Research progress of the mechanism of traditional Chinese medicine in the prevention and treatment of athero-sclerosis based on miRNA regulation of vascular endothelial cell injury / 中国药房

The injury of vascular endothelial cells is not only the initial condition to promote the occurrence of early atherosclerosis （AS） plaques， but also an important link in the pathogenesis of AS. The microRNA （miRNA）， as an important medium of intercellular communication and gene regulatory factor， can affect vascular endothelial function and participate in the development of AS. The molecular mechanism of miRNA’s multi-target intervention in vascular endothelial cell injury has become a hot topic in the research of cardiovascular diseases. Monomers of traditional Chinese medicines such as ginsenoside Rb2 and paeonol， as well as traditional Chinese medicine for resolving phlegm and removing blood stasis could regulate miRNA to improve endothelial cell inflammation； astragaloside Ⅳ， dihydromyricetin and notoginsenoside could target miRNA and inhibit vascular endothelial oxidative stress； Danhong injection， Jianpi qutan and huayu prescription and paeonol could affect endothelial autophagy through miRNA； resveratrol， Bushen huoxue formula and Bushen tongmai formula could inhibit vascular endothelial aging by miRNA； dendrobine played an active role in regulating miRNA and improving endoplasmic reticulum stress. In the future， more in- depth research is needed on the effectiveness， mechanism of action， diagnosis and treatment plans， and safety of targeted regulation of miRNA for AS therapy by traditional Chinese medicine.

Expression and role of deleted in malignant brain tumor protein 1 in acute respiratory distress syndrome rats induced by sepsis / 中华危重病急救医学

Objective:To observe the expression of deleted in malignant brain tumor protein 1 (DMBT1) in rat acute respiratory distress syndrome (ARDS) model induced by sepsis and its relationship with ARDS related biomarkers.Methods:Forty-eight healthy male rats were randomly divided into sham operation group (Sham group) and ARDS model group, and the rats in each group were further divided into three subgroups at 6, 12 and 24 hours after operation, with 8 rats in each subgroup. The rats in the Sham group were exposed to the cecum only, and sepsis induced ARDS model was reproduced by cecal ligation and puncture (CLP) in the ARDS model group. The general performance was observed at 6, 12, 24 hours after operation. Abdominal aortic blood of rats was collected, and the levels of DMBT1, surfactant-associated protein D (SP-D), vascular endothelial growth factor (VEGF), interleukins (IL-6, IL-10) in serum were determined by enzyme-linked immunosorbent assay (ELISA). The lung tissues were collected, and the lung wet/dry weight (W/D) ratio was determined. The lung tissue pathological changes were observed under light microscope after hematoxylin-eosin (HE) staining, and the lung tissue injury score was evaluated. The expression of DMBT1 protein in lung tissue was determined by Western blotting. The relationship between the serum DMBT1 and SP-D, VEGF, IL-6, IL-10, lung tissue injury score were analyzed by Pearson correlation analysis.Results:Rats in the ARDS model group showed obvious pathological manifestations after operation. The alveolar structure destruction, inflammatory cell infiltration, and alveolar hemorrhage were observed under microscope. Compared with the Sham group, the lung tissue injury score and the lung W/D ratio at 12 hours after operation in the ARDS model group were significantly increased (lung tissue injury score: 3.35±0.13 vs. 1.16±0.07, lung W/D ratio: 5.36±0.44 vs. 4.38±0.35, both P < 0.05), and pulmonary edema was present, which suggested that the ARDS model caused by CLP was successfully reproduced. The results of ELISA and Western blotting showed that the levels of serum DMBT1, SP-D, VEGF and IL-6 in the ARDS model group increased gradually with time, while the level of IL-10 increased first and then decreased. Compared with the Sham group, the levels of DMBT1 in serum and the expressions of DMBT1 protein in lung tissue in the ARDS model group were significantly increased from 6 hours after operation [serum (ng/L) : 231.96±19.17 vs. 187.44±10.19, lung tissue (DMBT1/β-actin): 2.05±0.19 vs. 0.93±0.25, both P < 0.05], and the levels of SP-D, VEGF, IL-6 and IL-10 in serum were significantly increased from 12 hours after operation [SP-D (ng/L): 73.35±8.05 vs. 43.28±5.77, VEGF (ng/L): 89.85±8.47 vs. 43.19±5.11, IL-6 (ng/L): 36.01±2.48 vs. 17.49±1.77, IL-10 (ng/L): 84.55±8.41 vs. 39.83±5.02, all P < 0.05]. Pearson correlation analysis showed that serum DMBT1 was positively correlated with serum SP-D, VEGF, IL-6, IL-10 and lung injury score at 12 hours and 24 hours in the ARDS model group (12 hours: r values were 0.946, 0.942, 0.931, 0.936, 0.748, respectively; 24 hours: r values were 0.892, 0.945, 0.951, 0.918, 0.973, respectively; all P < 0.05). Conclusion:DMBT1 is a novel early biomarker of ARDS by affecting alveolar epithelial cell, alveolar capillary permeability and inflammatory response.

