Chinese Journal of Experimental Traditional Medical Formulae
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    2021,27(11):1-8, DOI: 10.13422/j.cnki.syfjx.20211005
    Abstract:
    Objective To explore the progression of diabetic macrovascular disease and the effects of Didangtang at different doses on it.Method Four-week-old male apolipoprotein-E knockout (ApoE-/-) mice with diabetic macrovascular disease induced by exposure to high-fat diet combined with streptozotocin (STZ) were randomly divided into the model, simvastatin, as well as high-, medium-, and low-dose Didangtang groups. The age-matched ApoE-/- mice of the same batch only fed with a high-fat diet were classified into the ApoE-/- (model control) group, and C57BL/6 mice with the same genetic background receiving a regular diet into the normal group. The sampling was conducted at the 8th and 20th weeks of the experiment for observing the pathological characteristics of the aorta and the proportion of plaque area in mice of each group at different time points, followed by the comparison of blood glucose, blood lipid, and oxidized low-density lipoprotein (ox-LDL) levels. The aortic NOD-like receptor protein 3 (NLRP3) and cysteinyl aspartate-specific proteinase-1 (Caspase-1) protein expression was detected by Western blot assay, and the serum interleukin-1β (IL-1β), interleukin-18 (IL-18), interleukin-1α (IL-1α), and tumor necrosis factor-α (TNF-α) levels by enzyme-linked immunosorbent assay (ELISA).Result The comparison with the normal group revealed that the proportions of plaque area in the ApoE-/- group and the model group were increased (P<0.01), while the proportion of plaque area in each administration group was significantly reduced in contrast to that of the model group (P<0.05). The aortic NLRP3 and Caspase-1 protein expression levels as well as the serum IL-1β, IL-18, IL-1α, and TNF-α levels in the ApoE-/- group and the model group were significantly higher than those in the normal group (P<0.01). Compared with the model group, each administration group exhibited a significant reduction in aortic NLRP3 and Caspase-1 protein expression and serum IL-1β, IL-18, IL-1α, and TNF-α levels (P<0.05), with the strongest inhibitory effect detected in the medium-dose Didangtang group (P<0.05).Conclusion Didangtang directly alleviates diabetic macrovascular disease possibly by down-regulating NLRP3 and Caspase-1 protein expression and easing the inflammatory cascade.
    2021,27(11):9-18, DOI:
    Abstract:
    Objective To investigate the effect of Buyang Huanwutang (BHT) on proliferation and differentiation in neural stem cells (NSCs) after oxygen-glucose deprivation/reoxygenation (OGD/R) injury.Method NSCs isolated from the hippocampus of SD rats were cultured and randomly divided into a normoxia group, a model group, a BHT group, a rapamycin (Rapa) group, and a combination group [autophagy inhibitor 3-methyladenine (3-MA) combined with BHT]. The 20% blank serum was used in the normoxia group, and 20% BHT-medicated serum in the BHT group. The doses of Rapa and 3-MA were 1 μmol·L-1 and 5 mmol·L-1, respectively. The cells were subjected to OGD/R except those in the normoxia group. The cell morphology was observed under a light microscope. NSCs were confirmed by immunofluorescence detection of nestin expression. The viability and proliferation of NSCs were assessed by cell counting kit-8 (CCK-8) assay and 5-ethynyl-2-deoxyuridine (EdU) labeling, respectively. Furthermore, Ad-mCherry-GFP-LC3B fluorescence assay was performed to investigate autophagy. The effect of BHT on autophagy-related protein expression was detected by western blot assay. Brain derived neurotrophic factor (BDNF), β-tubulin Ⅲ, and glial fibrillary acidic protein (GFAP) were evaluated by immunofluorescence assay.Result OGD/R significantly reduced the cell viability of rat NSCs as compared with the normoxia group. Compared with the model group, the BHT group exhibited significantly improved viability of rat NSCs (P<0.01). BHT induced the production of autophagosomes in NSCs after OGD. The BHT group showed increased expression of microtuble-associated protein 1 light chain 3Ⅱ (LC3Ⅱ) and Beclin-1 (P<0.05,P<0.01) and slightly changed p62 compared with the normoxia group, and significantly up-regulated LC3Ⅱ and Beclin-1 (P<0.05,P<0.01) and down-regulated expression of p62 (P<0.01) compared with the model group. The Rapa group had similar effect as the BHT group (P<0.05,P<0.01), while the combination group inhibited the activity of autophagy (P<0.01). As indicated by the results of ad-mCherry-GFP-LC3B, compared with the normoxia group, the model group showed increased fluorescence intensity (P<0.01), and the BHT and Rapa groups could further increased the fluorescence intensity of autophagy (P<0.01), while the combination group inhibited autophagy activity (P<0.01). Immunofluorescence results revealed that compared with the normoxia group, the model group displayed significantly reduced positive cells of EdU, β-tubulin Ⅲ, GFAP, and BDNF (P<0.01), and the BHT and Rapa groups exerted similar protective and promoting effects (P<0.05,P<0.01), while the combination group partially blocked the neuroprotection and differentiation ability of BHT (P<0.05).Conclusion BHT pretreatment can effectively protect rat NSCs against OGD-induced injury and promoted proliferation and differentiation by up-regulating autophagy.
