OBJECTIVE: To explore the protective effect of palmitic acid against glutamate-induced apoptosis in PC12 cells. METHODS: PC12 cells were randomly divided into six groups: normal group, solvent group(DMSO 0.1%), model group (Glu 25 mmol · L^-1), nimodipine(NMP, 7 μmol · L^-1)-treated group or MK801 (10 μmol · L^-1) -treated group set as positive control group, palmitic acid groups of 100, 10, 1 μmol · L^-1 dose. Cell viability and membrane permeability were examined by MTT assay and LDH kit. Morphologic change of apoptosis were detected by AO/EB and Hoechst staining with fluorescence microscope. The apoptosis rates were measured by flow cytometry using fluorescein labeling FITC-Annexin V/PI. The expression of Bcl-2, Bax and GluR1 protein was assayed by Western blot methods. RESULTS: Compared with the model group, the results showed that palmitic acid could significantly improve the viability of PC12 cells damaged by glutamate, and reduce the permeability of cell membranes. After AO/EB staining, nuclear chromatin of the cells in model group were stained strongly into orange by EB. In palmitic acid - treated group, only few cells were stained into orange. Compared with model group, after the treatment of palmitic acid, slight blue fluroscence stained by Hoechst in nucleus had been emitted. Palmitic acid could increase the expression of Bcl-2 and inhibit the level of the expression of Bax protein and GluR1 protein. CONCLUSION: In summary, palmitic acid had protective effect on glutamate-induced apoptosis in PC12 cells through increasing the expression of Bcl-2 and inhibiting the expression of Bax and GluR1.

The relationship between the conformation of interferon-α (IFN-α) and its anti-viral activity were analyzed by circular dichroism (CD) and flow cytometry (FCM) techniques. The recombinant human IFN-α (rIFN-α2b and rIFN-α2a) were used. CD spectra from 190 nm to 240 nm indicated that two the IFN-α showed stable secondary structure at 65 degrees C, but unstable when the temperature was above 65 degrees C, and the change was irreversible. FCM data of the anti-viral activity of IFN-α indicated that the change of its secondary structures partly weakened its anti-viral activity. The rIFN-α2b and rIFN-α2a showed the same phenomenon. These data indicated that the conformation of IFN-α is one of the factors to influence its anti-viral activity and the combination of CD and FCM is a good method to analyze the relationship between the conformation of protein drugs and their biological activities in single cell level.
Antiviral Agents ; chemistry ; Circular Dichroism ; Flow Cytometry ; Humans ; Interferon-alpha ; chemistry ; Protein Structure, Secondary ; Recombinant Proteins ; chemistry

Objective: To investigate the chemical constituents of the whole plants of Artemisia rupestris. Methods: The chemical constituents were isolated and purified by silica gel, Sephadex LH-20 column chromatography and semi-preparative HPLC. The structures of the isolated compounds were identified by NMR and MS spectroscopic method. Results: Eighteen compounds were isolated and purified, which structures were identified as 2α-(2',4'-hexadiynoyl)-1,6-dioxaspiro [4,5]-deca-3-ene (1), chrysophanol (2), ethyl vanillate (3), methyl linolenate (4), 8,11-octadecadienoic acid methyl ester (5), eicosane coumaric acid ester (6), emodin (7), 2,5-di-tert-butylphenol (8), phellopterin (9), syringic acid (10), 7-methoxycoumarin (11), vanillin (12), imperatorin (13), oroxylin A (14), 5-hydroxy-3,4',6,7-tetramethoxyflavone (15), erucylamide (16), octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl) propanoate (17) and 3,5,3',4'-tetrahydroxy-6,7-dimethoxyflavone (18). Conclusion: Compounds 1, 4, 11-14 and 18 are isolated from A. rupestris for the first time, and all the other compounds were isolated from the genus Artemisia for the first time.

The aim of this paper was to observe the effect of gambogenic acid on angiogenesis of lung cancer and its preliminary mechanism. After culturing lung adenocarcinoma A549 cells, the conditioned medium was treated with gambogenic acid and then used to culture human umbilical vein endothelial cells (HUVECs) to establish the indirect contact cell co-culture system. A two-dimensional culture model of HUVEC was established with matrigel to observe the effect of gambogenic acid on angiogenesis. DAPI staining was used to observe the morphological changes in HUVEC cells after treatment with gambogenic acid under the fluorescence microscope. Annexin V-FITC/PI staining and flow cytometry analysis were used to determine gambogenic acid's effect on HUVEC cell apoptosis rate. The protein expressions of PI3K, p-PI3K, Akt, p-Akt were measured by Western blot. PTEN-siRNA was transfected into cells, and RT-PCR was used to detect the expression levels of PI3K and Akt genes. Gambogenic acid can significantly inhibit angiogenesis, and its inhibitory effect was dose-dependent. DAPI staining showed apoptotic morphological features of HUVEC cells under fluorescence microscope. Annexin V-FITC/PI staining showed that gambogenic acid induced apoptosis in HUVECs. The results of Western blot showed that the expressions of p-PI3K and p-Akt protein were down-regulated with gambogenic acid, while the expressions of PI3K and Akt protein was insignificant. The results of RT-PCR indicated that the expressions of PI3K and Akt protein were up-regulated by PTEN siRNA. Gambogenic acid can inhibit angiogenesis in lung cancer in vitro, and the mechanism of inhibiting angiogenesis may be related to the PI3K/Akt signaling pathway.
A549 Cells ; Apoptosis ; Coculture Techniques ; Human Umbilical Vein Endothelial Cells ; drug effects ; Humans ; Lung Neoplasms ; drug therapy ; pathology ; Neovascularization, Pathologic ; pathology ; PTEN Phosphohydrolase ; genetics ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Transfection ; Xanthenes ; pharmacology