Effect and mechanism of Danggui Buxue Decoction-containing serum in mitigating H9c2 cell injury caused by exposure to intermittent low oxygen / 中国中药杂志

This study aims to explore the effect and mechanism of Danggui Buxue Decoction(DBD)-containing serum in alleviating the H9c2 cell injury caused by the exposure to intermittent low oxygen. H9c2 cells were assigned into five groups: control(CON) group, intermittent low oxygen(IH) group, intermittent low oxygen plus DBD-containing serum(IH+DBD) group, intermittent low oxygen plus the autophagy enhancer rapamycin(IH+RAPA) group, and intermittent low oxygen plus DBD-containing serum and the autophagy inhibitor 3-methyladenine(IH+DBD+3-MA) group. Monodansylcadaverine(MDC) staining was employed to detect the changes of autophagosomes. Cell counting kit-8(CCK-8) assay was employed to determine the activity of myocardial cells, and lactate dehydrogenase(LDH) and creatine kinase(CK) kits were used to measure the LDH and CK levels in the cell culture, which would reflect the degree of cell damage. TdT-mediated dUTP nick-end labeling(TUNEL) staining was used to detect the apoptosis of myocardial cells, and JC-1 fluorescence probe to detect the changes in mitochondrial membrane potential. Western blot was employed to determine the expression levels of the autophagy-related proteins microtubule-associated proteins light chain 3Ⅱ(LC3Ⅱ), microtubule-associated proteins light chain 3Ⅰ(LC3Ⅰ), P62, Parkin and apoptosis related proteins pro caspase-3, caspase-3, B-cell lymphoma-2(Bcl-2), Bcl-2-associated X(Bax). The results showed that compared with the CON group, the IH group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated expression of P62. In addition, the IH group showed decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, and decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. Compared with the IH group, the IH+DBD and IH+RAPA groups showed increased fluorescence intensity of MDC staining, increased LC3Ⅱ/LC3Ⅰ ratio, up-regulated Parkin expression, and down-regulated P62 expression. In addition, the two groups showed increased cell survival rate, reduced content of LDH and CK in the culture medium, decreased number of TUNEL positive cells, and increased pro caspase-3/caspase-3 and Bcl-2/Bax ratios and mitochondrial membrane potential. The IH+DBD+3-MA and IH groups showed no significant differences in the above indicators. Compared with the IH+DBD group, the IH+DBD+3-MA group showed decreased fluorescence intensity of MDC staining, decreased LC3Ⅱ/LC3Ⅰ ratio, down-regulated Parkin expression, and up-regulated P62 expression. In addition, the group had decreased cell survival rate, increased content of LDH and CK in the culture medium, increased number of TUNEL positive cells, decreased pro caspase-3/caspase-3 and Bcl-2/Bax ratios, and declined mitochon-drial membrane potential. To sum up, DBD could promote the mitophagy, inhibit the apoptosis, and alleviated the injury of H9c2 cells exposed to low oxygen.

Role of lysosomal cathepsin B in endothelial cell injury induced by hypoxia / 中国药理学通报

Aim To investigate whether CTSB is involved in hypoxia-induced injury of cardiac microvascular endothelial cells.Methods A hypoxia-induced endothelial cell injury model was used.Cardiac microvascular endothelial cells were isolated from CTSB gene knockout mice.CTSB was overexpressed by adenovirus delivery system, and bafilomycin was used to block autophagy.ELISA was used to detect the release of inflammatory factors.Tunel staining was used to detect the number of cell apoptosis.caspase-3 kit was used to detect the activity of cell caspase-3.Cells were infected with LC3-GFP-mCherry double-labeled adenovirus todetect cell autophagy flow.Results CTSB gene knockout could significantly aggravate the inflammation and apoptosis of endothelial cells induced by hypoxia, and increased autophagy.Overexpression of CTSB reduced the inflammation and apoptosis of endothelial cells induced by hypoxia, and increased autophagy.But bafilomycin treatment could significantly offset the inhibitory effect of CTSB overexpression on cell inflammation and apoptosis and the protective effect on cell autophagy.Conclusions CTSB knockout aggravates inflammation and apoptosis induced by hypoxia in endothelial cells; while the overexpression of CTSB ameliorates endothelial cell injury induced by hypoxia.CTSB maintains normal autophagy degradation in endothelial cells.BAF blocks the protective effect of CTSB on endothelial cells by inhibiting autophagy degradation.