    2021,27(11):19-26, DOI: 10.13422/j.cnki.syfjx.20211001
    Abstract:
    Objective To study the effect of Gegen Qinliantang (GQT) on the structure of intestinal flora in dysbacterial diarrhea rats by 16S rRNA sequencing.Method Sixty healthy SD rats were randomly and equally divided into a control group, a model group, high-, medium-, and low-dose GQT groups, and a Bifidobiogen group. The rat model was induced in the five groups except the control group by administration of mixed antibiotics (178.6 mg·kg-1 cefradine and 31.25 mg·kg-1 gentamicin sulfate) according to the dose. Drug intervention was carried out in each group (7.02, 3.51, and 1.755 g·kg-1 GQT for the high-, medium-, and low-dose GQT groups, 0.125 g·kg-1 bifidobacterium capsules for the Bifidobiogen group, and sterile distilled water for the control and model groups) with a volume of 10 mL·kg-1 for seven days. Colon contents of rats were obtained under anesthesia. The extracted fecal DNA underwent 16S rRNA high-throughput sequencing and the results were analyzed.Result GQT was proved capable of adjusting the species number and Alpha and Beta diversity, improving the biological richness and diversity of the flora, and positively regulating three differential phyla (Firmicutes, Proteobacteria, and Bacteroidetes) and 14 differential genera (Bacteroides Parabacteroides Blautia, etc.) in rat model of dysbacterial diarrhea.Conclusion The present study confirmed the regulatory effect of GQT on intestinal flora of dysbacterial diarrhea rats, and revealed the physiological and pathological mechanism between intestinal flora and dysbacterial diarrhea.
    2021,27(11):27-34, DOI: 10.13422/j.cnki.syfjx.20211190
    Abstract:
    Objective To explore the effects and mechanism of Chinese classical prescription Dahuang Zhechongwan on silicosis in mice.Method Thirty-six male Kunming mice of SPF grade were randomized into the normal control group, model control group, tetrandrine (Tet, 0.039 mg·kg-1) group, as well as high- (1.560 g·kg-1), medium- (0.780 g·kg-1), and low-dose (0.390 g·kg-1) Dahuang Zhechongwan groups, with six mice in each group. Mice in all groups except for the normal control group underwent static inhalation of silica (SiO2) dust for 40 consecutive days to induce fibrosis. After 28 days of intervention with corresponding drugs, the mice were sacrificed to collect the serum and lung tissues, with the former used for detecting tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and hydroxyproline (HYP) levels by enzyme-linked immunosorbent assay (ELISA) and the latter for observing the pathological changes. Meanwhile, the protein and mRNA expression levels of p38 mitogen-activated protein kinase (p38 MAPK), nuclear transcription factor-κB (NF-κB), transforming growth factor-β1 (TGF-β1), α-smooth muscle actin (α-SMA), Smad2, Smad3, and Smad7 in the lung tissues were determined by Western blot and real-time polymerase chain reaction (Real-time PCR).Result Compared with the normal group, the contents of TNF-α, IL-1β, IL-6 and HYP in the model group were significantly increased, the difference was statistically significant(P<0.05,P<0.01); compared with the model group, the high-dose group of Dahuang Zhechongwan could significantly reduce the contents of TNF-α, IL-6 and HYP in the serum of mice(P<0.05, P<0.01), indicating that Dahuang Zhechongwan could reduce the lung inflammation of silicosis mice. At the same time, compared with the normal group, the protein and mRNA expression levels of p38 MAPK, NF-κB p65, TGF-β1α-SMA, Smad2 and Smad3 in the model group were significantly increased(P<0.05,P<0.01), while the protein and mRNA expression levels of Smad7 were significantly decreased(P<0.01); compared with the model group, the protein and mRNA expression levels of p38 MAPK, NF-κB p65, TGF-β1α-SMA, Smad2 and Smad3 in the high-dose Dahuang Zhechongwan group were significantly increased the protein and mRNA expression levels were significantly decreased(P<0.05,P<0.01), while Smad7 protein and mRNA expression levels were significantly increased(P<0.05,P<0.01).Conclusion Dahuang Zhechongwan ameliorates the alveolar inflammation, extracellular matrix (ECM) deposition, and fibrosis in mice with silicosis possibly by regulating the p38 MAPK/NF-κB/TGF-β1 pathway.
    2021,27(11):35-41, DOI: 10.13422/j.cnki.syfjx.20211193
    Abstract:
    Objective To investigate the molecular mechanism of Qiyu Sanlong prescription (QYSL) in inhibiting the "addiction" of lung cancer A549 cells to miRNA21.Method The human lung cancer A549 cells were routinely passaged and divided into the blank group, blank serum group, QYSL-containing serum group, and siRNA group. The prepared QYSL-containing serum was used for intervention, with the optimal concentration and action time determined in previous studies. The protein and mRNA expression levels of miRNA21 and related molecules in its target phosphatase and tensin homolog deleted on chromosome 10 (PTEN)/phosphatidylinositol 3-kinase (PI3K) signaling pathway were detected by real-time polymerase chain reaction (Real-time PCR) and Western blot assay.Result The comparison with the blank serum group revealed that the mRNA expression levels of miRNA21 in the QYSL-containing serum group and the siRNA group were decreased, while the PTEN mRNA expression in the QYSL-containing serum group was increased, showing significant differences (P<0.01). Compared with the blank serum, the QYSL-containing serum and siRNA significantly down-regulated PI3K and mammalian target of rapamycin (mTOR) mRNA expression (P<0.01), whereas the QYSL-containing serum did not change the mRNA expression of protein kinase B (Akt). The protein expression levels of PTEN in the QYSL-containing serum group and the siRNA group were obviously elevated in contrast to that in the blank serum group (P<0.05). Meanwhile, the protein expression levels of phosphorylated Akt (p-Akt) and phosphorylated mTOR (p-mTOR) evidently declined in the QYSL-containing serum group (P<0.05), but there was no significant reduction in total Akt and mTOR protein expression. The PI3K protein expression was slightly down-regulated, with no statistical significance.Conclusion QYSL inhibits the transcription of miRNA21, increases the expression of PTEN, and reduces the expression of key molecules in PI3K/Akt/mTOR signaling pathway, thus mildly inhibiting the "addiction" of lung cancer cells to oncogenes and blocking their proliferation.
