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OBJECTIVE@#To investigate the effects of LASS2/TMSG1 gene overexpression on proliferation and apoptosis of human lung cancer A549 cells and explore the possible mechanism.@*METHODS@#We examined LASS2/TMSG1 expression level in a previously constructed A549 cell line overexpressing LASS2/TMSG1 using Western blotting. The proliferation and apoptosis of the cells were detected using colony-forming assay, CCK-8 assay, Hoechst/PI double staining and flow cytometry. Fourteen nude mice were randomized into 2 groups (n=7) to receive subcutaneous injection of A549 cells with or without LASS2/TMSG1 overexpression on the back of the neck, and the cell proliferation in vivo was observed. The expression levels of p38 MAPK protein and p-p38 MAPK protein in the xenografts were detected with Western blotting. ELISA was used to detect the levels of ceramide and p38 MAPK protein in cultured A549 cell supernatants and the xenografts in nude mice.@*RESULTS@#Compared with the negative control cells, A549 cells with LASS2/TMSG1 overexpression had significantly lowered proliferation ability in vitro with increased early apoptosis rate (P < 0.05), and showed obvious growth inhibition after inoculation in nude mice(P < 0.05). Western blotting showed that in both cultured A549 cells and the xenografts in nude mice, LASS2/TMSG1 gene overexpression significantly increased the expression levels of p38 MAPK protein and p-p38 MAPK protein (P < 0.05); the results of ELISA also revealed significantly increased levels of ceramide and p38 MAPK protein in the cell supernatant andxenografts as well (P < 0.05).@*CONCLUSION@#Overexpression of LASS2/TMSG1 gene can significantly inhibit the proliferation and promote early apoptosis of human lung cancer A549 cells both in vitro and in vivo possibly by upregulating the expressions of ceramide and p38 MAPK protein to activate a signal transduction cascade.
Subject(s)
Animals , Humans , Mice , A549 Cells , Apoptosis , Cell Line, Tumor , Cell Proliferation , Lung Neoplasms , Membrane Proteins/metabolism , Mice, Nude , p38 Mitogen-Activated Protein Kinases/metabolism , Signal Transduction , Tumor Suppressor Proteins/metabolismABSTRACT
OBJECTIVES@#To study the effect and mechanism of low-level laser irradiation (LLLI) on lipopolysaccharide (LPS)-induced inflammatory injury of human periodontal ligament fibroblasts (hPDLFs).@*METHODS@#hPDLFs were inoculated into well plates and randomly divided into the normal group, LPS group, and LPS+LLLI group. The cells in the normal group were cultured in conventional medium. The hPDLFs in the LPS and LPS+LLLI groups were cultured in RPMI1640 medium containing 1 mg·L@*RESULTS@#Compared with the normal group, the LPS group showed increased apoptosis rate of hPDLFs and intracellular free Ca@*CONCLUSIONS@#LLLI has a protective effect on the inflammatory injury of hPDLFs induced by LPS, and the effect is most obvious when the irradiation intensity is 4 J·cm
Subject(s)
Humans , Cells, Cultured , Fibroblasts , Interleukin-1beta , Lasers , Lipopolysaccharides , Periodontal Ligament , Tumor Necrosis Factor-alphaABSTRACT
Aim To investigate the effect of the active ingredients of toad venom (bufalin and cinobufagin) combined with sorafenib on the growth of hepatocellular carcinoma HepG2 cells,and to explore the possible mechanism.Methods The rates of inhibition after treated with drugs 12,24,48 h were detected by MTT assay.The changes of cell morphology were detected by Hoechst 33342 fluorescent staining.The changes of cell cycle were detected by flow cytometry.The expressions of proteins such as Akt,p-Akt (Ser473),IκB,NF-κB,p-NF-κB p65,Bcl-2,Bax,cyclin A,PCNA were detected by Western blot.Results Bufalin,cinobufagin and sorafenib could inhibit the proliferation of HepG2 cells,presenting a dose-and time-dependent manner.Meanwhile,it could significantly increase the inhibitory rate of cells compared with those of single treatment,and they performed a synergistic activity in sorafenib combined with cinobufagin or bufalin by Jin Formula after 24 h treatment (P ＜ 0.01).The results of fluorescence staining showed the observation of the morphological features of nuclear condensation.Sorafenib induced the cell cycle G0/G1 phase arrest (P ＜0.01),and bufalin,cinobufagin and the combination treatment generated the cell cycle S phase arrest (P ＜0.01).The results of Western blot showed that the expressions of Akt,NF-κB were not obviously changed between control and all other treatment.The expression levels of p-Akt (Ser473),p-NF-κB p65,Bcl-2,PC-NA and cyclin A in combination treatment significantly decreased,and the expression levels of IκB and Bax significantly increased compared to those in single treatment (P ＜ 0.01).Conclusion The active ingredients of toad venom (bufalin and cinobufagin) combined with sorafenib performs a synergetic effect on the anti-cancer of HepG2 cells by down-regulating Akt/ NF-κB signaling pathway.
