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Objective::To investigate the effect of drug-containing serum of Jianpi Xiaoai prescription on protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in colorectal cells HCT116. Method::The HTC116 cells were treated by 15%concentration of drug-contained serum, and then the cell migration and invasion were detected by Transwell assay, the protein expression levels of Akt, phosphorylated protein kinase B (p-Akt), mTOR, phosphorylated mammalian target of rapamycin (p-mTOR), ribosomal protein S6 kinase, polypeptide1(S6K1), phosphorylated ribosomal protein S6 kinase, polypeptide1 (p-S6K1), 4E-binding protein1(4EBP1), and phosphorylated 4E-binding protein1(p-4EBP1) in HCT116 cells were detected by Western blot. The control group was treated by untreated serum (15%), and 10%fetal bovine serum(FBS). Result::As compared with the control group, the number of migration and invasion cells was significantly reduced in drug-contained serum group (P<0.01), the expression of Akt had no obvious decrease, p-Akt protein expression was significantly lowered in the drug-contained serum group (P<0.01), the expression of mTOR had no obvious decrease, but p-mTOR protein expression was significantly lowered in drug-contained serum group (P<0.01), the expression of S6K1 had no obvious decrease, but p-S6K1 protein expression was significantly lowered in the drug-contained serum group (P<0.01), the protein expression of 4EBP1 had no obvious decrease, but p-4EBP1 protein expression was significantly lowered in the drug-contained serum group (P<0.01). Conclusion::The anti-tumor mechanism and transfer of Jianpi Xiaoai prescription may be related to inhibiting the activation of Akt/mTOR signaling pathways in colorectal cancer.
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Objective:To observe the effect of polyphyllin Ⅰ on the expressions of forkhead box Q1（FOXQ1）and epithelial-mesenchymal transition (EMT)-related factors, in order to explore the possible mechanism of polyphyllin Ⅰ in inhibiting the metastasis of colon cancer. Method:After the treatment with 1.25，2.50 μmol·L-1 polyphyllin Ⅰ on HCT116 cells, Western blot and Real-time PCR were used to detect the expressions of FOXQ1，E-cadherin，Vimentin protein and mRNA. Result:Compared with the blank group, relative expressions of FOXQ1 protein and mRNA in low-concentration polyphyllin Ⅰ group were decreased, while relative expression of E-cadherin mRNA was increased, the differences were not statistically significant, and relative expressions of Vimentin protein and mRNA in low-concentration polyphyllin Ⅰ group were decreased (P<0.05)， and relative expression of E-cadherin protein in low-concentration polyphyllin Ⅰ group was increased (P<0.01). Compared with the blank group, relative expressions of FOXQ1, Vimentin protein and mRNA in high-concentration polyphyllin Ⅰ group were decreased，while relative expressions of E-cadherin protein and mRNA were increased (P<0.05, P<0.01). Compared with low-concentration polyphyllin Ⅰ group, relative expressions of Vimentin protein and mRNA in high-concentration polyphyllin Ⅰ group were decreased, but the difference was not statistically significant, relative expressions of E-cadherin protein and mRNA in high-concentration polyphyllin Ⅰ group were increased, whereas relative expressions of FOXQ1 protein and mRNA were decreased (P<0.05, P<0.01). Conclusion:Mechanism of polyphyllin Ⅰ inhibiting the metastasis of colon cancer may be related to the decrease of FOXQ1 and Vimentin expressions, and the up-regulation of E-cadherin.