    2021,27(11):42-51, DOI: 10.13422/j.cnki.syfjx.20211006
    Abstract:
    Objective To investigate the therapeutic effect and mechanism of modified Wenjingtang on endometriosis (EM) rats with kidney deficiency and blood stasis.Method Healthy non-pregnant female Sprague-Dawley (SD) rats of SPF grade were randomly divided into the blank group and experimental group. After being modeled via soaking in ice water and subcutaneous injection of epinephrine hydrochloride, the ones in the experimental group were further divided into the sham operation group and EM model group, with the former only undergoing laparotomy and the latter further receiving autologous endometrial transplant for triggering EM. The successfully modeled rats with EM due to kidney deficiency and blood stasis were randomized into the positive drug (danazol, 63 mg·kg-1) group and low- (5 g·kg-1), medium- (10 g·kg-1), and high-dose (20 g·kg-1) modified Wenjingtang groups. The corresponding drugs were administered by gavage, once per day, for four weeks. Then the ectopic and eutopic endometrial tissues were stained with hematoxylin-eosin (HE) to observe the histopathological changes. The protein and mRNA expression levels of cysteinyl aspartate-specific proteinase-8 (Caspase-8), matrix metalloproteinase-9 (MMP-9), N-cadherin, and E-cadherin were detected by immunohistochemistry (IHC), Western blotting, and real-time polymerase chain reaction (Real-time PCR), respectively.Result The IHC and Western blot revealed that the protein expression levels of MMP-9 and N-cadherin in eutopic and ectopic endometrial tissues of the model group were significantly increased as compared with those in the sham operation group (P<0.01), while the levels of Caspase-8 and E-cadherin was significantly decreased (P<0.01). Compared with the model group, the danazol and low-, medium-, and high-dose modified Wenjingtang groups exhibited obviously down-regulated MMP-9 and N-cadherin protein expression in eutopic and ectopic endometrial tissues (P<0.05,P<0.01), but up-regulated Caspase-8 and E-cadherin (P<0.05, P<0.01). Real-time PCR uncovered that the mRNA expression levels of MMP-9 and N-cadherin in eutopic and ectopic endometrial tissues of the model group were significantly elevated as compared with those in the sham operation group (P<0.01), whereas the levels of Caspase-8 and E-cadherin significantly declined (P<0.01). The comparison with the eutopic endometrial tissue in the model group showed that the mRNA expression levels of MMP-9 and N-cadherin in the danazol group and high- and medium-dose modified Wenjingtang groups were significantly down-regulated, while those of Caspase-8 and E-cadherin were significantly up-regulated (P<0.05,P<0.01).Conclusion Modified Wenjingtang alleviates the immunosuppression and blocks the angiogenesis in EM rats with kidney deficiency and blood stasis syndrome by regulating the expression of such cytokines as Caspase-8, MMP-9, N-cadherin, and E-cadherin, thus exerting the therapeutic effect against EM. The above-mentioned micro-indicators are potential markers reflecting the disease (EM), syndrome (kidney deficiency and blood stasis), and pathological mechanisms (immunosuppression and angiogenesis).
    2021,27(11):52-62, DOI: 10.13422/j.cnki.syfjx.20211007
    Abstract:
    Objective To explore the intervention effect of Yuxuebi tablet (YXB) on collagen-induced arthritis (CIA) in rats and its anti-inflammatory mechanism.Method Following CIA modeling, the rats in the drug administration groups were separately treated with intragastric administration of YXB (0.1, 0.2, and 0.4 g·kg-1) and methotrexate (MTX, 0.4 mg·kg-1), once a day. The incidence of CIA, mechanical pain threshold (MPT) and cold pain threshold (CPT) were evaluated once every three days. After continuous administration for 30 days, the peripheral blood of rats was collected for the determination of platelet (PLT) count and fibrinogen (FIB) content. The hematoxylin-eosin (HE) staining was conducted to analyze the pathological changes in joint tissues. The protein expression levels of interleukin (IL)-1β, IL-8, nuclear transcription factor-κB (NF-κB) p65, phosphorylated NF-κB (p-NF-κB) p65, Ras, and Raf-1 in joint tissues of CIA rats were detected by immunohistochemistry (IHC) and Western blot. The rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) were induced by tumor necrosis factor-α (TNF-α, 10 μg·L-1in vitro and then subjected to transwell migration/invasion assay, followed by the detection of protein expression levels of Ras, Raf-1, and p-NF-κB p65 in RA-FLS by Western blot.Result Compared with the control group, the model group exhibited an increased incidence of CIA, significantly decreased MPT (P<0.05,P<0.01), elevated CPT (P<0.01) and PLT and FIB in the peripheral blood, worsened histopathological score of joints, enhanced RA-FLS migration and invasion, and up-regulated inflammatory factors (P<0.01). The comparison with the model group revealed that YXB at different doses obviously reduced the incidence of CIA, increased MPT, down-regulated CPT and PLT and FIB in the peripheral blood (P<0.05,P<0.01), ameliorated the pathological changes like synovial hyperplasia and bone and cartilage destruction (P<0.05,P<0.01), and inhibited RA-FLS migration and invasion. Besides, the low-, medium-, and high-dose YXB reversed the IL-1β, IL-8, Ras, Raf-1, and p-NF-κB p65 expression in joint tissues of CIA rats to different extents, as well as the protein expression of Ras, Raf-1 and p-NF-κB p65 in RA-FLS (P<0.05,P<0.01).Conclusion YXB reduces the incidence of CIA, ameliorates the clinical symptoms of RA and the pathological changes in joint tissues, and inhibits the formation of synovium, which may be attributed to its inhibition against Ras/Raf-1/NF-κB signaling pathway.