AIM: To investigate the protective mechanism of gastrodin against hydrogen peroxide (H

Manipulations are essential to the acupuncture curative effect. With the technique limited, the researches on acupuncture manipulations can only be carried out with respect to the senses and experience of doctors and patients, thus seriously impeding the modernization and internationalization of acupuncture. Both the modern integration technology of transducer and the biomechanical principles are applied to develop a detection system that can measure the interaction force between the manipulator and the manipulated patient on the needle at each kind of manipulation. Through clinical practice, the force waveforms acting on needle with the manipulations of symmetrical twirling-rotating and symmetrical lifting-inserting were recorded so as to realize the quantitative, objective and real-time detection of the force during the acupuncture process, which provided a new experimentation means and analysis method for the improvement of clinical curative effect and quantitative research of acupuncture and meridians.
Acupuncture ; instrumentation ; Biosensing Techniques ; instrumentation ; Equipment Design ; Humans ; Mechanics ; Medicine, Chinese Traditional ; Needles ; Software ; Software Design

Objective: To explore the effects of minimally invasive intracranial hematoma removal in the treatment of cerebral hemorrhage, and its influence on neurological functional recovery, serum levels of high-sensitivity C-reactive protein (hs-CRP), interleukin-6 (IL-6), interleukin-8(IL-8), tumor necrosis factor-alpha (TNF-α). Methods: From January 2016 to December 2017, 100 patients with cerebral hemorrhage admitted to Zhejiang Xin'an International Hospital were selected and randomly divided into two groups according to the digital table, with 50 cases in each group.The control group was given routine symptomatic treatment, the observation group received minimally invasive intracranial hematoma removal combined with conventional treatment.The curative effect, restoration of nerve function, the levels of hs-CRP, IL-6, IL-8 and TNF- were observed in the two groups. Results: After treatment, the blood loss and edema volume around the hematoma in the two groups were declined significantly (t=5.74, 9.32, 7.41, 9.32, all P<0.05), and the improvements of the observation group was better than those of the control group (t=8.29, 5.28, all P<0.05). The excellent and good rate of the observation group was 90%(45/50), which was significantly higher than 72%(36/50) of the control group (χ²=3.62, P<0.05). After treatment, the NDS scores of the two group were significantly lower than those before treatment (t=4.64, 5.75, all P<0.05), the GCS scores of the two groups were significantly improved (t=5.41, 7.86, all P<0.05). The NDS score of the observation group was significantly lower than that of the control group (t=5.31, P<0.05), the GCS score of the observation group was significantly higher than that of the control group(t=3.84, P<0.05). After treatment, the levels of inflammatory factors in the two groups were significantly reduced compared with those before treatment (t=3.27, 3.75, 3.38, 3.61, 5.74, 4.39, 6.52, 8.26, all P<0.05), the levels of inflammatory factors in the observation group were significantly lower than those in the control group (t=4.37, 3.92, 8.52, 4.28, all P<0.05). Conclusion Minimally invasive removal of intracranial hematoma combined with conventional treatment in the treatment of patients with cerebral hemorrhage can obtain satisfactory clinical effect, can promote neural functional recovery, improve inflammatory factor levels (hs-CRP, IL-6, IL-8, TNF alpha), it is worthy of application.

