ABSTRACT
@#To effectively screen treating pigmentation disorders drug, a new transgenic zebrafish drug screening model was constructed. Green fluorescent protein(GFP)were drived by tyrosinase-related protein 1a (tyrp1a)promoter specific expression in melanocyte. Effect of N-Phenylthiourea(PTU)and alpha-melanaocyte stimulating hormone(α-MSH)on the GFP expression and melanogenic of tyrp1a: eGFP zebrafish were photographed under the steromicroscope. Results showed that PTU could significantly inhibit the melanogenic and expression of GFP of zebrafish. As compared with control group, α-MSH treatment resulted in marked stimulation of body pigmentation and expression of GFP. In conclusion, a new transgenic zebrafish drug screening model was successfully established, which can be used for treating pigmentation disorders.
ABSTRACT
The purpose of this study was to establish a drug screening method for small molecules extracted from traditional Chinese medicines (TCM) that have neuronal differentiation promoting effects, using P19 embryonic carcinoma cell as a cell-based model. First, the constructed plasmid (pTα1-Luc) was transfected into P19 cells to establish a screening model. Second, several TCMs were screened using the established model and all-trans-retinoic acid as a positive control. Finally, the underlying molecular mechanism was explored using immunofluorescence staining, qT-PCR, and Western blot analysis. Our results indicated that the drug screen model was established successfully and that both honokiol and hyperoside induced P19 differentiation into neurons, with the possible molecular mechanism being modulating the Wnt signaling pathway. In conclusion, the drug screening model developed in the present study provides a rapid, cell-based screening platform for identifying natural compounds with neuronal differentiation effects.
Subject(s)
Animals , Mice , Biphenyl Compounds , Pharmacology , Cell Differentiation , Cell Line, Tumor , Cell Proliferation , Drug Evaluation, Preclinical , Methods , Drugs, Chinese Herbal , Pharmacology , Embryonal Carcinoma Stem Cells , Lignans , Pharmacology , Neurons , Quercetin , Pharmacology , Tretinoin , Physiology , Wnt Signaling PathwayABSTRACT
Objective: To establish a rapid high-throughput screening model with positvive transcription elongation factor (p-TEFb) as target for screening the inhibitors of HIV. Methods: Double cross plasmid of BD-Tat and AD-CyclinT1 was established and transformed into yeast AH109 and Y187, respectively. AH109/pGBKT7-Tat was mated with Y187/pGADT7-CyclinT1 and the diploids were subjected to autoactivation, toxicity test, and reporter gene assay. The screening model based on double hybrid system was established. Results: Two out of 20 kinds of Chinese materia medica possessing immunity-enhancing effects were identified as primary hits, the anti-HIV activity of these two positive drugs was already demonstrated by other researchers through detecting the suppression effects on HIV duplication in vitro. Conclusion: The system established in this paper can be used for rapid high-throughput screening anti-HIV chemicals or herbs. Further research for the obtained two positive Chinese materia medica, Cibotii Rhizoma and Coptidis Rhizom should be carried out to isolate the effective component for disrupting the interaction between HIV Tat and CyclinT1.
ABSTRACT
AIM: To find small molecule organic mimetics of G-CSF, according to the transcription regulating effect of the important transcription factors-STAT3 (signal transducers and activators of transcription3) in hematopoiesis growth factor's signal transductional passageway, we established a drug screening model targeting transcription regulating of STAT3. METHODS: A recombinat vector pTKS3-CAT was constructed. Then pTKS3-CAT and pTK-Hyg were transfected into NFS-60 cells expressing G-CSF receptor with lipofectamine 2000. Cells derived from hygromycin-resistant colonies were tested for CAT activity, and positive colonies were isolated. At the same time, the sensitivity and the specialty of the model were tested. RESULTS: A dose-dependent expression of CAT gene with half-maximal induction by rhG-CSF at 0.044 nmol·L-1 was observed. After treating with rhEPO CAT activity couldn't be tested. CONCLUSION: The expression of CAT gene could be strongly induced by its special ligands in drug screening model. This model can be used to assay CAT from extracts of cells grown with CAT-ELISA method in microtiter wells and then to screen small molecular compounds with G-CSF-like activity.