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Vasculogenic mimicry (VM) is a process by which aggressive tumor cells generate non-endothelial cell-lined channels in malignant tumors including hepatocellular carcinoma (HCC).It has provided new insights into tumor behavior and has surfaced as a potential target for drug therapy.The molecular events underlying the process of VM formation are still poorly understood.In this study,we attempted to elucidate the relationship between Notch4 and VM formation in HCC.An effective siRNA lentiviral vector targeting Notch4 was constructed and transfected into Be17402,a HCC cell line.VM networks were observed with a microscope in a 3 dimensional cell culture system.Cell migration and invasion were evaluated using wound healing and transwell assays.Matrix metalloproteinases (MMPs) activity was detected by gelatin zymography.Furthermore,the role of Notch4 inhibition in Be17402 cells in vivo was examined in subcutaneous xenograft tumor model of mice.The results showed that downregulation of Notch4 destroyed VM network formation and inhibited migration and invasion of tumor cells in vitro (P＜0.05).In vivo,tumor growth was also inhibited in subcutaneous xenograft model (P＜0.05).The potential mechanisms might be related with down-regulation of MT1-MMP,MMP-2,MMP-9 expression and inhibition of the activation of MMP2 and MMP9.These results indicated that Notch4 may play an important role in VM formation and tumor invasion in HCC.Related molecular pathways may be used as novel therapeutic targets for HCC antiangiogenesis therapy.
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OBJECTIVE@#To investigate the seroepidemiology and genetic characterization of hepatitis E virus (HEV) in western Yunnan Province.@*METHODS@#Questionnaire survey was conducted among 1638 residents in western Yunnan Province using stratified sampling method. Enzyme-linked immunosorbent assay was used to detect serum anti-HEV IgG and IgM. HEV RNA was extracted from patients with serum anti-HEV IgM positive. The open reading flame 2 (ORF2) of HEV that was amplified by nested RT-PCR was sequenced and compared with standard HEV genotypes 1-4.@*RESULTS@#Serum anti-HEV positive was found in 13.92% (228/1638) residents. The HEV infection rate in males was significantly higher than that in females with a ratio of 1.47 (P<0.01). 20-30 and 30-40 years old young men showed the highest incidence, 20.57% and 20.78%, respectively. While 10-20 and 20-30 years old young women exhibited the highest infection rate, 11.85% and 15.60%, respectively. According to occupation, the highest HEV infection rate was observed in farmers (20.35%) and migrants (16.50%). We isolated 10 individual HEV isolates from 31 patients with serum anti-HEV IgM positive. Homology analysis and phylogenetic analysis indicated that these 10 HEV isolates belonged to HEV genotype 4 with the homology of 78.65%-94.71%.@*CONCLUSIONS@#The HEV infection rate is high in western Yunnan Province. HEV genotype 4 is the leading cause of HEV infection and young farmers and migrants are the main infected population.
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Objective: To investigate the seroepidemiology and genetic characterization of hepatitis E virus (HEV) in western Yunnan Province. Methods: Questionnaire survey was conducted among 1638 residents in western Yunnan Province using stratified sampling method. Enzyme-linked immunosorbent assay was used to detect serum anti-HEV IgG and IgM. HEV RNA was extracted from patients with serum anti-HEV IgM positive. The open reading flame 2 (ORF2) of HEV that was amplified by nested RT-PCR was sequenced and compared with standard HEV genotypes 1-4. Results: Serum anti-HEV positive was found in 13.92% (228/1638) residents. The HEV infection rate in males was significantly higher than that in females with a ratio of 1.47 (. P<0.01). 20-30 and 30-40 years old young men showed the highest incidence, 20.57% and 20.78%, respectively. While 10-20 and 20-30 years old young women exhibited the highest infection rate, 11.85% and 15.60%, respectively. According to occupation, the highest HEV infection rate was observed in farmers (20.35%) and migrants (16.50%). We isolated 10 individual HEV isolates from 31 patients with serum anti-HEV IgM positive. Homology analysis and phylogenetic analysis indicated that these 10 HEV isolates belonged to HEV genotype 4 with the homology of 78.65%-94.71%. Conclusions: The HEV infection rate is high in western Yunnan Province. HEV genotype 4 is the leading cause of HEV infection and young farmers and migrants are the main infected population.
