ABSTRACT
Aim To explore the antidepressant mecha-nism and laws of traditional Chinese formula Yueju Pill by taking drug pair as the breakthrough point. Meth-ods On the basis of anti-depressant activities of Yueju Pill, the combination of different herbs was obtained by the successively disassembling, and the key drug pair was obtained through the acute administration of Yueju Pill in mice. In addition, chronic unpredictable mild stress model was established to further verify the anti-depressant effect of key drug pair, and to explore its molecular mechanism. Results The drug pair of zhizi and chuanxiong was necessary to anti-depression effect of Yueju Pill, and the immobility time of TST and FST was significantly reduced. As expected, the expres-sions of IL-6 and TNF-a and p-NF-kBp65, P-IkBa were obviously lower than those in model group, but the expressions of BDNF and TrkB were up-regulated than those in model group. Conclusions The drug pair of zhizi and chuanxiong is necessary for traditional Chinese formula Yueju Pill for the antidepressant effect. It is assumed that the antidepressant effect and mechanism of zhizi and chuanxiong are connected with cytokine IL-6, TNF-a and protein expressions of p-NF-kBp65, P-IkBa, and BDNF.
ABSTRACT
The vitamin K epoxide reductase complex subunit 1 (VKORC1), the rate-limiting enzyme for vitamin K recycling, is significantly down-regulated in the kidneys of urolithiasis patients. This study searched for direct evidence to define the inhibitory activity of VKORC1 against calcium oxalate (CaOx) crystal formation. In the experiment of VKORC1 overexpression, HK-2 cells were transfected with the pFLAG-CMV-7.1-VKORC1 plasmid as a pFLAG-CMV-7.1-VKORC1 transfection group or the pFLAG-CMV-7.1 plasmid as a pFLAG-CMV-7.1 control group. In the experiment of VKORC1 knockdown, HK-2 cells were transfected with the PGPU6/GFP/Neo-VKORC1shRNA-2 as a PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group or the PGPU6/GFP/Neo-shRNA-NC plasmid as a PGPU6/GFP/Neo-shRNA-NC control group. The expression of VKORC1 in HK-2 cells was detected by real-time quantitative PCR and Western blotting. The CaOx crystal formation was observed under the laser-scanning confocal microscope. It was found that the expression levels of VKORC1 mRNA and protein were significantly higher in the pFLAG-CMV-7.1-VKORC1 transfection group than in the pFLAG-CMV-7.1 control group (P<0.01). The number of CaOx crystals in HK-2 cells incubated in fluorescently labeled CaOx monohydrate (COM) crystal medium for 48 h was 14±4 per field (100×) in the pFLAG-CMV-7.1-VKORC1 transfection group and 26±5 per field (100×) in the pFLAG-CMV-7.1 control group respectively under the laser-scanning confocal microscope. The amount of CaOx crystal aggregation and formation in the pFLAG-CMV-7.1-VKORC1 transfection group was significantly reduced as compared with the pFLAG-CMV-7.1 control group (P<0.05). The expression levels of VKORC1 mRNA and protein were significantly lower in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group than in the PGPU6/GFP/Neo-shRNA-NC control group (P<0.05). The number of CaOx crystals in HK-2 cells incubated in fluorescently labeled COM crystal medium was 65±11 per field (100×) in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group and 24±6 per field (100×) in the PGPU6/GFP/Neo-shRNA-NC control group respectively under the laser-scanning confocal microscope. The amount of CaOx crystal aggregation and formation in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group was significantly increased as compared with the PGPU6/GFP/Neo-shRNA-NC control group (P<0.05). These findings suggested that the VKORC1 protein could inhibit CaOx salt crystallization, adhesion and aggregation. This research would help us to understand the mechanisms involving the interaction between crystallization and epithelial cells and the formation of CaOx.