Effect of Ranae Oviductus Protein Hydrolysate on Ethanol-induced L-02 Cell Injury / 中国实验方剂学杂志

Objective:To study the protective effect and mechanism of Ranae Oviductus protein hydrolysate （ROPH） on the expression of pathway-related proteins in ethanol-induced L-02 cell injury. Method:The ROPH was prepared by compound enzymatic hydrolysis. L-02 cell injury model was induced with 400 mmol·L^-1ethanol. Cell viability was detected by cell counting kit-8 （CCK-8） assay. Cell cycle and apoptosis were examined by flow cytometry. JC-1/Hochest staining was employed for qualitative investigation. The expression of related proteins in apoptosis， mitogen-activated protein kinase （MAPK） signaling pathway， and pyroptosis in L-02 cells was detected by Western blot. Result:The results of the CCK-8 assay showed that 400 mmol·L^-1ethanol could induce L-02 cell injury within 12 hours. Compared with the blank group， the model group showed decreased viability of L-02 cells （<italic>P</italic><0.01）， elevated percentage of the cell cycle in the G₀/G₁ phase （<italic>P</italic><0.01）， increased total cell apoptosis rate （<italic>P</italic><0.01）， reduced mitochondrial membrane potential （<italic>P</italic><0.01）， up-regulated expression of apoptosis-related proteins ［B-cell lymphoma-2 （Bcl-2）-associated X protein （Bax）， Cytochrome C （Cyt C）， and cysteine-dependent aspartate specific protease-3 （Caspase-3）］ （<italic>P</italic><0.05， <italic>P</italic><0.01） and MAPK signaling pathway-related proteins ［C-Jun amino-terminal kinase （JNK） and p38 MAPK］ （<italic>P</italic><0.05， <italic>P</italic><0.01）， and potentiated expression of pyrolysis-related proteins Caspase-1 and interleukin-1<italic>β </italic>（IL-1<italic>β</italic>） （<italic>P</italic><0.05）. Compared with the model group， the ROPH treatment group exhibited improved cell cycle arrest （<italic>P</italic><0.05， <italic>P</italic><0.01）， diminished total cell apoptosis rate （<italic>P</italic><0.01）， elevated mitochondrial membrane potential in a dose-dependent manner， down-regulated expression of Bax， Cyt C， and Caspase-3 proteins （<italic>P</italic><0.05， <italic>P</italic><0.01）， up-regulated expression of Bcl-2 protein （<italic>P</italic><0.05， <italic>P</italic><0.01）， and a downward trend in expression of proteins related to MAPK signaling pathway and pyrolysis （<italic>P</italic><0.05， <italic>P</italic><0.01）. Conclusion:ROPH could inhibit oxidative stress-triggered liver injury in ethanol-induced cells by improving mitochondrial membrane potential， reducing the expression of proteins in the mitochondria-mediated apoptosis pathway， and inhibiting the expression of proteins related to the MAPK signaling pathway and pyrolysis pathway to reduce the mitochondrial dysfunction and inflammatory response in ethanol-induced L-02 liver cells and inhibit oxidative stress， thereby exerting a therapeutic role in alcoholic liver injury.

Cyclovirobuxine D ameliorates aldosterone-induced primary neonatal rat cardiac myocyte injury via Ca / 中国药理学通报

Aim To investigate the protective effect of cyclovirobuxine D(CVB-D) on aldosterone (ALD)-induced primary neonatal rat cardiac myocytes (PNRC-Ms) injury and the possible mechanism. Methods PNRCMs were extracted by trypsin, and the PNRCMs injury model was established by ALD (10 μmol · L

Protective Effect of Salvianolic Acid B on Arsenic Trioxide-induced HepaRG Human Hepatocyte Injury / 中国实验方剂学杂志

Objective::To investigate the protective effect of salvianolic acid B on HepaRG hepatocyte injury induced by arsenic trioxide (As2O3 ) and its mechanism. Method::HepaRG cells were incubated with 5μmol·L-1 As2O3 for 24 h to induce hepatocyte injury. The cells were divided into control group, model group, salvianolic acid B 10 μmol·L-1 group, salvianolic acid B 10 μmol·L-1+ As2O3 group, salvianolic acid B 5 μmol·L-1+ As2O3 group, and salvianolic acid B 2.5 μmol·L-1+ As2O3 group. HepaRG cells were preincubated with salvianolic acid B for 2 h and then incubated with As2O3 for 24 h. At the end of the incubation, cell viability was detected by thiazolyl blue tetrazolium bromide assay, apoptosis was observed by Hoechst33342 fluorescence staining, apoptosis rate was detected by annexin V-FITC/propidium iodide double staining flow cytometry, and mitochondrial membrane was observed by JC-1 fluorescence staining. Western blot was used to detect the protective effect of expressions of relevant proteins Bcl-2, Bax, Akt, p-Akt on salvianolic acid B in the liver. Result::As2O3 concentration-dependently reduced the survival rate of HepaRG cells(P<0.01), salvianolic acid B had no effect on normal cell viability for 2 h, pre-incubation with salvianolic acid B(5, 10 μmol·L-1) for 2 h significantly increased the decreased cell survival rate caused by As2O3 (P<0.01). As2O3 significantly increased hepatocytes apoptosis rate(P<0.01), while pre-incubation with salvianolic acid B(10 μmol·L-1) deceased apoptosis rate(P<0.01). Incubation with As2O3 for 24 h caused decrease of mitochondrial membrane potential, pre-incubation with salvianolic acid B maintained mitochondrial membrane potential, indicating that the anti-apoptotic effect of salvianolic acid B were related to the mitochondrial pathway modulation. Western blot analysis showed that salvianolic acid B promoted the ratio of Bcl-2/Bax and promoted p-Akt/Akt compared with As2O3 group(P<0.01). Conclusion::Salvianolic acid B has a protective effect on hepatocyte injury induced by As2O3, and its mechanism is related to maintenance of mitochondrial function and inhibition of hepatocyte apoptosis.