    2021,27(11):63-75, DOI: 10.13422/j.cnki.syfjx.20210627
    Abstract:
    Objective To investigate the hepatotoxicity of different doses of geniposide on the liver of rats and the effects on bile acid profile in serum, liver tissue and feces.Method The 60 Sprague Dawley rats, half male and half female, were randomly divided into 5 groups according to body weight: blank group and four different doses (50, 100, 200, 400 mg·kg-1) geniposide groups, 12 rats in each group. The rats were treated by gavage once a day for 7 consecutive days, and the serum, liver and cecal contents were collected on the 8th day of treatment. The activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), the contents of albumin (ALB), total bilirubin (TBIL), total bile acid (TBA), creatinine (Crea) and carbamide (Urea) were detected in each group. The sections of liver tissue were stained with hematoxylin-eosin(HE), and the protein expressions of cytokeratin 7(CK7) and cytokeratin 19(CK19) were detected by immunohistochemistry. The protein expressions of CK7 and CK19 in the liver tissue were detected by Western blot. And the mRNA expressions of cholesterol 7α-hydroxylase (CYP7A1), cholesterol 27α-hydroxylase ( CYP27A1) and cholesterol 12α-hydroxylase (CYP8B1) were detected by real-time PCR. The contents of 18 kinds of bile acids in serum, liver and cecal contents were determined by ultra-performance liquid chromatography-mass spectrometry(UPLC-MS).Result Compared with the control group, TBIL level in each dose of geniposide group was increasesd significantly (P<0.01). ALT, AST activity and TBA content in 400 mg·kg-1 geniposide group were increased significantly (P<0.05, P<0.01). HE staining showed that, compared with control group, there was bile duct reaction in the portal area and inflammatory cells infiltrate around bile duct in 200 mg·kg-1 and 400 mg·kg-1 geniposide groups, especially 400 mg·kg-1. The expressions of CK7 and CK19 in liver tissue of 400 mg·kg-1 geniposide group were significantly higher than those in the control group (P<0.05, P<0.01). Compared with the control group, the contents of glycoursodeoxycholic acid (GUDCA) and glycohyodeoxycholic acid (GHDCA) in liver tissue of 400 mg·kg-1 geniposide group decreased significantly (P<0.05, P<0.01), the contents of sodium taurochenodeoxycholate (TCDCA), hyodeoxycholic acid (HDCA), cholic acid (CA) and chenodeoxycholic acid (CDCA) in liver tissue increased significantly (P<0.01), the contents of glycocholic acid hydrate (GCA), glycochenodeoxycholic acid (GCDCA), glycodeoxycholic acid hydrate (GDCA), glycocholic acid (GLCA), tauroursodeoxycholic acid (TUDCA), GUDCA, GHDCA, ursodeoxycholic (UDCA) and taurolithocholic acid (TLCA) decreased, the proportions of TCDCA, HDCA, CA, CDCA and deoxycholic acid (DCA) in liver tissue increased, the contents of GHDCA and lithocholic acid (LCA) in serum decreased significantly (P<0.01), while sodium taurohyodeoxycholate hydrate (THDCA), taurocholic acid (TCA), GCA, TCDCA, UDCA, CA, CDCA, DCA in serum decreased significantly (P<0.05, P<0.01). The contents of CA, UDCA, CA, CDCA and DCA increased significantly (P<0.05), the ratio of CA/DCA increased significantly (P<0.05), and the ratio of CA and CDCA increased by 19.60% and 4.63%, respectively; Compared with the control group, the contents of all bile acids in cecal contents of 400 mg·kg-1 were decreased, and the contents of GCA, UDCA, HDCA, GCDCA, GDCA, TLCA, GLCA, CDCA, DCA and LCA were decreased significantly (P<0.05, P<0.01). In addition, real-time PCR results showed that the mRNA expressions of CYP7A1, CYP27A1 in the 400 mg·kg-1 geniposide group were significantly higher than those in the control group (P<0.05, P<0.01).Conclusion The 400 mg·kg-1 geniposide can cause obvious hepatotoxicity in rats, and the bile acid profile in liver, serum and excrement changes significantly, and the changes of the each bile acid in liver, serum and feces are different. However, the causal relationship between the gardenoside-induced liver injury and the changes in bile acid profile are not clear. It needs to be further studied.
    2021,27(11):76-82, DOI: 10.13422/j.cnki.syfjx.20211192
    Abstract:
    Objective To detect the toxicity of water-eluted fraction from Siegesbeckiae Herba (SWEF) at different concentrations against MRC-5 human embryonic lung fibroblasts and its impacts on the expression of α7 nicotinic acetylcholine receptor (α7nAChR) and inflammatory factors, so as to figure out the active components responsible for toxicity and efficacy.Method The toxicities of SWEF at 1, 6, 10, 20, and 50 g·L-1 against MRC-5 cells were determined by cell counting kit-8 (CCK-8) assay combined with flow cytometry and Trypan blue staining. The changes in α7nAChR expression and inflammatory factor levels before and after α7nAChR gene silencing were detected to reveal the pharmacodynamic effect of SWEF on MRC-5 cells.Result SWEF (≥6 g·L-1) obviously inhibited the viability of MRC-5 cells (P<0.01) and promoted their apoptosis and necrosis (P<0.01), with the half-maximal inhibitory concentration (IC50) being 6.03 g·L-1. The determination of α7nAChR expression and inflammatory factor levels in MRC-5 cells showed that SWEF contained α7nAChR agonist-like substance, which enhanced α7nAChR mRNA and protein expression (P<0.05, P<0.01) and decreased the inflammatory factor levels (P<0.05, P<0.01). SWEF down-regulated the inflammatory factors possibly by re-regulating α7nAChR mRNA expression, exhibiting a negative correlation between them (P<0.01).Conclusion SWEF (≥6 g·L-1) is highly toxic to MRC-5 cells. Pharmacodynamic studies have confirmed that α7nAChR agonist-like substance contained in SWEF was responsible for the elevated α7nAChR expression and reduced inflammatory cytokines. It is inferred that excessive α7nAChR agonist-like substance may trigger the toxicity of Siegesbeckiae Herba.