AIM: To investigate the effects and protective mechanism of Tianma Gouteng (TGY) against rotenone (Rot) damage in PC12 cells. METHODS: PC12 cells were treated with Rot to establish nerve injury model and cell survival rate was determined by MTT colorimetry to determine the optimal modeling concentration of Rot and effective intervention concentration of TGY. Lipid reactive oxygen species (ROS) were detected by flow cytometry and fluorescence intensity was observed by inverted fluorescence microscope. The biochemical methods were used to detect, superoxide dismutase (SOD), glutathione (GSH) levels and activity and the contents of malondialdehyde (MDA). Transmission electron microscopy observed morphological change of mitochondria and protein expression of glutathione peroxidase 4 (GPX4), long chain lipoyl-coa synthase 4 (ACSL4) and lysophospholipid cholinyltransferase 3 (LPCAT3) were detected by western blot. RESULTS: The survival rate of cells treated with 0.6 μmol/L Rot for 24 h was close to 50%(56.7%±9.9%). Pretreatment with TGY for 12 h could inhibit the damage of Rot. At the same time, the leakage rate of lactate dehydrogenase (LDH) was reduced in a dose-dependent manner. Lipid ROS content increased after the treatment of Rot, whereas pretreatment with TGY effectively reduced lipid ROS content, decreased MDA level and increased SOD activity and GSH level in damaged cells in cells damaged by Rot. Transmission electron microscopy showed that the mitochondria of PC12 cells were shrunk after the damage of 0.6 μmol/L Rot and the mitochondrial morphology of PC12 cells was improved to some extent after preprotection of TGY compared with normal group. Western blot results showed that TGY pretreatment could increase the expression of GPX4 and reduce the expression of ACSL4 and LPCAT3 after damage of Rot to a certain extent. CONCLUSION: TGY can improve nerve damage by up-regulating the expression of GPX4 protein, down-regulating the expression of ACSL4 and LPCAT3 protein to inhibit the oxidation of unsaturated fatty acids, reduce the level of lipid peroxidation and ROS content.

OBJECTIVE: To investigate the effects of Naoluo Xintong Decoction (NLXTD) on pyroptosis and angiogenesis of brain microvascular endothelial cells (BMECs) and explore the possible mechanisms in rats with oxygen-glucose deprivation/ reperfusion (OGD/R). METHODS: Rat BMECs with or without caspase-1 siRNA transfection were cultured in the presence of 10% medicated serum from NLXTD-treated rats (or blank serum) and exposed to OGD/R. CCK-8 assay, Transwell chamber assay, and tube formation assay were used to assess proliferation, migration, and tube-forming abilities of the cells. The activity of lactate dehydrogenase (LDH) in the culture supernatant was determined using a commercial assay kit, and the levels of inflammatory factors IL-1β and IL-18 were detected with ELISA. The cellular expressions of pro-caspase-1, caspase-1, NLRP3, Gasdermin D, and angiogenesis-related proteins VEGF and VEGFR2 were detected using Western blotting. RESULTS: The BMECs showed obvious injuries after OGD/R exposure. Compared with the blank serum, the medicated serum significantly improved the cell viability, migration ability, and lumen-forming ability (P < 0.01) and lowered the levels of IL-1β and IL-18 and the LDH release (P < 0.01) of the cells with OGD/R exposure. Western blotting showed that in the BMECs exposed to OGD/R, the medicated serum strongly upregulated the expression of VEGF and VEGFR2 proteins (P < 0.01) and reduced the protein expressions of pro-caspase-1, caspase-1, NLRP3, and Gasdermin D (P < 0.01), and transfection of the cells with caspase-1 siRNA further promoted the expressions of VEGFR2 protein in the cells (P < 0.01). CONCLUSION NLXTD can improve the proliferation, migration, and tube- forming ability and promote angiogenesis of BMECs with OGD/R injury probably by inhibiting the caspase-1/Gasdermin D pathway in pyroptosis, alleviating cell injury, and upregulating the expressions of VEGF and VEGFR2.
Animals ; Rats ; Endothelial Cells ; Caspase 1 ; Gasdermins ; Interleukin-18 ; NLR Family, Pyrin Domain-Containing 3 Protein ; Vascular Endothelial Growth Factor A ; Reperfusion Injury ; Brain ; Angiogenic Proteins ; Glucose

Heart failure is a complex clinical syndrome，which is the final result of compensatory failure of heart injury caused by various reasons. Long-term persistent cardiac stress leads to mitochondrial dysfunction，which in turn further damages cardiomyocytes and leads to disease progression. Timely removal of damaged mitochondria in cardiomyocytes and maintaining a good living environment of viable mitochondria is not only an effective means to protect cardiomyocytes，but also a new way to prevent and treat heart failure and ventricular remodeling. Mitochondrial quality control is a series of cellular activities for mitochondria to maintain their structural and functional stability，including oxidative stress response，regulation of mitochondrial dynamics，mitochondrial autophagy，intracellular calcium regulation and so on. Traditional Chinese medicine（TCM） mostly uses drugs of replenishing Qi and activating blood circulation in the treatment of chronic heart failure，and Qi and mitochondria are similar in function. According to TCM，the performance of the body as "static，descending and inhibitory" in the case of Qi deficiency can also be compared with the energy defect of mitochondria. The classical method of tonifying qi and activating blood circulation in TCM can be applied here. In recent years，TCM takes mitochondria as the target and carries out many related experimental studies from the point of view of myocardial energy supply. It is found that Chinese herbs for replenishing Qi and activating blood circulation can participate in regulating the quality control mechanism of intracellular mitochondria with multiple targets and links. It is proved by experiments that Chinese herbs for replenishing Qi and activating blood circulation can exert myocardial protective effect through this mechanism.