Sal b attenuates cardiomyocyte injury by regulating priming of nlrp3 inflammasomes / 中国药理学通报

Aim To investigate whether Sal B alleviates hypoxic-induced cardiomyocyte injury by regulating the priming of NLRP3 inflammasomes. Methods The effects of Sal B on growth of H9C2 cells were examined by CCK8 assay,then the appropriate concen tration of Sal B was selected. The expression level of LDH was detected by Microliter assay. The expression levels of cTn/IL-lp were measured by Elisa assay. The protein and mRNA levels of TLR4/Myd88/I-RAK1/NF-kB/NLRP3 were detected by Western blot and qPCR. Results Hypoxia intervention notably reduced the viability of H9C2 cells and increased the expression levels of cTn/IL-IP. Besides,the protein and mRNA expression levels of TLR4/Myd88/IRAK1/NF-kB/NLRP3 were significant uP-regulated after hypoxia intervention. However, the viability of H9C2 cells increased, the secretion levels of LDH/cTn/IL-1 p were reduced,and the protein and mRNA levels of TLR4/Myd88/IRAK1/NF-KB/NLRP3 were significant inhibited after pretreated with Sal B. Conclusion Sal B attenuates cardiomyocyte injury by regulating the priming of NLRP3 inflammasome.I.

Research progress on the mechanism of endothelial cells injury in diabetic nephropathy / 解放军医学杂志

Diabetic nephropathy (DN) is one of the diabetic microvascular complications characterized by progressive protein uria and renal failure, which may eventually develop into end-stage renal disease (ESRD). Glomerular endothelial cells are one of the important components of glomerular filtration barrier. The structural and functional integrity of these cells are closely related to the maintenance of glomerular filtration function. Dysfunction of glomerular endothelial cells can lead to proteinuria, glomerulosclerosis and interstitial fibrosis, further damaging renal function and accelerating the progression of DN. The mechanisms leading to endothelial dysfunction include glucose metabolism disorder, oxidative stress, abnormal angiogenesis, inflammation and endothelial transdifferentiation. In recent years, it has been proposed that mechanisms such as intercellular communication, epigenetics and exosomes may be involved in the injury of diabetic nephropathy. In present paper, the research progress of related mechanisms was reviewed on the damage of glomerular endothelial cells in DN.

Paired Immunoglobulin-like Receptor B Inhibition in Müller Cells Promotes Neurite Regeneration After Retinal Ganglion Cell Injury in vitro / 神经科学通报·英文版

In the central nervous system (CNS), three types of myelin-associated inhibitors (MAIs) have major inhibitory effects on nerve regeneration. They include Nogo-A, myelin-associated glycoprotein, and oligodendrocyte-myelin glycoprotein. MAIs possess two co-receptors, Nogo receptor (NgR) and paired immunoglobulin-like receptor B (PirB). Previous studies have confirmed that the inhibition of NgR only results in a modest increase in regeneration in the CNS; however, the inhibitory effects of PirB with regard to nerve regeneration after binding to MAIs remain controversial. In this study, we demonstrated that PirB is expressed in primary cultures of retinal ganglion cells (RGCs), and the inhibitory effects of the three MAIs on the growth of RGC neurites are not significantly decreased after direct PirB knockdown using adenovirus PirB shRNA. Interestingly, we found that retinal Müller cells expressed PirB and that its knockdown enhanced the regeneration of co-cultured RGC neurites. PirB knockdown also activated the JAK/Stat3 signaling pathway in Müller cells and upregulated ciliary neurotrophic factor levels. These findings indicate that PirB plays a novel role in retinal Müller cells and that its action in these cells may indirectly affect the growth of RGC neurites. The results also reveal that PirB in Müller cells affects RGC neurite regeneration. Our findings provide a novel basis for the use of PirB as a target molecule to promote nerve regeneration.

Paired Immunoglobulin-like Receptor B Inhibition in Müller Cells Promotes Neurite Regeneration After Retinal Ganglion Cell Injury in vitro / 神经科学通报·英文版

