本文關(guān)鍵詞： 桑寄生水煎液 肢芽 Tbx BMP-　出處：《毒理學(xué)雜志》2017年02期 　論文類型：期刊論文

【摘要】：目的探討桑寄生水煎液對(duì)小鼠胚胎肢芽生長(zhǎng)發(fā)育及T-框蛋白2(Tbx2)和骨形成蛋白2(BMP-2)mRNA和蛋白表達(dá)的影響,評(píng)價(jià)桑寄生的胚胎發(fā)育毒性。方法利用超高高效液相色譜儀鑒定藥材桑寄生,常規(guī)法制成水提液;取E13(embryos on day 13)前肢芽隨機(jī)分為低、中、高劑量組和空白對(duì)照組,分別加入0.5、1和2 mg/ml不同濃度的桑寄生水煎液和雙蒸水懸浮培養(yǎng),72 h后收獲肢芽,測(cè)量肢芽長(zhǎng)度后,阿利新藍(lán)染色用Neubert評(píng)分系統(tǒng)進(jìn)行評(píng)分以評(píng)價(jià)肢芽軟骨的發(fā)育情況;采用實(shí)時(shí)熒光定量PCR和免疫組化法分別檢測(cè)肢芽中Tbx2和BMP-2 mRNA和蛋白的表達(dá)水平。結(jié)果 (1)經(jīng)UPLC分析,槲皮苷峰面積比為38.45%,可鑒定所用藥材為桑寄生。(2)肢芽長(zhǎng)度測(cè)量:低、中、高劑量組和空白對(duì)照組的長(zhǎng)度分別為1091.18±94.32、1107.07±80.29、1118.76±97.49和1080±89.54μm,各劑量組與空白對(duì)照組比較差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。(3)Neubert評(píng)分:低、中、高劑量組和空白對(duì)照組總評(píng)分分別為223.24±28.32、208.24±28.32、213.30±30.59和201.7±28.94。高劑量組評(píng)分最高,與空白對(duì)照組比較差異有統(tǒng)計(jì)學(xué)意義(P0.05),低和中劑量組與空白對(duì)照組比較差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。(4)實(shí)時(shí)熒光定量PCR和免疫組化結(jié)果顯示:各劑量桑寄生水煎液均上調(diào)了Tbx2和BMP-2 mRNA和蛋白的表達(dá)水平,其中高劑量組與空白對(duì)照組比較差異有統(tǒng)計(jì)學(xué)意義(P0.05),而低和中劑量與空白對(duì)照組比較差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論各劑量桑寄生水煎液對(duì)小鼠胚胎肢芽的生長(zhǎng)發(fā)育未體現(xiàn)出胚胎發(fā)育毒性,高劑量可誘導(dǎo)Tbx2和BMP-2 mRNA和蛋白表達(dá)促進(jìn)肢芽軟骨的發(fā)育。
[Abstract]:Objective to investigate the effects of water extract of Mori parasiticus on the growth and development of mouse embryonic limb buds and the expression of T- box protein 2 (Tbx2) and bone morphogenetic protein 2BMP 2 (BMP-2) mRNA and protein, and to evaluate the embryonic developmental toxicity of Mori parasiticus. Methods the parasitism of Morus mulberry was identified by ultrahigh performance liquid chromatography (UHP-HPLC). The forelimb buds of E13 embryo on day 13 were randomly divided into low, middle and high dose groups and blank control group. The limb buds were harvested after 72 h suspension culture with different concentrations of 0. 5 and 2 mg/ml water solution and double distilled water, respectively. After the length of limb bud was measured, Alisin blue staining was used to evaluate the development of limb bud cartilage by Neubert scoring system. The expression levels of Tbx2 and BMP-2 mRNA and protein in limb buds were detected by real-time fluorescence quantitative PCR and immunohistochemistry respectively. Results: by UPLC analysis, the peak area ratio of quercetin was 38.45, which could be identified as mulberry parasite. The length of the high dose group and the blank control group were 1091.18 鹵94.32n 1107.07 鹵80.29v 1118.76 鹵97.49 and 1080 鹵89.54 渭 m, respectively. There was no significant difference between the high dose group and the blank control group in the score of Neubert: the total score of low, middle, high dose and blank control group was 223.24 鹵28.32n 208.24 鹵28.32n 213.30 鹵30.59 and 201.7 鹵28.94.The total score of the high dose group was the highest. Compared with the blank control group, the difference was statistically significant (P 0.05), but there was no significant difference between the low and middle dose groups and the blank control group (P 0.05). The results of real-time fluorescence quantitative PCR and immunohistochemistry showed that all the doses of the decoction were upregulated. The expression level of Tbx2 and BMP-2 mRNA and protein, There was significant difference between the high dose group and the blank control group, but there was no significant difference between the low and middle dose groups and the blank control group. Conclusion there is no significant difference in the growth and development of embryonic limb bud of mice treated with different doses of mulberry parasitic decoction. The toxicity of embryonic development, High dose could induce the expression of Tbx2 and BMP-2 mRNA and protein to promote the development of limb bud cartilage.
【作者單位】： 遵義醫(yī)學(xué)院珠海校區(qū)生物工程系;華南師范大學(xué)生命科學(xué)學(xué)院;香洲區(qū)人民醫(yī)院婦產(chǎn)科;
【基金】：貴州省中醫(yī)藥管理局中醫(yī)藥、民族醫(yī)藥科學(xué)技術(shù)研究項(xiàng)目(QZYY2013-98)
【分類號(hào)】：R285.5

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