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Objective:To observe the effect of Jianpi Xiaoai prescription on the activation of normal human embryonic lung fibroblasts （HFL1） into tumor-associated fibroblasts （CAFs） induced by human colon cancer cells （HCT116） derived exosomes. Method:SD rats were gavaged with 13.1 g·kg-1 of Jianpi Xiaoai prescription to prepare drug-containing serum， and HCT116 cell exosomes-containing 10% exosomes-free serum and 20% Jianpi Xiaoai prescription drug serum were isolated by ultra-high speed centrifugation. The particle size distribution of exosomes were detected by Nanoparticle tracking analyzer （Zetaview）， and the exosomes' marker proteins apoptotic transfer gene 2 interaction protein X （Alix）， heat shock protein 70 （HSP70）， and tumor-susceptibility gene 101 （TSG101） were identified by Western blot， and the uptake of exosomes labeled with cell membrane staining kit （PKH67） by HFL1 was observed by fluorescence microscope. HFL1 cells were divided into six groups： the blank group， the transforming growth factor-β1 （TGF-β1） group， the TGF-β1 combined with HCT116 exosomes of 2 mg·L-1 group， the TGF-β1 combined with HCT116 exosomes of 4 mg·L-1 group， the TGF-β1 combined with Jianpi Xiaoai prescription exosomes of 2 mg·L-1 group， and the TGF-β1 combined with Jianpi Xiaoai prescription exosomes of 4 mg·L-1 group， and all groups were cultivated for 48 h. Western blot and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） were used to determine the protein and mRNA expressions of α-smooth muscle actin （α-SMA）. Result:The particle size distribution detected by Zetaview was mainly between 50-100 nm， and the exosomes were verified based on the expressions of marker proteins Alix， HSP70 and TSG101. After co-incubation of HFL1 cells with exosomes， a large number of exosomes were absorbed by HFL1 cells under fluorescence microscope. Compared with the blank control group， the protein and mRNA expressions of α-SMA in the TGF-β1 group and TGF-β1 combined with HCT116 exosome groups were increased （P<0.01）. Compared with the TGF-β1 combined with HCT116 exosome groups， the protein and mRNA expressions of α-SMA were decreased in the TGF-β1 combined with Jianpi Xiaoai prescription exosome groups （P<0.01）. Conclusion:Human colon cancer cell exosomes combined with TGF-β1 can induce the activation of HFL1 into CAFs， and Jianpi Xiaoai prescription can reduce the activation of HFL1 by affecting the expressions of α-SMA， thus antagonizing the lung metastasis of colon cancer.
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Objective To observe the effects of Jianpi Xiaoai Prescription on epithelial-mesenchymal transition of colorectal cancer cell SW620 induced by TGF-β1; To discuss its possible mechanism of action for prevention and treatment of colorectal cancer. Methods Colorectal cancer cells SW620 were cultured in vitro. By conducting CCK-8 and Transwell experiments, the proliferation, invasion and migration of colorectal cancer cell (SW620) were detected. qRT-PCR and Western blot experiments were applied to verify the mRNA and protein expression level of E-cadherin and Vimentin. Results Jianpi Xiaoai Prescription showed in vitro inhibitory effect on the proliferation, invasion and migration of colorectal cancer cell SW620. Compared with blank group, the expression of E-cadherin in TGF-β1 induced group was reduced and Vimentin was increased (P<0.05); Meanwhile, the expression of E-cadherin in Jianpi Xiaoai Prescription group was increased and Vimentin was decreased when compared with TGF-β1 induced group (P<0.05). Conclusion Jianpi Xiaoai Prescription can inhibit the proliferation, invasion and migration of colorectal cancer cell by reverting the epithelial-mesenchymal transition of colorectal cancer cell induced by TGF-β1, with a purpose to achieve the goal of preventing and treating the recurring and migration of colorectal cancer.
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It is important to establish animal models of Parkinson disease (PD) similar to clinical features of human disease. Rotenone can readily penetrate the blood-brain-barrier and cytomem-branes due to its strong lipophilic ability. Rotenone models of PD can not only simulate behavioral changes in patients with PD, but also bear a strong resemblance to the human disease characteristics and pathological process of PD. Based on to researches at home and abroad in recent years, this paper summarizes and analyzes the modeling methods and toxic mechanisms of rotenone-induced PD. These methods include stereotaxis, intravenous injection, abdominal injection, subcutaneous injection, microdialysis drug, intragastric administration, subcutaneous embedded slow-release microspheres and exposure to drugs. The In vitro model invotves SH-SY5Y cells, PC12 cells and DA neurons. The toxic mechanisms involveα-synuclein abnormal aggregation, mitochondrial dysfunction, the generation of reactive oxygen species, damage to the antioxidant defense system, nerve cell apoptosis, and autophagy.