In the central nervous system (CNS), three types of myelin-associated inhibitors (MAIs) have major inhibitory effects on nerve regeneration. They include Nogo-A, myelin-associated glycoprotein, and oligodendrocyte-myelin glycoprotein. MAIs possess two co-receptors, Nogo receptor (NgR) and paired immunoglobulin-like receptor B (PirB). Previous studies have confirmed that the inhibition of NgR only results in a modest increase in regeneration in the CNS; however, the inhibitory effects of PirB with regard to nerve regeneration after binding to MAIs remain controversial. In this study, we demonstrated that PirB is expressed in primary cultures of retinal ganglion cells (RGCs), and the inhibitory effects of the three MAIs on the growth of RGC neurites are not significantly decreased after direct PirB knockdown using adenovirus PirB shRNA. Interestingly, we found that retinal Müller cells expressed PirB and that its knockdown enhanced the regeneration of co-cultured RGC neurites. PirB knockdown also activated the JAK/Stat3 signaling pathway in Müller cells and upregulated ciliary neurotrophic factor levels. These findings indicate that PirB plays a novel role in retinal Müller cells and that its action in these cells may indirectly affect the growth of RGC neurites. The results also reveal that PirB in Müller cells affects RGC neurite regeneration. Our findings provide a novel basis for the use of PirB as a target molecule to promote nerve regeneration.

Clinical and biochemical features of bile duct injury type of drug-induced liver injury / 临床肝胆病杂志

ObjectiveTo investigate the pathological and biochemical features of each pathological subtype of bile duct injury type of drug-induced liver injury (DILI), and to verify the value and significance of pathological classification of DILI. MethodsA retrospective analysis was performed for the clinical data of 112 patients with bile duct injury type of DILI who were admitted to Shijiazhuang Fifth Hospital from January 2006 to January 2016 and China-Japan Friendship Hospital from October 2003 to June 2014. According to the pathological subtype, the patients were divided into mixed hepatitis group with 40 patients, cholestatic hepatitis group with 40 patients, and simple cholestasis group with 32 patients, and the three groups were compared in terms of types of drugs used, course of disease, R value, and peak values, changing trend, time to peak, and recovery time of liver biochemical indices. The independent-samples Kruskal-Wallis H test was used for comparison of continuous data between multiple groups, and the Mann-Whitney U test was used for further comparison between two groups; the chi-square test was used for comparison of categorical data between groups. ResultsAmong the drugs inducing DILI, traditional Chinese medicine and Western medicine each accounted for half of the cases of bile duct injury type of DILI. Traditional Chinese medicine mainly included the drugs for osteoarthropathy, intervertebral disc bulge, alopecia, calculus-removing and cholagogic treatment, Yang-tonifying therapy, and skin diseases; 26 patients (65%) in the cholestatic hepatitis group had DILI caused by traditional Chinese medicine, while 16 patients (40%) in the mixed hepatitis group and 13 (40.6%) in the simple cholestasis group had such DILI. Antibiotics and antipyretic and analgesic drugs were the most common Western medicines for DILI. The mixed hepatitis group had the highest peak values of ALT and AST and R value, followed by the cholestatic hepatitis group and the simple hepatitis group (χ2=54.77, 44.21, and 5195, all P＜0.001), and there were no significant differences in the peak values of the other liver biochemical parameters between the three groups (all P＞0.05). In the mixed hepatitis group and the cholestatic hepatitis group, the time to peak of TBil was longer than that of ALT. There were no significant differences in course of disease, time to peak of liver biochemical parameters, and recovery time between the three groups (all P＞0.05). ConclusionEach subtype of bile duct injury type of DILI has unique clinical and biochemical features, and an understanding of such features may help to accurately judge clinical typing, pathological changes of targets, and degree of injury.

The role of apolipoprotein C4 in vascular endothelial cell injury in hypertensive disorder of pregnancy / 中华检验医学杂志

Objective Find abnormal changes of plasma lipid metabolism-related proteins before 20 weeks of gestation in patients with hypertensive disorder of pregnancy(HDP), and preliminarily investigate the role of plasma apolipoprotein C4 elevation in HDP. Methods A nested case-control study was used. The plasma were collected from pregnant women who underwent routine prenatal examination in Guangzhou Women and Children's Medical Center from November 2014 to March 2017. Label-free mass spectrometry was used to detect the differences in plasma lipid metabolism-related proteins before 20 weeks of gestation between 12 pairs of HDP patients and normal controls, and different 48 pairs of samples were used for verification. The protein with the most significant difference multiples was screened to study its effects on monolayer permeability and nitric oxide secretion of endothelial cells. One-way ANOVA was used for comparison between groups, and P<0.05 was considered as statistically significant difference. Results Compared with the control, the lipid metabolism-related proteins, APOC4, Fatty acid-binding protein 4 (FABP4), Apolipoprotein E (APOE), Apolipoprotein C3 (APOC3) and Beta-2-glycoprotein 1(APOH) raised to 1.94, 1.82, 1.59, 1.55 and 1.38 times, phospholipid transfer protein (PLTP) decreased to 0.78 times in plasma before 20 weeks of pregnancy of patients with HDP (t value were 2.499, 2.497, 2.081, 2.098, 2.426 and 2.564, respectively, P<0.05). Cell experiments results showed that 50 ng / ml APOC4 significantly increased 20％ HUVEC single layer cell permeability to FITC-labeled dextran (F=455.4, P<0.01), and significantly decreased the level of nitric oxide in the supernatant of HUVEC culture by 25％ (F=61.92, P<0.01). Conclusions Before diagnosis, plasma protein levels involved in lipid metabolism in HDP patients have been changed, resulting in abnormal lipid metabolism. APOC4 can increase the permeability of vascular endothelial cells, inhibit endothelial source of NO secretion, cause endothelial dysfunction.