    2021,27(11):83-88, DOI:
    Abstract:
    Objective To investigate the effect and mechanism of saikosaponin A (SSA) on the reversal of cisplatin (DDP) resistance in human lung cancer cell line A549/DDP.Methods The resistance of A549 and A549/DDP cells to DDP and the inhibitory effects of SSA against the proliferation of A549 and A549/DDP cells were detected using cell counting kit-8 (CCK-8). The apoptosis rates of A549/DDP cells treated with SSA or DDP or SSA combined with DDP and the changes in reactive oxygen species (ROS) were determined by flow cytometry. The mRNA expression levels of C-myc, B-cell lymphoma 2 (Bcl-2) and cysteinyl aspartate-specific protease-3 (Caspase-3) were detected by real-time polymerase chain reaction (Real-time PCR), followed by the determination of β-catenin transcriptional activity using the TopFish dual-luciferase reporter assay system and the measurement of β-catenin protein expression in A549/DDP cells by Western blot.Results The results of CCK-8 assay showed that the DDP resistance of A549/DDP cells was 12.82 times that of A549 cells (P<0.05). SSA inhibited the viability of A549 cells with the half maximal inhibitory concentration (IC50) being 34.9 μmol·L-1, and also suppressed the viability of A549/DDP cells in a concentration-dependent manner. Since the inhibition rate of 20 μmol/L SSA against A549/DDP cells was less than 10%, the reversal concentration was set at 20 μmol/L. Flow cytometry revealed that compared with the control, DDP alone increased the apoptosis rate of A549/DDP cells (P<0.05), stimulated the accumulation of intracellular ROS (P<0.05), down-regulated the mRNA expression levels of C-myc and Bcl-2 in A549/DDP cells, up-regulated Caspase-3 mRNA expression, and reduced the transcriptional activity of β-catenin (P<0.05). Compared with the DDP group, the SSA+DDP group exhibited obviously increased apoptosis of A549/DDP cells, enhanced accumulation of intracellular ROS, down-regulated C-myc and Bcl-2 mRNA expression, up-regulated Caspase-3 mRNA expression (P<0.05), and weakened β-catenin transcription (P<0.05). DDP combined with SSA better decreased the β-catenin protein expression in contrast to that of control or DDP (P<0.05).Conclusions SSA enhances the sensitivity of A549/DDP cells to DDP possibly by inhibiting the activation of Wnt/β-catenin pathway.
    2021,27(11):89-97, DOI: 10.13422/j.cnki.syfjx.20211004
    Abstract:
    Objective To observe the effect of Yiqiyangyin Huoxuetongluo prescription on janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway and cell apoptosis in rats with diabetic nephropathy (DN), and to explore the mechanism of its intervention in DN.Method A total of 100 SD rats were randomly divided into an experimental group (n=80) and a normal group (n=20). The DN model was induced by high-sugar and high-fat diet combined with intraperitoneal injection of streptozotocin (STZ) in the experimental group, and confirmed by the pathological changes of kidney tissues in rats (three in each group) observed under light and electron microscopes. The model rats were randomly divided into a model group (normal saline, equal volume), low-, medium-, and high-dose (5.775, 11.550, and 23.100 g·kg-1) Yiqiyangyin Huoxuetongluo prescription groups, and an irbesartan group (irbesartan tablets, 0.016 g·kg-1). After drug intervention (i.g., once a day for 16 consecutive weeks), the 24-hour urine total protein (UTP), serum total cholesterol (TC), triglyceride (TG), creatinine (SCr), and blood urea nitrogen (BUN) levels of the rats were measured. Western blot was used to detect the protein expression of JAK2/STAT3 signaling pathway. Immunohistochemistry was used to determine the protein expression of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), zonula occludens-1 (ZO-1), and actinin-4 in rat kidney tissues.Result Compared with the normal group, the model group exhibited elevated UTP, serum TC, TG, BUN, and SCr levels (P<0.05), severe pathological damage of rat kidney tissues, up-regulated expression of phospho-JAK2 (p-JAK2), phospho-STAT3 (p-STAT3), and Bax, increased renal cell apoptosis, and diminished expression of Bcl-2, ZO-1, and actinin-4 (P<0.05). Compared with the model group, the Yiqiyangyin Huoxuetongluo prescription group and the irbesartan group showed dwindled UTP, serum TC, TG, BUN, and SCr levels (P<0.05), relieved pathological damage of rat kidney tissues, down-regulated p-JAK2, p-STAT3, and Bax expression, and up-regulated expression of Bcl-2, ZO-1, and actinin-4 (P<0.05).Conclusion Yiqiyangyin Huoxuetongluo prescription can reduce renal cell apoptosis and improve the prognosis of DN by inhibiting the activation of JAK2/STAT3 signaling pathway.
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    2013,19(20):246-250, DOI: 10.11653/syfj2013200246
    [Abstract] (6526) [HTML] (83) [PDF 854.77 K] (15572)
    Abstract:
    Objective: To investigate the intervention effect and potential mechanism of Qingre Huashi decoction(QHD) for preventing the onset of colon cancer by observing SGC-7901 cell proliferation and apoptosis along with the expression of CD14 gene, tumor necrosis factor alpha and interleukin 1 beta. Method: SGC-7901 cells were divided into control group, intervention group. In vitro, the control group was offered fetal calf serum. The intervention group was fed with serum containing QHD for 5%,10%,15%,20%,30%. The multiplication, proliferation and apoptosis of SGC-7901 cell were detected by MTT and flow cytometry respectively. The expression of CD14, TNF-α and IL-1β gene was assayed by immunohistochemistry and real-time PCR. Result: Compared with control group, the survival of SGC-7901 cell was decreased in QHD group(P<0.05). In QHD group, the cells in G0/G1 phase were increased but the S phase cell was on the decline. Those changes were correlated with the culturing content. There is a significant difference between QHD group and control group in those changes as mentioned above(P<0.05). Especially, the effect of middle concentration serum containing QHD on inducing cell apoptosis and down regulating the expression of CD14, TNF-α, IL-1β was obvious (P<0.05). Conclusion: QHD exerts a distinct effect on preventing the onset of colon cancer by mediating SGC-7901 cell S time, cell composition and regulating the expression of CD14, TNF-α and IL-1β and inducing cell apoptosis.
