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AIM:To screen and verify the effective ingredient of traditional Chines medicine (TCM) Q0409 in improving learning and memory ability of mice.METHODS:The mouse learning and memory impairment model was induced by intraperitoneal injection of scopolamine.The mice in each group were given the corresponding drug by gavage at the same time for 14 d in succession.Morris water maze test was used to measure the learning and memory ability of the mice, and then the hippocampal tissue homogenate was taken to determine the activity of acetylcholinesterase (AChE).The animals were divided into 8 groups according to L8(27) orthogonal table.The variance analysis and factorial analysis were used to analyze the pharmacological effects of seven kinds of single herbal in TCM Q0409 and determine the screening ingredients.The animals were divided into 6 groups according to the results of the preliminary screening results, and further testing and validation of TCM Q0409 screening ingredients were performed to get the final simplified ingredients.RESULTS:Three medicinal herbs of Polygalae, Panax ginseng and Acori graminei rhizome were screened by the orthogonal results of Morris water maze test and the activity of AChE in mouse hippocampal tissues.The simplified ingredients of TCM Q0409 were obtained through the variance results of Morris water maze test and the activity of AChE in mouse hippocampal tissues.CONCLUSION:Polygala and ginseng were eventually determined as simplified ingredients of TCM Q0409 and it was verified that they improve the learning and memory ability of the mice with learning and memory impairment.
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AIM: To observe the effects of berberine (Ber) on enterocyte apoptosis in septic mice and its pos-sible mechanism.METHODS: Male C57BL/6 mice (8 ~10 weeks old) were randomly divided into sham group, cecal ligation and puncture (CLP) group, CLP +Ber group and sham +Ber group.The mice in CLP group underwent CLP ope-ration, and the mice in sham groups suffered a similar operation except the ligation and puncture.After the sham or CLP operation, the mice were administered intragastrically with distilled water or berberine (50 mg/kg) within 2 h.After 20 h, the mice were killed with excess pentobarbital sodium and the ileum tissues were removed.The histological changes of the intestine were observed and the enterocyte apoptosis was examined by determining the protein level of cleaved caspase-3. Furthermore, mitochondrial Bax, cytoplasm cytochrome C (Cyt C) and the total proteins of Bcl-2, Fas, FasL and Fas-as-sociated protein with death domain (FADD) were examined by Western blot.The mRNA expression of tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) was measured by real-time PCR.RESULTS: The extensive ileum injuries, including remarkably increased leukocytes and necrosis of intestinal villus were observed 20 h after CLP.In CLP group, the protein levels of cleaved caspase-3, cytoplasm Cyt C, as well as Fas, FasL were significantly increased, but the Bcl-2 level was decreased.Bax translocation into mitochondria was promoted.However, FADD was not changed significantly.The mRNA expression of TH and DBH was also increased sharply in CLP group.On the contrary, treatment with berberine made a considerable alleviating alteration in the ileum of the septic mice.CONCLUSION: Treatment with berberine pro-vides protective effects on intestinal injury in septic mice by reducing enterocyte apoptosis, and its possible mechanism may be involved in the inhibition of the endogenous and exogenous apoptosis pathways.
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AIM:To study the expression of glycine receptorα1 subunit in neonatal rat myocardial cells and to investigate the effect of lipopolysaccharide (LPS), hypoxia/reoxygenation, isoproterenol (ISO) and high concentration of glucose (HG) on the expression of glycine receptorα1 subunit in the neonatal rat myocardial cells.METHODS:Neonatal rat myocardial cells were cultured in vitro.The expression of glycine receptorα1 subunit was detected by Western blotting. The neonatal rat myocardial cells were treated with LPS (20 mg/L), ISO (100 μmol/L) or high concentration of glucose (25 mmol/L) for 24 h, or were exposed to hypoxia for 3 h followed by reoxygenation for 3 h.Subsequently, the cell viabil-ity was measured by CCK-8 assay, and the expression of glycine receptorα1 subunit was determined by Western blotting. RESULTS:The expression of glycine receptor α1 subunit in the neonatal rat myocardial cells was positively detectable by Western blotting.Compared with control group, no significant difference of the cell viability ( P>0.05) in LPS group, ISO group, hypoxia/reoxygenation group and HG group was observed.The expression of glycine receptor α1 subunit was in-creased (P<0.01) in LPS group, ISO group and hypoxia/reoxygenatio group, but decreased (P<0.01) in HG group. CONCLUSION:Glycine receptorα1 subunit exists in the neonatal rat myocardial cells.A certain concentration of LPS or ISO, or hypoxia/reoxygenation for a certain period upregulate the expression of glycine receptorα1 subunit, but HG down-regulates the expression of glycine receptor α1 subunit in cultured neonatal rat myocardial cells.