Protective effect and mechanism of total flavonoids of Fructus Livistonae on acetaminophen-induced liver injur / 中草药

Objective: To research the effects of total flavonoids of Livistona chinensis (TFFL) on normal liver LO2 cells induced by acetaminophen (APAP), and study the liver protective mechanism on liver injury induced by acetaminophen in vitro. Methods: The effects of TFFL on LO2 cells and cell activity induced by acetaminophen were determined by MTT method; The apoptosis rate of LO2 cells induced by APAP was researched by Flow Cytometry combine with staining agent Hoechst 33342; The levels of malondialdehyde (MDA), aspartate aminotransferase (AST), reduced glutathione (GSH), and superoxide dismutase (SOD) were examined; The expressions of iNOS and nitrotyrosine tubulin (NT) in APAP-induced LO2 hepatocyte injury cells was studied by Western blotting method. Results: TFFL had no toxicity to LO2 cells at a given concentration which can promote the proliferation. At the same time, experiment results showed that TFFL prophylactically reduced the apoptosis of LO2 cells induced by APAP. TFFL decreased the level of AST and MDA, increase the level of GSH and SOD, and inhibit the expressions of iNOS protein and NT protein. Conclusion: TFFL has a protective effect on LO2 cells injury induced by APAP. The possible protective mechanism of TFFL is related to the inhibition of oxidative stress and the nitro stress.

Crocetin promotes autophagy in injured rat hepatocytes induced by lipopolysaccharide and D-galactosamine / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the effect of crocetin on autophagy in rat hepatocytes exposed to lipopolysaccharide (LPS) and D-galactosamine (D-gal) and explore the mechanism.</p><p><b>METHODS</b>Cultured rat hepatocytes were exposed to LPS (1 mg/L) and Dgal (60 mg/L) to induce cell injury and treated with crocetin, 3MA, or crocetin+3MA. Twelve hours after the treatments, the cells were examined for levels of ALT, AST and LDH in the supernatant using ELISA. LC3 fluorescence in the cells following immunofluorescence staining was observed using fluorescence microscopy. Autophagosomes in the cells were observed by transmission electron microscopy, and the cellular expressions of LC3, p62 and SIRT1 were detected using Western blotting.</p><p><b>RESULTS</b>The levels of ALT, AST and LDH in the hepatocytes were elevated after LPS- and D-gal-induced injury, reached the highest levels after 3MA treatment, but were decreased significantly by crocetin treatment. LC3 fluorescence increased obviously in the injured hepatoctyes, and the increment was the most obvious in crocetin-treated cells; LC3 fluorescence was decreased significantly after 3MA treatment. Cell injury induced obvious increase in autophagy in the hepatocytes, and the number of autophagosomes increased significantly after crocetin treatment but was reduced significantly after 3MA treatment. The cell injury caused an obvious up-regulation of LC3 and SIRT1 expression and down-regulated p62 expression. LC3 and SIRT1 expression levels were the highest and the expression of p62 was the lowest in cells with crocetin treatment. 3MA treatment significantly reduced the expression of LC3 and SIRT1 and increased the expression of p62 in the injured cells.</p><p><b>CONCLUSIONS</b>Autophagy is increased in injured rat hepatocytes, and crocetin can promote autophagy in the injured cells to reduce further cell injury.</p>

Research progress on pharmacological effects of Polygonati Rhizoma / 中草药

As a traditional Chinese herbal medicine, Polygonati Rhizoma widely distributed in most areas south of the Yangtze River. It has the function of nourishing liver and kidney, prolonging life and so on. Importantly, it is a Taoist Holy medicine since ancient times. Polygonati Rhizoma has high medicinal value and nutritional value because it contains polysaccharides, saponins, flavonoids, lignin, amino acids, quinones, vitamins, alkaloids and a variety of trace elements and so on. The domestic research institutions have carried out a deeper exploration, while its research is still at an early stage for foreign countries. At present, the experimental studies are mainly concentrated on the polysaccharides, ethanol, the extracts of saponins or the aqueous extracts of Polygonati Rhizoma. The experimental type is mainly based on the animal experiments and the clinical researches of Polygonati Rhizoma or its compound preparations. Various Polygonati Rhizoma preparations have been widely used in clinic, such as Polygonati Rhizoma Oral Liquid, Polygonati Rhizoma Tea, Cistanche and Polygonati Rhizoma Granules, Polygonati Rhizoma Zanyu Capsules, Polygonati Rhizoma essence oil patch and so on, which play different roles in individual products. In this paper, a comprehensive analysis was carried out on the basis of the latest experimental research on Polygonati Rhizoma, and its utility value was summed up from various angles, which provides a reference for the deep development and application of the Polygonati Rhizoma.