    2014,20(5):54-56, DOI: 10.11653/syfj2014050054
    [Abstract] (7711) [HTML] (67) [PDF 406.88 K] (9559)
    Abstract:
    Objective: To establish a high performance liquid chromatographic method (HPLC) for simultaneous determination of rubrofusarin gentiobioside, casside, cassiaside C in Cassia obtusifolia. Method: The HPLC with Dionex Acclaim C18(4.6 mm×250 mm, 5 μm) column was used, acetonitrile-THF-1% acetic acid (17 :2 :81) was used as a mobile phase, with flow rate of 1 mL·min-1, column temperature at 30 ℃ and detection wavelength at 278 nm. Result: Rubrofusarin gentiobioside, casside, cassiaside C showed good linearity (r=0.999 9) in the range of 0.218-1.09, 0.188-0.94, 0.200-1.000 μg, regressive equation were Y=54.017X-0.344 6 (r=0.999 9),Y=78.63X-0.354(r=0.999 9),Y=74.098X-0.433 5 (r=0.999 9)respectively;the average recoveries of the method were 98.43%, 101.89%, 102.46%, RSD were 1.75%, 1.10%, 1.15%. Conclusion: The method was proved to be simple, rapid, sensitive, precise, reliable and repeatable. It can be applied to the quality control of Semen Cassia.
    2015,21(11):7-10, DOI: 10.13422/j.cnki.syfjx.2015110007
    [Abstract] (5614) [HTML] (92) [PDF 534.15 K] (9505)
    Abstract:
    Objective: To investigate correlation between content and anti-proliferative effect against lung carcinoma cell line SPC-A-1 and A549 of active ingredients in Coicis Semen from five places. Method: HPLC-ELSD and UV spectrophotometry were employed to determine contents of glycerol trioleate and total triglycerides in Coicis Semen,respectively.In addition,MTT assay was performed to explore anti-proliferative effect of Coicis Semen extract from various origins towards SPC-A-1 and A549 cells. Result: The content of glycerol trioleate in Coicis Semen from Hebei,Liaoning,Fujian,Guizhou,Shandong province was 0.91%,1.14%,0.82%,0.77%,1.09%,the content of total triglycerides was 14.83,10.88,7.85,5.26,7.71 mg·g-1,respectively.IC50(median inhibitory concentration) against the lung carcinoma cell line A549 were 2.33,2.77,3.15,3.24,3.08 g·L-1,IC50 against SPC-A-1 were 1.64,2.15,2.63,3.01,2.99 g·L-1,respectively. Conclusion: All samples originated from five provinces are in line with standards of the 2010 edition of Chinese Pharmacopoeia.However,samples from Hebei province have the highest content of total triglycerides and the strongest anti-lung cancer activity,suggesting a positive correlation between active ingredients content and pharmacological activity.
    2014,20(9):15-18, DOI: 10.13422/j.cnki.syfix.2014090015
    [Abstract] (7196) [HTML] (82) [PDF 1.24 M] (8755)
    Abstract:
    Objective: To optimize extraction technology of Jiedu Tongluo particles. Method: Based on single factor tests, central composite design-response surface methodology was applied to optimize extraction process with soaking time, decoction time and solvent amounts as independent variables, while composite score of extraction amounts of baicalin, quercetin and isorhamnetin as dependent variable.SPSS software was used to fit multivariate linear equation and second-order polynomial equation for experimental data, response surface and contour plot were delineated according to best-fit mathematic models by Design Expert 8.0.0 statistic analysis software. Result: Optimum extraction process was as follows:soaking time 90 min, decoction time 85.63 min, the amount of solvent 9.21 times;Composite score was (93.2±0.051) %, which had deviation of -2.32% with the predicted value of 94.30%. Conclusion: Model established by central composite design-response surface methodology had good predictability, which was suitable for optimizing extraction process of Jiedu Tongluo particles.
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    [Abstract] (10354) [HTML] (103) [PDF 0.00 Byte] (6808)
    Abstract:
    【Abstract】Objective To study the mechanism of the chinese medicine Weichangshu on mitochondrial kinetic energy in diabetic mellitus rats stomach by measuring the changes of morphology, structure, function and protein of mitochondria. Methods: Standard Sprague Dawley rats with a mean weight of 150-170g were randomly divided into 4 groups, including a control group, DM model group, DM WeichangshuⅠgroup, DM Weichangshu Ⅱ group. The mitochondrial morphology and structure were detected using transmission electron microscopy. The changes of mitochondrial transmembrane potential, respiratory chain and mitochondrial protein were detected using flow cytometry, fluorescence labeling and Western Blot respectively. Results: Weichangshu could ameliorate the mitochondrial morphology, structure and respiratory chain in diabetic mellitus rats stomach. Mitochondrial transmembrane potential, ATP level and activity of cytochrome C oxidase were increased. The release of ROS was decreased. The expression of Bcl-2 up-regulated and bax down-regulated. Conclusion: Weichangshu could promoted the gastrointestinal motility, which might be related with the regulation of mitochondrial kinetic energy.