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AIM:To observe the effect of B-HT933, a selective α2-adrenoceptor agonist, on lipopolysaccha-ride ( LPS )-induced TNF-αproduction in neonatal rat cardiomyocytes and to explore the underlying mechanisms . METHODS:The neonatal rat cardiomyocytes were cultured .The localization of α2A-adrenoceptor in the cardiomyocytes was examined by immunofluorescence staining .The cardiomyocytes were exposed to LPS or/and B-HT933 for different time.The level of TNF-αin the supernatants and the mRNA expression of TNF-αwere detected by ELISA and real-time PCR, respectively.In addition, LPS-associated signal molecules in the cardiomyocytes were also examined by Western blotting.RESULTS: Immunofluorescence staining showed that α2A-adrenoceptors were localized in the cardiomyocytes . LPS stimulated TNF-αproduction in the cardiomyocytes in a dose and time-dependent manner .B-HT933 pretreatment sig-nificantly inhibited the expression of TNF-αat mRNA and protein levels in LPS-treated cardiomyocytes .Furthermore, LPS exposure induced IκBαand p38 phosphorylation in cardiomyocytes and only IκBαphosphorylation was prevented by B-HT933 treatment.CONCLUSION:α2A-adrenoceptors are present in neonatal rat cardiomyocytes and its agonist B -HT933 inhibits LPS-induced TNF-αproduction in cardiomyocytes via suppressing IκBαphosphorylation .
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AIM:To explore the preliminary mechanism of senegenin ( Sen) on inhibiting hypoxia/reoxygenation ( H/R)-induced apoptosis of primary cortical neurons .METHODS:The cultured cortical neurons were randomly divided in-to normal group (control group), model group (H/R group), Sen+H/R group and Sen group.Flow cytometry was used to evaluate the effect of Sen on H/R-induced cell apoptosis .The protein levels of JNK , p-JNK, c-Jun, p-c-Jun, Bcl-2 and Bax were assessed by Western blotting .RESULTS:The apoptotic rate in H/R group was obviously higher than that in control group (P<0.05), while the apoptotic rate in Sen +H/R group was obviously lower than that in H/R group (P<0.05), suggesting that the model of apoptosis was established successfully .The results of Western blotting showed that Sen increased the expression of JNK and c-Jun, inhibited the phosphorylation of JNK and c-Jun (P<0.05), increased the protein level of Bcl-2 and inhibited the protein level of Bax in H/R treated primary cortical neurons (P<0.05).CONCLUSION:Sen has a protective effect against H/R-induced neuronal apoptosis by increasing the expression of JNK and c-Jun, inhibiting the phosphorylation of JNK and c-Jun, increasing the protein level of Bcl-2 and decreasing the protein level of Bax .
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RNA interference(RNAi)is a process by which introduced small interfering RNA(siRNA)can cause the specific degradation of mRNA with identical sequences. The human herpes simplex virus type 1(HSV-1)RR is composed of two distinct homodimeric subunits encoded by UL39 and UL40, respectively. In this study, we applied siRNAs targeting the UL39 and UL40 genes of HSV-1. We showed that synthetic siRNA silenced effectively and specifically UL39 and UL40 mRNA expression and inhibited HSV-1 replication. Our work offers new possibilities for RNAi as a genetic tool for inhibition of HSV-1 replication.
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Herpes simplex virus type 1 (HSV-1) is a commonly occurring human pathogen worldwide. There is an urgent need to discover and develop new alternative agents for the management of HSV-1 infection. Tripterygium hypoglaucum (level) Hutch (Celastraceae) is a traditional Chinese medicine plant with many pharmacological activities such as anti-inflammation, anti-tumor and antifertility. The usual medicinal part is the roots which contain about a 1% yield of alkaloids. A crude total alkaloids extract was prepared from the roots of T. hypoglaucum amd its antiviral activity against HSV-1 in Vero cells was evaluated by cytopathic effect (CPE) assay, plaque reduction assay and by RT-PCR analysis. The alkaloids extract presented low cytotoxicity (CC_(50) = 46.6 μg/mL) and potent CPE inhibition activity, the 50% inhibitory concentration (IC_(50)) was 6.5 μg/mL, noticeably lower than that of Acyclovir (15.4 μg /mL). Plaque formation was significantly reduced by the alkaloids extract at concentrations of 6.25 μg/mL to 12.5 μg/mL, the plaque reduction ratio reached 55% to 75% which was 35% higher than that of Acyclovir at the same concentration. RT-PCR analysis showed that, the transcription of two important delayed early genes UL30 and UL39, and a late gene US6 of HSV-1 genome all were suppressed by the alkaloids extract, the expression inhibiting efficacy compared to the control was 74.6% (UL30), 70.9% (UL39) and 62.6% (US6) respectively at the working concentration of 12.5μg/mL. The above results suggest a potent anti-HSV-1 activity of the alkaloids extract in vitro.
