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The Anti-Proliferation Effect of Saponins from Tribulus Terrestris L. on Renal Carcinoma Cell Line 786-O

Author: YangHuangJian
Tutor: ZuoWeiZuo
School: East China Normal University
Course: Botany
Keywords: Tribulus Terrestris Renal tumors Apoptosis Cell Cycle Bcl-2 Ca2 Mitochondrial membrane potential RP-HPLC Save Time Trace elements Glycation endproducts Fuzzy Clustering Analysis
CLC: R737.11
Type: Master's thesis
Year: 2006
Downloads: 102
Quote: 0
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Abstract


Tribulus (Tribulus terrestris L.) as set out in our pharmacopoeia into traditional Chinese medicine, the main medicinal parts of many pharmacological efficacy studies, while terrestris whole plant medicine knowledge has just begun. Early research group found that the whole plant Tribulus terrestris saponins (saponins from Tribulus terrestris L. referred STT) can effectively reduce high blood sugar pathological state level work glycosylation basis, suggesting that this intervention on STT glycation endproducts (Advanced Glycation End P roducts , referred to as AGEs) malignant lesions of diabetic nephropathy resulting potential effects of an in-depth tracking, this is intended to human renal cancer 786-0 as target cells, cellular and molecular level from the level of in vitro expanded STT inhibitory effects on the kidney and its mechanism of action. First, using the MTT assay, SRB France observed STT 786-0 renal carcinoma cells in vitro for growth inhibition, and compare STT human embryonic kidney cells in vitro effects, the results show: 25 ~ 100μg/ml of STT for 786 - 0 cells was significantly inhibited and dose-dependent manner; 25 ~ 200μg/ml of STT for human embryonic kidney cell cytotoxicity was significantly less than on 786-0 cytotoxicity. To further proven STT tumor suppressor mechanism study wright staining, acridine orange staining and transmission electron microscopy role in tumor cells STT cell morphology and ultrastructure changes; agarose gel electrophoresis and the use of diphenylamine Determination of DNA fragmentation; application propidium iodide (PI) staining was detected by flow cytometry STT on 786-0 cell cycle; combined immunofluorescence, flow cytometry to STT on 786-0 cyclin expression and the role of cell STT antiapoptotic protein Bcl-2 levels and intracellular calcium ion concentration, mitochondrial membrane potential differences. The results showed that: ① 786-0 cells after action by the STT was early apoptotic characteristics: smaller cell shrinkage, nuclear condensation, uptake, nucleolus cracking, condensed chromatin margination; But agarose gel electrophoresis and flow cytometry cytometry showed that, STT 786-0 induced apoptosis rate is not high. ② STT 786-0 cells by down-regulating cyclin D1 protein levels, thus affecting 786-0 cell cycle regulation, namely reduced G 0 / G 1 phase cells, the cells DNA synthesis by blocking; G 2 / M phase cells decreased, reducing cell mitosis. ③ anti-apoptotic protein Bcl-2 downregulation is STT one of the molecular mechanisms of tumor suppressor role. ④ STT 786-0 can cause loss of mitochondrial transmembrane potential, activation of the mitochondrial pathway, which in turn induce 786-0 to death. ⑤ STT significantly increased intracellular Ca 2 levels, while intracellular Ca 2 level changes can enhance STT on 786-0 cells in vitro, in the medium Join 2m mol / L CaCl 2 could significantly inhibit STT 786-0 cells induced DNA fragmentation, and EGTA or ZnSO 4 does not affect the STT-induced cell DNA 786-0 degree of fragmentation, suggesting that activation of calcium-dependent STT can endonuclease (such as DNase Ⅰ or DNase Ⅹ, etc.), causing a series of biochemical reactions. The content of the material basis of efficacy reliable and strict quality standards, is an important part of modern medicine. For this reason, the relevant indications exist for STT effective dose swings, established in our laboratory by RP-HPLC [Kromasil C18 (250mm × 4.6mm, 5um) and a refractive index detector combined with technical support, with 100 there are differences, STT monomer pharmacodynamic track found Haike sapogenin STT in vitro anti-tumor effect is the main material foundations. Has been consensus, herbal medicine is also trace elements contained in the material basis of efficacy important part. Studies based on traditional Chinese medicine theory of coordination chemistry theoretical guidance for the whole herb Tribulus resources in the field of information blind spots, combined with the laboratory material extract on six intervention of AGEs formed by using fuzzy cluster analysis of these types of materials trace elements associated with its efficacy. By inductively coupled plasma atomic emission spectroscopy (Inductively Coupled Plasma-Atomic Emission Spectrometry, referred to ICP-AES) determination of copper, magnesium, zinc, manganese, chromium, molybdenum, vanadium, zinc binding / copper and zinc / manganese ratio, data poor method of statistical indicators standardization; plot method to calculate the number of objects to be classified among the correlation coefficient to determine the domain of the fuzzy clustering analysis matrix. The results showed: several pairs to inhibit the formation of AGEs, prickly pear fruit marc flavonoids and flavonoids proximity effect; correlation coefficient λ = 0.75, the above two substances similar clustering. STT on far less inhibited the formation of AGEs other materials only in the GXL system effective, but in vitro anti-tumor effect on terrestris extract significantly stronger than other materials. The two saponins isolated STT containing ligand group such as a hydroxyl, and trace elements may be present with bit effect, STT contained in the organic components, and type of trace elements 786-0 and inhibition of cell proliferation effect, interference effect of the formation of AGEs other closely related.

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CLC: > Medicine, health > Oncology > Genitourinary tumors > Urinary tumors > Kidney,renal pelvis tumor
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