    2012,18(24):39-41, DOI:
    [Abstract] (8576) [HTML] (135) [PDF 419.69 K] (5583)
    Abstract:
    Objective:To optimize matrix prescription of Guyanxiao cataplasm. Method: Matrix prescription was optimized by orthogonal test with initial viscosity,sustained adhesive strength and general sensory as comprehensive evaluation index,the amount of NP700,CMC-Na, glycerol and titania were chosen as factors. Result: Optimal matrix prescription was:NP-700, CMC-Na, glycerol and titania of 3,0.6,16,0.8g,respectively. Conclusion: Guyanxiao catapasm with good appearance and adhesion could be prepared by optimized matrix formulation.
    2013,19(8):46-50, DOI: 10.11653/syfj2013080046
    [Abstract] (1644) [HTML] (50) [PDF 1.16 M] (5323)
    Abstract:
    Objective: To optimize formulation of ursolic acid self-microemulsion drug delivery system using central composite design/response surface methodology. Method: With ratio of surfactants and cosurfactants and oil content as independent variables,particle size and saturated drug doading were taken as dependent variables,optimum formulation was screened by SAS9.1.3 software,and verified. Result: Optimized formulation of ursolic acid self-microemulsion was oil phase(MCT)-emulsifier(HS15)-cosurfactants(alcohol) 12.5:62.5:25. Conclusion: This optimized formulation of ursolic acid self-microemulsion drug delivery system could be obtained exactly and conveniently by central composite design response surface methodology,and established model had a reliable predictability.
    2014,20(8):15-18, DOI: 10.13422/j.cnki.syfix.2014080015
    [Abstract] (5341) [HTML] (92) [PDF 981.94 K] (5104)
    Abstract:
    Objective: To optimize ultrafine grinding process of Chuanxiong Rhizoma. Method: Taking feeding amount, moisture content and grinding time as independent variables, average particle size as dependent variable, Box-Behnken response surface methodology was adopted to optimizeultrafine grinding process of Chuanxiong Rhizoma, its powder properties and hygroscopicity before and after smashed were investigated. Result: Optimum ultrafine grinding process was as follows:feeding amount1.3 kg, moisture content 4%, grinding time 19 min;Deviation between the predicted value(18.357 7 μm) and the measured value(19.187 μm) of average particle size was only 4.5%.Response angles of ordinary and ultrafine powder were (49.4±0.03) and (50.2±0.02) °, bulk densities were (0.429±0.003) and (0.363±0.005) g·mL-1, tap densities were (0.609±0.05) amd (0.556±0.10) g·mL-1, moisture equilibrium times were about 80 h. Conclusion: Optimized ultrafine grinding process was reasonable and feasible, preparation technology of ultrafine powder of Chuanxiong Rhizoma should add flow aid and other auxiliary materials in order to improve fluidity of powder, ultrafine grinding process should strictly control humidity.
    2012,18(24):24-27, DOI:
    [Abstract] (5279) [HTML] (64) [PDF 525.29 K] (4748)
    Abstract:
    Objective:To optimize matrix formulation of Sanhuang Sanyu cataplasm. Method: Taking appearance and adhesion of cataplasm as evaluation indexes,matrix formulation of Sanhuang Sanyu cataplasm was optimized by single factor test and orthogonal test. Result: Optimized matrix formulation of Sanhuang Sanyu cataplasm was NP-700-K90-glycerol(0.5∶0.4∶6).0.02 g·mL-1 solution was received by adding carbomer-u10NF into mixed extraction concentrated liquid of Rheum palmatum and Scutellaria baicalensis,swelling overnight, as A phase,0.5 g sodium polyacrylate and 0.4 g PVP were added to 6 g glycerol as B phase,aluminum chloride and citric acid were added to water as C phase,single extraction concentrated liquid of Phellodendron chinense was as D phase,four phases was fully mixed with A-B-C-D(10∶3∶3∶5), stirred 15 min, coating,covered with anti-sticking layer. Conclusion: Optimized matrix formulation of Sanhuang Sanyu cataplasm was simple,stable and feasible,which could be applied to industrial production.
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    [Abstract] (3278) [HTML] (59) [PDF 0.00 Byte] (4673)
    Abstract:
    Fracture healing is a process that involves the proliferation and differentiation of cells and tissues and can be stimulated by drugs. Traditional medicine has a reputation in the therapy on bone fracture. It is believed that herbal formulae promotes bone repair through multiple functions of “anti-inflammation”, “promoting blood circulation” and “stimulating bone formation”. This study aims to engineer a herbal formula under the guidance of Chinese medicine theories, then define the effective dosage ratio between the herbs and demonstrate the biological effects of the formulae on different parameters of healing using relevant in vitro assays. Six herbs which are frequently used for fractures were combined into five formulae according to the “Uniform Design” method. Assays of nitric oxide (NO) inhibition in mouse macrophages, human umbilical vein endothelial cells (HUVEC) and rat osteoblasts proliferation were used to reflect the “anti-inflammation inflammatory”, “circulation promotion” and “bone formation stimulation” functions. The results showed that all the formulae expressed significant activities of NO inhibition, and HUVEC cell and osteoblasts proliferation. Formula 5 with the largest doses (containing 30g Radix et Rhizoma Rhei, 9g Fructus Gardeniae, 9g Radix et Rhizoma Notoginseng, 9g Flos Carthami, 30g Ramulus Sambucus Willamsii, 15g Radix Dipsaci) demonstrated the best results in all the three assays.
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    [Abstract] (2842) [HTML] (69) [PDF 0.00 Byte] (4635)
    Abstract:
    Abstract:Objective: to investigate the absorption mechanism of palmatine chloride by using Caco-2 monolayer mode1.Methods: Caco-2 cell monolayer model was used to investigate the bi-directional transport of palmatine chloride. The effect of time, drug concentration, temperature, pH and inhibitors on the absorption of palmatine chloride was studied. Drug concentration was measured by HPLC and the apparent permeability coefficients (Papp) were calculated. Results: The apical to basolateral permeability (Papp) of palmatine chloride was bigger than the basolateral to apical permeability(Papp).Transport of palmatine chloride was time and concentration dependent, and it was also effected by P-glycoprotein inhibitor, pH and temperature. Conclusion:The absorption of palmatine chloride in Caco-2 cell model was an active transportation mediated by transporter, which mainly located in the apical side of Caco-2 cell monolayer.