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The primary action of corticotropin releasing hormone (CRH) is stimulation of the synthesis and release of adrenocorticotropic hormone and β-endorphin from the pituitary in response to stress. In addition, a number of studies indicate that CRH exerts other physiological actions within the central nervous system which are independent of the pituitary. These include increased body temperature and thermogenesis. However, the intracellular mechanism responsible for pyrogenic action of CRH is still unclear. The purpose of these studies was to determine whether or not cAMP was involved in the pyrogenic action of CRH in the rat. Intracerebroventricular (icv) microinjection of CRH (2.5 μg, 5.0 μg, 10 μg) caused increases in colonic temperature and hypothalamus cAMP level in conscious rats. The pyrogenic effects of CRH were abolished or markedly inhibited by prior injection (icv) of an adenylate cyclase inhibitor, 2，，3，-dideoxyadenosine (DDA, 30 μg) or an inhibitor of cAMP-dependent protein kinase, adenosine-3，，5，-(cyclic) monophosphorothionate (Rp-cAMPs, 15 μg). This is the first report demonstrating the pyrogenic effcet of centrally administration of CRH on the rat via the cAMP-mediated protein kinase signal transduction pathway.
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AIM and METHODS:To investigate the effect of electrical stimulation of VSA on the firing of thermosensitive neurons in preoptic anterior hypothalamus (POAH), the firing rate of thermosensitive neurons in POAH of 20 New Zealand white rabbits was recorded by using extracellular microelectrode techinque. RESULTS:(1)Electrical stimulation of ventral septal area (VSA) caused a significant increase in firing rate of warm-sensitive neurons in the preoptic area of the anterior hypothalamus(POAH).(2) The firing rate of cold-sensitive neurons was decreased remarkably in the POAH by electrical stimulation of VSA. CONCLUSION:VSA may play a controlling role in the thermoregulation through altering the firing rate of thermosensitive neurons in the POAH.
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AIM: To investigate whether glycine receptor is involved in the protection of glycine against(anoxia/reoxygenation) injury in cardiomyocytes by detecting oxygen free radical metabolism,apoptosis and intracellular calcium overload.METHODS: The neonatal rat cardiomyocytes were cultured and exposed to anoxia and reoxygenation((A/R)) in the presence of glycine receptor antagonist,glycine or in free chloride buffer.The superoxide dismutase(SOD) activity,the contents of malondialdehyde(MDA) and nitric oxide(NO),the intracellular free calcium concentration and the apoptotic rate in the cardiomyocytes were determined.RESULTS: SOD activity and NO content in cardiomyocytes were lower,but MDA content,intracellular free calcium concentration and apoptotic rate in cardiomyocytes were higher in A/R group than those in control.Pretreatment with glycine inhibited the above changes caused by A/R,which was reversed by strychnine treatment and in the free chloride medium.CONCLUSIONS: Glycine inhibits free radical production,attenuates calcium overload,decreases apoptotic rate and increases SOD activity and NO release in cardiomyocytes exposed to(A/R).These findings suggest that glycine exerts a protective effect against A/R injury via glycine receptor and glycine protects the neonatal rat cardiomycytes from A/R-induced injury in a chloride-dependent manner.