Protectiveeffects of endovascular cooling treatment on post-resuscitation syndrome and its mechanism / 中华危重病急救医学

Objective To investigate the protective function of endovascular cooling method on post-resuscitation syndrome (PRS) in porcine cardiac arrest (CA) model and its mechanism.Methods Ventricular fibrillation (VF) was electrically induced and untreated for 8 minutes in 15 healthy male porcines, cardiopulmonary resuscitation (CPR) was then initiated. All successful recovery animals were randomly divided into two groups by random number table. In normal temperature group, the core temperature was maintained at (38.0±0.5) ℃ for 12 hours. In mild hypothermia group, the mild hypothermia treatment was initiated at 5 minutes after successful resuscitation, the treatment of rapid endovascular cooling was performed to reach the target cooling temperature of (33.0±1.0) ℃, and then maintained until 6 hours after resuscitation. Rewarming was implemented at the rate of 0.7 ℃/h until the body temperature reached (38.0±0.5) ℃. Hemodynamic parameters including heart rate (HR), mean arterial blood pressure (MAP), cardiac output (CO) were continually monitored. Right femoral vein blood was collected before VF and 1, 2, 4, 6, 12 and 24 hours after resuscitation, respectively, and the serum concentrations of E-selectin, soluble thrombomodulin (sTM), and interleukin-1β(IL-1β) were determined with enzyme linked immunosorbent assay (ELISA). The survival of porcines at 24 hours after resuscitation was observed, and the neurological deficit score (NDS) was calculated for the surviving porcines. All animals were sacrificed, and brain, heart and lung tissues were collected, after hematoxylin and eosin (HE) staining, the histopathology changes were evaluated under a light microscopy.Results After 8-minute VF, 14 porcines were resuscitated successfully, 7 porcines in normal temperature group and 7 in mild hypothermia group respectively, with the resuscitation success rate of 93.3％. There was no significant difference in body weigh, core temperature, hemodynamics, or blood lactate as well as duration of CPR and the number of defibrillations between the two groups. The core temperature of normal temperature group was maintained at (38.0±0.5) ℃, while in mild hypothermia group, the hypothermia was reduced to the hypothermia range (33.0±1.0) ℃until 6 hours, then rewarmed to normothermia gradually [(38.0±0.5) ℃]. Compared with those before VF, HR was significantly increased after resuscitation in both groups, and MAP and CO were decreased, then they tended to normal. There was no significant difference in hemodynamic parameter at all time points between the two groups. Compared with those before VF, the levels of E-selectin and sTM in serum of the two groups were increased significantly at 1 hour after resuscitation, and they were decreased gradually after reaching the peak at 6 hours, and IL-1β was increased continuously with time. There was no significant difference in E-selectin (μg/L:1.34±0.52 vs. 1.60±0.61), sTM (μg/L: 19.13±0.34 vs. 19.24±0.73), or IL-1β (ng/L: 25.73±0.87 vs. 25.32±0.25) before VF between normal temperature group and mild hypothermia group (allP> 0.05). The levels of E-selection, sTM and IL-1β in mild hypothermia group were significantly lower than those in normal temperature group from 2 hours after resuscitation [E-selection (μg/L): 11.15±2.73 vs. 16.04±3.23, sTM (μg/L): 49.67±3.32 vs. 62.22±1.85, IL-1β (ng/L): 140.51±6.66 vs. 176.29±18.51, allP< 0.05], and E-selection decreased to the baseline level at 12 hours (μg/L: 1.17±0.65 vs. 1.60±0.61,P > 0.05). The 24-hour survival rates of two groups were both 100％. The NDS score of mild hypothermia group was obviously lower than that of normal temperature group (150.0±6.6 vs. 326.4±12.3,P < 0.05). In normal temperature group, neuronal cell necrosis was observed in the cerebral cortex at 24 hours after resuscitation, and nucleus was deeply stained. The myocardial necrosis and alveolar collapse was found. Meanwhile the infiltration of inflammatory cell could be found in the myocardium and alveolar. The brain, lung and myocardium injury were significantly milder in mild hypothermia group as compared with those in normal temperature group.Conclusions The intravascular cooling therapy was a safe and effective method for inducing mild hypothermia after resuscitation. This cooling effect was fast and reliable, and the rewarming speed was controllable and stable. The protective mechanism of mild hypothermia on PRS may be related to inhibiting systemic inflammatory response and reducing vascular endothelial cell injury.