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    [Abstract] (5716) [HTML] (61) [PDF 0.00 Byte] (4566)
    Abstract:
    Abstract:Objective To observe the effect of Dendrobium with different doses on ALT, AST, TC in the serum of chronic alcoholic liver injury mice. Methods Male ICR mice were divided into nine groups randomly: normal control group, model control group, positive control group, fresh Dendrobium groups with three doses(3g/kg,6g/kg,9g/kg), dried Dendrobium groups with three doses(0.45g/kg,0.90g/kg,1.35g/kg), gastric infusion for 65 days. The mice mode1 of chronic alcoholic 1iver injury was established by filling the stomach with alcohol after the administration for 65 days, once a day. The blood was taken to separate into serum and the activity of ALT, AST and the content of TC in the serum were determined. Results Compared with the normal control group, the ALT, AST, TC in the serum of model control group increased. Compared with model control group, fresh Dendrobium (3g/kg) and dried Dendrobium (0.45g/kg) could decrease ALT in the serum;fresh Dendrobium (3g/kg, 6g/kg) and dried Dendrobium (0.45g/kg) could decrease AST, TC in the serum; dried Dendrobium (0.90g/kg, 1.35g/kg) could decrease AST in the serum. Conclusion Fresh and dried Dendrobium could improve effectively the activity of ALT, AST and decrease the content of TC in the serum of chronic alcoholic 1iver injury mice.
    2014,20(23), DOI:
    Abstract:
    Objective: To establish HPLC fingerprint of Tibetan medicine Pterocephali Herba, and compare it of Pterocephali Herba from different place, provided a new methods for its quality evaluation. Method: Adoption the method of HPLC to determine 10 groups of Pterocephali Herba from different place. Chromatographic Column was Boston Symmetrix(250×4.6mm,5μm), mobile phase was Methanol-0.1 mol.L-1 ammonium acetate solution(81:19),velocity was 0.8mL.min-1, the detection wavelength was 210nm, column temperature was 30℃. And compared the similarity of Pterocephali Herba from different place. Result: Taking advantage of < USP fingerprint similarity evaluation system (2004A version)> to establish common mode of HPLC fingerprint of Tibetan medicine Pterocephali Herba, and marked 101 characteristic peaks. The 10 groups samples had high similarity. Conclusion: This method had good repeatability, precision, and stability, providing scientific basis for quality control of Pterocephali Herba.
    2014,20(23), DOI:
    Abstract:
    Objective To establish a sensitive and specific HPLC method for quality control of Ilex hainanensis Merr..Methods HPLC method was applied for quality assessment of Ilex hainanensis Merr.. HPLC analysis was performed on a HanbonC18 column (250mm×4.6mm,5 m).The mobile phase consisted of acetonitrile (solvent B) and watercontaining0.1% (v/v) phosphoric acid (solvent A) at a constant flow rate of 1.0ml/min.An increasing linear gradient (v/v) of solvent B was used. The injection volume was 10μl. The column temperature was set at 25℃. The chromatogram s were monitored at 330nm. Results In different varieties of processing methods of Ilex hainanensis Merr. showing 9 characteristic peaks. It was established from its prepared samples of Ilex hainanensis Merr..Conclusion The characteristic chromatogram of Ilex hainanensis Merr. with high specificity can be used to control its quality as well as the consistency of processing products.
    2014,20(23), DOI:
    Abstract:
    Objective: To establish TLC fingerprint of antibacterial extraction of Pilea aquarum Dunn. Methods: The antibacterial activity extraction of Pilea aquarum Dunn was confirmed by antibacterial activity research, and then the TLC fingerprint of 10 batches of Pilea aquarum Dunn from Guizhou was analyzed comparatively. Results: The ethyl acetate extraction of Pilea aquarum Dunn is antibacterial activity. Another, the TLC chromatogram of 10 groups of drugs were very similar, which had 8 spots and 8 corresponding peaks, but the intensity of each peak was different. Conclusion: The method is simple and stabilized, and can be used for the quality control of Pilea aquarum Dunn.
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    [Abstract] (2849) [HTML] (45) [PDF 0.00 Byte] (4017)
    Abstract:
    Objecive To discuss the relationship between water-soluble sugar and cold-heat properties and furtherly reveal correlations of four properties and material basis.Methods Ten cold herbs and heat herbs were selected for the determination,and dealed with the water-soluble sugar with hydroxylamine hydrochloride,pyridine and acetic anhydride.Measuring Gas chomatography-mass(GC-MS) fingerprints of water-soluble sugar,then determining the water-soluble components of 20 herbs by mass spectrometry and analyzing the results by Fisher method.Results A discriminant function had been established by Fisher.The Fisher discriminant analysis could distinguish the cold and heat properties about 20 herbs that the group back substitution is 100%.From the results,it confirmes that there is difference between the cold and heat properties in the GC-MS fingerprints by Fisher method,which shows that there are different water-soluble sugar in the cold and the heat herbs.Conclusion The Fisher analysis could differentiate the cold and heat properties of herbs,and it is an effective statistical tool to study the relationship between the cold and heat properties and water-soluble sugar.It concludes that the water-soluble sugar is one of the material basis of four properties of herbs.

Periodical information

    ISSN:1005-9903
    CN:11-3495/R
    Editor in chief:
    Proprieter:Tong Yan
    Impact factor:
    Publish house:Chinese Journal of Experimental Traditional Medical Formulae

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