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AIM: To investigate the mechanisms by which siduqing, a Chinese medicine, protects against lipopolysaccharide (LPS)-induced acute renal dysfunction. METHODS: Mice were divided randomly into control, LPS, siduqing treatment and siduqing groups, and treated intragastrically with siduqing at a dose of (1 000) g/L (0.2 (mL/10 g) body weight) or distilled water (0.2 (mL/10) g body weight) twice a day for 3 days, LPS (30 mg/kg) or normal saline was injected intraperitoneally on day 3, followed by intragastrical administration with siduqing at a dose of (1 000) g/L (0.2 (mL/10 g) body weight) or distilled water (0.2 (mL/10 g) body weight). Blood was collected for determining urea nitrogen (BUN) and creatinine (Cr) contents, renal tissue for examining superoxide dismutase (SOD) activity and malondialdehyde (MDA) content. In addition, electron microscopy was used to examine the ultrastructure changes in kidney, and RT-PCR was performed to detect renal intercellular adhesion molecule-1 (ICAM-1) mRNA expression. RESULTS: LPS significantly increased serum urea nitrogen (BUN) and creatinine (Cr) contents, and produced an obvious pathological change in renal ultrastructure, which were significantly attenuated by siduqing treatment. Moreover, siduqing treatment increased renal SOD activity, also markedly suppressed an increase in renal MDA production and ICAM-1 mRNA expression induced by LPS. CONCLUSION: These results suggest that siduqing protects against LPS-induced acute renal injury through inhibiting ICAM-1 mRNA expression, enhancing renal SOD activity and attenuating oxidant stress.
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AIM: To study the influence of glycine(GLY) on lipopolysaccharide-binding protein(LBP) mRNA expression induced by LPS. METHODS: The level of LBP mRNA expression in liver tissues of rats was examined by RT-PCR,and the effects of glycine on LBP mRNA expression in liver tissues of rats induced by LPS were investigated. RESULTS: The level of LBP mRNA expression in hepatic tissue of rats in the LPS group was significantly higher than that in the control group( P
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AIM: The present study was undertaken to explore the effects of ?-MSH on partial biological activities of LPS. METHODS:Colorimetric method was used for the measurement of hydrogen peroxide(H_2O_2) production in mouse peritoneal macrophages, the apoptosis of polymorphonuclear leukocytes(PMNs) and the binding of LPS to monocytes were studied with flow cytometry. RESULTS: It was found that LPS strongly stimulated macrophages to release H_2O_2. When macrophages were cultured with ?-MSH in the presence of LPS, the H_2O_2 release was markedly suppressed (P0.05). In the presence of LPS, however, ?-MSH significantly promoted the apoptosis of PMNs (P
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AIM: To explore effects of glycine and polymyxin B mixture (Gly/PMB) on endotoxin-induced acute phase response in vivo . METHODS: Model of acute phase response was reconstructed by endotoxin (ET) in rabbits. Specimens of blood were collected at 1 hour after the highest body temperature. Leukocyte count, serum C-reactive protein and trace element were also detected. RESULTS: Pretreatment of half-dose Gly/PMB significantly inhibited acute phase response induced by ET ( P 0.05). CONCLUSION: These results suggested that glycine enhanced the inhibitory effect of polymyxin B on ET-induced acute phase response. The advantage of glycine and polymyxin B mixture was decreasing dosage and side effects of polymyxin B. [
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AIM:To investigate the effect of dimethyl sulfoxide on endotoxin -induced fever in rabbits. METHODS:Test substances were administered intravenously into a marginal ear vein. Temperature responses were measured using rectal thermistor probes. RESULTS:Endotoxin (0.4 ?g/kg) stimulated a biphasic rise in colonic temperature. Pretreatment with dimethyl sulfoxide significantly attenuated the biphasic febrile response to endotoxin (0.4 ?g/kg) of conscious rabbits in a dose-dependent manner. Rabbits treated with dimethyl sulfoxide 10 min after injection of endotoxin developed fever, which was significantly milder compared with rabbits treated with an equal volume of normal saline. No significant differences were found between thermal response index of rabbits injected with dimethyl sulfoxide and those with normal saline. CONCLUSION:Dimethyl sulfoxide inhibit endotoxin-induced fever in rabbits.