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Objective To study changes and correlations in ultrasound acoustic parameters, bone density and microstructure of the cancellous bone at different stages of decalcification. Methods Fifteen defatted porcine cancellous bone specimens were decalcified at different decalcification stages, and the bone density, microstructure and acoustic parameters were measured by Micro CT and ultrasound system, respectively, before and after the decalcification. Correlations between acoustic parameters, bone density and microstructure were investigated. Results With the loss of calcium in bone specimens, BMD (bone mass density), BS/TV and BV/TV decreased continuously. Microstructure parameters SMI and BS/BV increased, while Tb.Th and Tb.N decreased with Tb. Sp increasing. Degree of anisotropy (DA) increased. Acoustic parameter SOS increased at first, and then decreased, with nBUA slightly decreasing. High correlation was found between acoustic parameters, BMD and bone microstructure parameters. Conclusions Ultrasound acoustic parameters are correlated with BMD and bone microstructure. This study may provide some reference information for the early diagnosis of osteoporosis based on ultrasound.
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Pericardial fluid is a kind of serous fluid in pericardial cavity. Because blood undergoes postmortem changes such as autolysis and putrefaction, vitreous humor is limited,cerebrospinal fluid is easily mixed with blood, pericardial fluid, on the other hand, exists in a closed cavity and can be hardly contaminated by postmortem changes, and also is easily obtained. Pericardial fluid not only plays an important role in clinic practice, but also is widely applicable in forensic practice. This paper briefly presented the properties of pericardial fluid and its clinical significance. It reviewed biochemical changes in decedents died of heart diseases, drowning and asphyxia, and explored the significance in medico-legal investigation. Moreover, application of pericardial fluid in forensic serology, forensic toxicological analysis and other fields were also discussed. Pericardial fluid analysis may provide important information for determination of the cause of death with further investigation concerning forensic applicability of pericardial fluid.
Subject(s)
Humans , Asphyxia/pathology , Atrial Natriuretic Factor/metabolism , Biomarkers/metabolism , Calcium/metabolism , Drowning/pathology , Forensic Pathology , Heart Diseases/pathology , L-Lactate Dehydrogenase/metabolism , Magnesium/metabolism , Myocardium/metabolism , Natriuretic Peptide, Brain/metabolism , Pericardium/metabolism , Postmortem Changes , Troponin I/metabolismABSTRACT
This study examined the mRNA expression of NALP3 in the spleen of the mice with hypersplenism due to portal hypertension(PH).The mouse hypersplenism models were established by oral administration of tetrachloromethane(2 mL/kg/week for 12 weeks by oral gavage).All the mice were randomly divided into a control group and an experimental group.The blood routine test was conducted,spleen index was calculated and spleen was histologically examined.Portal vein sera were taken for detection of the level of uric acid.The mRNA expressions of NALP3 and IL-1β in the spleen were detected by reverse transcriptase-polymerase chain reaction(RT-PCR).The results showed that the platelet count was significantly lower in the experimental group[(674±102)× 109/L]than in the control group[(1307± 181)× 109/L](P<0.05),while the spleen index was significantly higher[(9.83±1.36)μg/g]in the experimental group than in the control group[(4.11±0.47)μg/g](P<0.05).The histopathological changes of spleen followed the pattern of congestive splenomegaly.No significant difference was found in the uric acid level in the portal vein between the control group and the experiment group.The mRNA expressions of NALP3 and IL-1β were up-regulated significantly in the spleen in the experimental group as compared with those in the control group(P<0.05).It was concluded that NALP3 and IL-1β may play important roles in the pathogenesis of hypersplenism.