Protective effect and mechanism of Sestrin2 in the development of pulmonary alveolar type II epithelial cell injury induced by cigarette smoking / 中国比较医学杂志

Objective To explore the role of Sestrin2 in pulmonary alveolar type II epithelial cell injury induced by cigarette smoking and its mechanism of action. Methods The cell injury model was induced by cigarette smoke extract (CSE)in the human pulmonary alveolar type II epithelial A549 cells. The generation of ROS was detected by DCFDA fluorescence probe. The levels of inflammatory factors TNF-α and IL-8 were determined by ELISA, and the expression of Sestrin2 and the peroxiredoxin,Prx-SO2/3H,was detected by Western blot. In addition,all the events were also measured in the A549 cells which were transfected with Sestrin2 siRNA and treated with azithromycin. Results After the CSE treatment,the expression of Sestrin2 in the A549 cells was decreased, the expression of Prx-SO2/3H was increased, the ROS production,secretion of cytokines TNF-α and IL-8 were increased(P < 0.05). These changes were partly reducedby azithromycin, indicating that azithromycin significantly relieved CSE-induced oxidative stress and inflammatory injury.Silencing of Sestrin2 in the A549 cells result ed in an increase of Prx-SO2/3 H expression, ROS production and the secretionof the cytokines TNF-α and IL-8. However, oxidative stress and inflammatory injury were not alleviated with the addition ofazithromycin in the Sestrin2 siRNA silencing A549 cells. Conclusions Sestrin2 plays an protective role in the pulmonaryalveolar type II epithelial cell injury induced by cigarette smoking through negatively regulating the level of intracellularROS via catalyzing the reduction of the hyperoxidized peroxiredoxin Prx-SO2/3 H.

Kaempferol alleviates ox-LDL-mediated endothelial cell injury via regulating AMPK/Nrf2/HO-1 signaling pathway / 中国免疫学杂志

Objective:To investigate the effects of kaempferol on the oxidized low density lipoprotein(ox-LDL)-induced injury in human umbilical vein endothelial cell (HUVECs) and to explore its underlying molecular mechanism.Methods: HUVECs were randomly divided into 6 groups:control group,ox-LDL group,Kaempferol+ox-LDL group,Kaempferol+ox-LDL+Compound C group, Kaempferol+ox-LDL+si-Nrf2 group and Kaempferol+ox-LDL+si-HO-1 group.The cell activity was measured by MTT assay.The cell apoptosis was analyzed by flow cytometry.The protein expression were measured by Western blot.The ROS levels were evaluated by flow cytometry.Results: Compared with control group,ox-LDL treatment decreased the cell viability,increased the cell apoptosis,up-regulated the protein levels of cleaved-caspase 3 and down-regulated the Bcl-2 level,elevated the expression of TNF-α,IL-1β,IL-6, vascular cell adhesion molecule (VCAM1),intercellular adhesion molecule (ICAM1) and selectin E (E-selectin),promoted the reactive oxygen species (ROS) generation and reduced the superoxide dismutase(SOD) levels,and decreased the protein levels of p-AMPK,Nrf2 and HO-1.Compared with the ox-LDL group,kaempferol treatment reversed the ox-LDL-induced cell activity inhibition, apoptosis and oxidative stress damage,and increased the protein levels of p-AMPK,Nrf2 and HO-1.Compound C,si-Nrf2 and si-HO-1 inhibited AMPK/Nrf2/HO-1 signaling pathway,elevated the production of ROS and thus inhibiting the protective effects of kaempferol on ox-LDL-treated HUVECs.Conclusion:Kaempferol alleviates ox-LDL-induced endothelial cell injury,which may be related with the activation of AMPK/Nrf2/ HO-1 signaling pathway.

Role and mechanism of Angiotensin Ⅱ-induced ChemR23 in podocyte injury / 中华肾脏病杂志

Objective To observe the expression of ChemR23 induced by Angiotensin Ⅱ (Ang Ⅱ) in podocyte and its role in renal injury.Methods Conditionally immortalized mice podocytes were cultured in vitro.Immunofluorescence was used to observe the sub-cellular location of ChemR23.The expressions of ChemR23,Nephrin and Podocin stimulated by different concentrations of Ang Ⅱ were detected by qRT-PCR and Western blotting.Lentivirus targeting ChemR23 was used.The expressions of Nephrin and Podocin and the phosphorylation state of NF-κB P65 were detected by Western Blot.The inhibitor of NF-κB P65 was added to the cultural medium for 2 h before Ang Ⅱ stimulation.The effect of NF-κB P65 inhibitor on Ang Ⅱ-induced expression of Nephrin and Podocin was detected by Western Blot.Results It is showed that ChemR23 was located in cytosol and membrane.Compared with the normal control,the expression of ChemR23 was significantly increased by Ang Ⅱ in mRNA and protein level,while the expressions of Nephrin and Podocin were decreased (P ＜ 0.05).When using Lentivirus vector to interfere the expression of ChemR23,Ang Ⅱ-repressed expressions of Nephrin and Podocin were restored (P ＜ 0.05).Western Blot showed the level of phosphorylated NF-κB P65 was significantly increased by Ang Ⅱ stimulation (P ＜ 0.05),which could be inhibited by interfering the expression of ChemR23.When adding the NF-κB P65 inhibitor,the low expression of Nephrin and Podocin induced by Ang Ⅱ stimulation was restored (P＜0.05).Conclusions Ang Ⅱ can induce ChemR23 expression,which activates NF-κB P65 signaling pathway,and then inhibits the expressions of Nephrin and Podocin.Targeting ChemR23 is a potential way to alleviate podocyte injury caused by Ang Ⅱ.