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AIM:To study the protection of Glycyl-L-Glutamine(Gly-Gln) against myocardial ischemia/reperfusion(I/R) injury in the isolated rat heart.METHODS:A model of myocardial ischemia-reperfusion injury was established with a Langendorff apparatus.Thirty male SD rats were randomly divided into four groups:control group,Gly-Gln group,I/R group and I/R+Gly-Gln group.Both I/R and I/R+Gly-Gln group were pre-perfused for 30 min,followed by 20 min ischemia and 40 min reperfusion.During reperfusion I/R+Gly-Gln group was perfused with Gly-Gln perfusate.Control group was kept perfused for 90 min.Gly-Gln group Gly-Gln perfusate was also kept perfused for 90 min.The left ventricular end-diastolic pressure(LVEDP),left ventricular developed pressure(LVDP),?dp/dtmax,heart rate(HR),monophasic action potentials(MAP) was measured during perfusion.The coronary effluent fluid was collected at different certain times.The activities of lactic dehydrogenase(LDH) and creatine kinase(CK) were determined.RESULTS:The isolated rat heart function decreased severely after 20 min ischemia and 40 min reperfusion(I/R):the LVEDP increased and the LVDP,?dp/dtmax decreased.But the LVEDP decreased and the LVDP,?dp/dtmax increased in I/R+Gly-Gln group compared with I/R group.Moreover,the activities of LDH and CK in the coronary effluent fluid decreased remarkably in I/R+Gly-Gln group compared with I/R group.CONCLUSION:Gly-Gln can play a protective role against myocardial I/R injury in isolated rat hearts via maintaining the left ventricular function and decreasing the release of LDH and CK.
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0.05).The expression of glycine receptor ?1 subunit mRNA was increased(P
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AIM: To investigate the differences of Alzheimer's disease(AD)-related parameters in the SAM-P/8, the SAM-R/1 and the Kunming mice.METHODS: The changes of ethology, neurobiochemistry (true choline esterase, TchE), ultrastructure and gene expression(gene chips) were determined in mice of three groups: SAM-P/8 mice (n=14), SAM-R/1 mice (n=14) and Kunming mice (n=14), which were 6 months old[weight(20?5)g].RESULTS: The SAM-P/8 mice had the inabilities of learning and memory compared with the SAM-R/1 mice and the Kunming mice (P
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AIM: To study the expression of glycine receptor (GlyR) in adult rat cardiac tissue and cultured cardiomyocytes from neonatal rat hearts. METHODS: Total RNA and membrane protein were extracted from cardiac tissue of adult rats and primary cultured cardiomyocytes from neonatal rat hearts, ?1 and ? subunits mRNA and protein of GlyR were detected with the method of reverse transcription nest DNA polymerase chain reaction (RT-PCR) and Western blotting. RESULTS: The mRNA and protein expression of ? subunit were identified in the adult rat heart tissue and cultured cardiomyocytes, similar to the sequence and immunogenicity generated from GlyR ? subunit of the rat spinal cord; for ?1 subunit, mRNA was found in only cultured cardiomyocytes. CONCLUSION: These data demonstrate that adult rat cardiac tissue and cultured cardiomyocytes from neonatal rat hearts express mRNA and protein of GlyR subunits similar to the GlyR that expresses in the spinal cord, suggesting that GlyR exists in the membrane of the rat cardiomyocytes.
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AIM: To investigate the protective effects of berberine against liver injury induced by lipopolysaccharide in mice and the mechanisms underlying its protective effect.METHODS: The male mice were divided randomly into control,berberine group,LPS group and berberine treatment group.Mice were administered intragastrically with distilled water(0.01 mL/g) or(5 g/L) neutral sulfate berberine(0.01 mL/g) once a day for 5 days and injected intraperitoneally with normal saline or LPS(0.02(mL/g),28 mg/kg)at 1 h after gavage on day 5.Blood was collected for determining alanine aminotransferase(ALT) and aspartate aminotransferase(AST) activities,the content of tumor necrosis factors-?(TNF-?) at 10 h and 2 h after LPS or normal saline injection,respectively.Furthermore,the liver tissue was processed,and histological changes and ultrastructure in liver were observed with light and electron microscopy,malondialdehyde(MDA) content and superoxide dismutase(SOD) activity in liver were also detected.RESULTS: Both ALT and AST activities in serum in LPS group were higher than those in control and berberine treatment group.LPS increased the serum TNF-? content at 2 h after injection,which was reversed by berberine pretreatment.The histological examination showed that LPS caused severe hepatic cell edema,degeneration,apoptosis and even necrosis,and ultrastructure observation demonstrated that LPS induced mitochondrial swelling,condensation and margination of chromatin,irregular nuclear envelope in hepatocytes.The above pathological changes produced by LPS were attenuated by berberine pretreatment.Moreover,MDA contents in liver tissue were higher in LPS group than control and berberine treatment group,but there were no significant difference in SOD activity between berberine treatment and LPS group.CONCLUSION: Berberine has a protective effect on LPS-induced liver injury in mice,the mechanisms may be related to its decreasing the production of TNF-?,inhibiting lipid peroxidation and protecting mitochondria.