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Summary: The effect of target-directed regulation of the uncoupling protein-2 (UCP-2) gene expression on the ischemia-reperfusion injury of hepatocytes under different conditions was investigated. The expression plasmid and RNAi plasmid targeting UCP-2 gene were constructed and transfected into normal hepatocytes and fatty liver cells, respectively. The expression of UCP-2 mRNA was detected by real time PCR. The cells were divided into normal cell group (NCG), group of normal cells transfected with empty vector (EVNCG), group of normal cells transfected with expression plasmid (EPNCG), fatty liver cell group (FCG) and group of fatty liver cells transfected with RNAi plasmid (RPFCG). The ischemia-reperfusion model in vitro was established. One, 6, 12 and 24h after reperfusion, Annexin V/PI flow cytometry was used to measure cell necrosis rate, apoptosis rate and survival rate. Simultaneously, the intracellular ATP, ROS and MDA levels were determined. The resuits showed that 1, 6, 12 and 24h after ischemia-reperfusion, the intracellular ROS, MDA and ATPlevels and cell survival rate in EPNCG were significantly lower, and cell necrosis rate significantly higher than in NCG and EVNCG, but there was no significant difference in apoptosis rate among NCG, EVNCG and EPNCG (P005). Six, 12 and 24h after reperfusion there was no significant difference in ROS, MDA levels and apoptosis rate between FCG and RPFCG (P0.05), but the ATP level and survival rate of cells in RPFCG were higher than in FCG (P<0.05). It was concluded that down-regulation of the UCP-2 gene expression in steatotic hepatocytes could alleviate the ischemia-reperfusion injury of liver cells.
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This study is to investigate the protein and mRNA expressions of pro-inflammatory and anti-inflammatory cytokines in U937 foam cells and effects of Ginkgo biloba extract (GbE) on the cytokines. U937 cells were cultured with different concentrations of GbE (0.1, 1, and 10 microg x L(-1)), and stimulated by 100 mg x L(-1) oxidized low density lipoprotein (ox-LDL) for 24 h. The expressions of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) in culture solution were detected by enzyme-linked immunosorbant assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that incubated with 100 mg x L(-1) ox-LDL for 24 h, the U937 cells became foam cells, the protein or mRNA expressions of IL-1beta, TNF-alpha, IL-10, and its receptor IL-10R in U937 foam cells were higher markedly than those in normal U937 cells. When the cells were pretreated with GbE (0.1, 1, and 10 microg x L(-1)), the increases of IL-1beta and TNF-alpha in U937 foam cells were remarkably inhibited, but IL-10 expression increased greatly. Especially when cells were pretreated with 10 microg x L(-1) GbE, the protein and mRNA expressions of IL-1beta and TNF-alpha were markedly lower than those in U937 foam cells. The protein expression of IL-10 and mRNA expressions of IL-10 and its receptor IL-10R were markedly higher than those in U937 foam cells. GbE inhibited production of pro-inflammatory cytokines IL-1beta and TNF-alpha, but up-regulated the production of anti-inflammatory cytokine IL-10 and its receptor IL-10R in U937 foam cells, which might be related with its anti-atherosclerotic actions.
Subject(s)
Humans , Drugs, Chinese Herbal , Pharmacology , Foam Cells , Metabolism , Ginkgo biloba , Chemistry , Interleukin-10 , Genetics , Interleukin-1beta , Genetics , Lipoproteins, LDL , Plants, Medicinal , Chemistry , RNA, Messenger , Metabolism , Receptors, Interleukin-10 , Genetics , Tumor Necrosis Factor-alpha , Genetics , U937 CellsABSTRACT
Objective To investigate the distribution regularity of TCM science basic syndrome for multilple sclerosis (MS}. Methods Through retrospective analysis for general data of case 223 patients and the information of TCM diagnosis with the method of case analysis and modem mathematical statistic, the syndrome types of the case collected were established. Results General fatigue, weariness and no desire to speak, quadriplegia, megrim, acroanesthesia and so on, which reflect the main symptoms of MS, with occurrence frequency of above 60 percent. Five basic symptoms: Syndrome of haemostasis blocking collaterals (18.0%), dark purple tongue is the most important factor; Syndrome of dampness-heat blocking collaterals (17.2%), yellow greasy furred tongue is the important factor; Syndrome of yin-asthenia and obstruction of collateral (14.8%), rapid and thining pulse is the important factor; Syndrome of endogenous wind of liver and obstruction of collateral due to wind-phlegm (14.1%), physical convulsion is the important factor; Syndrome of asthenia of both qi and blood, and channel qi unfavorable (35.9%), feeble pulse is the important factor. Conclusion The location of MS is channels and collaterals. The characteristic of MS is haemostasis, moist heat, deficiency of yin and wind-sputum, deficiency of qi and blood. General fatigue, weariness and no desire to speak, quadriplegia, megrim, acroanesthesia and so on are the main characteristics of MS.
