utilized a codon-optimized green fluorescent protein (GFP) as reporter gene, which expression is in a Rev-dependent manner. Any compound that inhibits the Rev-mediated RNA nuclear export is identified by reducing emission of GFP. The Z' score of this model is 0.8220. Three thousands compounds were screened and the positive rate was 9.3% with a cutoff at 50% inhibition. IMB7C7, one of the positive compounds, efficiently inhibits viral production from HIV-1 infected cells.