Subject(s)
Humans , Apoptosis , Blotting, Western , Calcium Oxalate , Chemistry , Metabolism , Pharmacology , Cell Line , Crystallization , Dose-Response Relationship, Drug , Flow Cytometry , Gene Expression , Green Fluorescent Proteins , Genetics , Metabolism , Microscopy, Confocal , RNA Interference , Reverse Transcriptase Polymerase Chain Reaction , Time Factors , Transfection , Vitamin K Epoxide Reductases , Genetics , MetabolismABSTRACT
The vitamin K epoxide reductase complex subunit 1 (VKORC1), the rate-limiting enzyme for vitamin K recycling, is significantly down-regulated in the kidneys of urolithiasis patients. This study searched for direct evidence to define the inhibitory activity of VKORC1 against calcium oxalate (CaOx) crystal formation. In the experiment of VKORC1 overexpression, HK-2 cells were transfected with the pFLAG-CMV-7.1-VKORC1 plasmid as a pFLAG-CMV-7.1-VKORC1 transfection group or the pFLAG-CMV-7.1 plasmid as a pFLAG-CMV-7.1 control group. In the experiment of VKORC1 knockdown, HK-2 cells were transfected with the PGPU6/GFP/Neo-VKORC1shRNA-2 as a PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group or the PGPU6/GFP/Neo-shRNA-NC plasmid as a PGPU6/GFP/Neo-shRNA-NC control group. The expression of VKORC1 in HK-2 cells was detected by real-time quantitative PCR and Western blotting. The CaOx crystal formation was observed under the laser-scanning confocal microscope. It was found that the expression levels of VKORC1 mRNA and protein were significantly higher in the pFLAG-CMV-7.1-VKORC1 transfection group than in the pFLAG-CMV-7.1 control group (P<0.01). The number of CaOx crystals in HK-2 cells incubated in fluorescently labeled CaOx monohydrate (COM) crystal medium for 48 h was 14±4 per field (100×) in the pFLAG-CMV-7.1-VKORC1 transfection group and 26±5 per field (100×) in the pFLAG-CMV-7.1 control group respectively under the laser-scanning confocal microscope. The amount of CaOx crystal aggregation and formation in the pFLAG-CMV-7.1-VKORC1 transfection group was significantly reduced as compared with the pFLAG-CMV-7.1 control group (P<0.05). The expression levels of VKORC1 mRNA and protein were significantly lower in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group than in the PGPU6/GFP/Neo-shRNA-NC control group (P<0.05). The number of CaOx crystals in HK-2 cells incubated in fluorescently labeled COM crystal medium was 65±11 per field (100×) in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group and 24±6 per field (100×) in the PGPU6/GFP/Neo-shRNA-NC control group respectively under the laser-scanning confocal microscope. The amount of CaOx crystal aggregation and formation in the PGPU6/GFP/Neo-VKORC1shRNA-2 transfection group was significantly increased as compared with the PGPU6/GFP/Neo-shRNA-NC control group (P<0.05). These findings suggested that the VKORC1 protein could inhibit CaOx salt crystallization, adhesion and aggregation. This research would help us to understand the mechanisms involving the interaction between crystallization and epithelial cells and the formation of CaOx.
ABSTRACT
Objective To study the epidemic tendency of emerging influenza A (H1N1) in mainland China, and to explore the different patterns of spread on the disease under the following contexts: (1) To stop the temperature screening program at the border areas of the country; (2)To stop measures of prevention and control on those identified cases and their close contacts; (3) To strengthen programs for the foreign immigrants on 'home quarantine'. Methods Under relevant parameters and information on the transmission link from different reference data, the patterns of influenza spread were simulated by Monte Carlo method. Results The temperature screening on border could inhibit the transmission of influenza A (H1N1) to some extent, so that after 3 months the cumulative number of cases will be reduced by 21.5% (1718 cases) and transmission speed of influenza A (H1N1) in mainland China will be delayed by about 4 days. Furthermore, taking positive measures of prevention and control could efficiently slow down the epidemic, so that after 3 months the cumulative number of cases will be reduced by 93.4%(about 90 thousand cases) and it would be delayed by about 15 days if influenza A (H1N1) spreads to the whole country. In addition, if the immigrants were able to practise quarantine measures consciously by themselves at home the effect of prevention and control against influenza A(H1N1) would be more significant. If 30%, 60% and 90% of immigrants would take quarantine measures home consciously, after 3 months the cumulative number of cases will be reduced by about 15% (about 940 cases), 34% (about 2230 cases) and 64% (about 4180 cases), respectively. Also, influenza A (H1N1) spreads to the whole country will be delayed by about 4 days, 10 days and 25 days, respectively. It is difficult to curb fully the development of the epidemic by taking existing control measures, and influenza A (H1N1) may spread to almost all provinces after about 3 months. Conclusion The effects of existing prevention and control measures were objectively assessed and the results showed the necessity and effectiveness of these measures against the transmission of influenza A (H1N1) , in the mainland of China.