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Objective: This article is aimed at investigating the effect of Jianpi Xiaoai Recipe (JXR) on the mRNA expression of vascular endothelial growth factor (VEGF) and endostation (ES) in the tissue in nude mice with colorectal cancer metastasis. Methods: Eight BALB/c-nu nude mice were randomly selected as the control group. The remaining mice, after establishing the metastatic tumor by injecting in the appendix of 50 μL human colon carcinoma HCT116 cells, then 32 survival nude mice were taken after 3 d, and divided into JXR (low- and high-dose, 10 and 40 g/kg), 5-FU (0.13 g/kg), and model groups. After the mice were ig or ip given corresponding medicine and physiological saline for 4 weeks, the mRNA expression levels of VGEF and ES in the metastatic tumor tissue of colon, liver, lung, and cecum were evaluated by qRT-PCR method. Results: The mRNA expression of VEGF was higher in all groups except the control group, and the highest was in the model and 5-FU groups (P < 0.01). Compared with the model group, the mRNA expression level in two JXR groups were significantly lower (P < 0.05, 0.01). ES transcription expression was lower in each group than that in the control group, the lowest was in the model and 5-FU groups (P < 0.01). Compared with the model group, the mRNA expression levels in the two JXR groups were significantly higher (P < 0.05, 0.01). Among these four organs, the expression level of VEGF in cecum was lowest, while the mRNA expression levels of ES are nearly the same. Conclusion: JXR is possible to make the mRNA expression level of VEGF decreased, but the mRNA expression level of ES increased in the metastatic tumor tissue of colon, liver, lung, and cecum, so as to resist the metastasis of colorectal cancer.
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<p><b>AIM</b>To establish a sensitive and specific HPLC method for controlling the quality of total glycosides from Swertia franchetiana H. Smith.</p><p><b>METHODS</b>HPLC method was applied for quality and quantitative assessment of the pharmaceutical extracts from Swertia franchetiana H. Smith. The preparation of sample, the HPLC column, mobile phase, elution mode (isocratic or gradient) and gradient program were optimized in order to obtain HPLC profile. The HPLC system consisted of a SPD-1OAvp pump, SPD-M1OAVP photodiode-array detector (PAD), SIL-10ADVP auto injector. Data were acquired and processed with the CLASS-VP6.1 workstation. HPLC analysis was performed on a Kromasil C18 column (250 mm x 4. 6 mm ID, 5 microm) with methanol and water as mobile phase. The column temperature was set up at 40 degrees C and the flow-rate was 1 mL x min(-1). The reference solution of chemical standards and sample were injected into HPLC system, separately.</p><p><b>RESULTS</b>The HPLC chromatographic fingerprinting of the total glycosides, showing 16 characteristic peaks which were partitioned into three parts: one peak in 0-10 min of retention time, nine peaks containing main 1-7 peaks in 10-15 min of retention time, 6 peaks in 15-30 min of retention time, was established from 10 lots of their products. By comparison of the retention time and the on-line UV spectra and their molecule weights of chemical standards, peak 1-7 were identified as swertiamarin (1), gentiopicroside (2), sweroside (3), isoorientin (4), swertisin (5), isoswertisin (6) and swetianolin (7), respectively. The ratios of peak area between 1-16 were in their extent. Moreover, comparison of the HPLC profiles of the total glycosides, the extracts prepared using another process and the plant indicated that they were closely related to each other.</p><p><b>CONCLUSION</b>The HPLC profiles and quantitative assessment of the total glycosides from Swertia franchetiana H. Smith with high specificity can be used to control their quality and assure lot to lot consistency.</p>