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<p><b>AIM</b>To determine the effect of lysophosphatidylcholine (LPC) on the expression of vascular endothelial growth factor (VEGF) in human umbilical veins endothelial cell line (ECV304) and the inhibitory effect of 2, 3, 5, 4'-tetrahydroxystilbene-2-O-beta-D-glucoside (ST I) in vitro.</p><p><b>METHODS</b>Exposure to 2.5 mg x L(-1) LPC or LPC + ST I for 24 hours, VEGF protein was determined by enzyme-linked immunosorbent assay (ELISA). Meanwhile, VEGF mRNA expression in ECV304 was examined by in situ hybridization. VEGF165 mRNA was examined by RT-PCR and Realtime RT-PCR.</p><p><b>RESULTS</b>LPC upregulated VEGF protein and VEGF mRNA expression in the ECV304 cells. ST I was shown to markedly inhibit the LPC-induced increase of VEGF protein and VEGF165 mRNA (P < 0.001).</p><p><b>CONCLUSION</b>LPC can induce a strong expression of VEGF in ECV304 cells and ST I can inhibit it.</p>
Subject(s)
Humans , Cells, Cultured , Endothelial Cells , Metabolism , Glucosides , Pharmacology , Lysophosphatidylcholines , Plants, Medicinal , Chemistry , Polygonum , Chemistry , RNA, Messenger , Genetics , Stilbenes , Pharmacology , Umbilical Veins , Cell Biology , Vascular Endothelial Growth Factor A , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To analyse the effects of buyang huanwu decoction (BYHWT) on differentially expressed genes during cerebral ischemia/reperfusion in rats with DNA microarray.</p><p><b>METHOD</b>cDNA microarray chips containing 512 cDNAs were made by Biostar Genechip Inc. Sprague-Dawley rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) with an filament. Saline or BYHWT was given p.o. after onset of cerebral ischemia and brains were removed after 24 h of recirculation for mRNAs isolation. A differential measurment of mRNAs from post-ischemic and BYHWT treated animals was performed with microarray.</p><p><b>RESULT</b>Up-and down-regulated genes were 69 and 80 in ischemic group. Up-and down-regulated genes were 25 and 6 in BYHWT treated group.</p><p><b>CONCLUSION</b>BYHWT regulates the differential expression genes after focal brain ischemia/reperfusion in rats, due to its mechanism of protecting cerebral ischemia/reperfusion injury.</p>
Subject(s)
Animals , Male , Rats , Brain Ischemia , Genetics , Drugs, Chinese Herbal , Pharmacology , Gene Expression Profiling , Neuroprotective Agents , Pharmacology , Oligonucleotide Array Sequence Analysis , Rats, Sprague-Dawley , Reperfusion Injury , GeneticsABSTRACT
<p><b>AIM</b>To evaluate the inhibitory effect of simvastatin via investigating the overall expression level of proteins in the artery of atherosclerotic rabbit.</p><p><b>METHODS</b>Experimental model was established by feeding the rabbits a high fat diet (cholesterol 0.5 g.kg-1.d-1, lard 0.5 mL.kg-1.d-1) for 8 weeks. Then simvastatin (5 mg.kg-1) were fed for 4 weeks to the rabbits. The overall protein levels were measured using two-dimensional gel electrophoresis and a PDQUEST data processing.</p><p><b>RESULTS</b>Twenty nine protein spots showed significant quantitative changes in comparison with the normal and the diseased rabbits. Furthermore, after the diseased rabbit having taken simvastatin, an obvious decay of symptom of fatty liver was observed, and the intensity of most spots has not been back-regulated.</p><p><b>CONCLUSION</b>Simvastatin facilitates the metabolism of fat in the blood, but the lesion of the internal wall of the atherosclerotic artery cannot be restored.</p>
Subject(s)
Animals , Male , Rabbits , Arteries , Pathology , Arteriosclerosis , Drug Therapy , Metabolism , Pathology , Diet, Atherogenic , Fatty Liver , Pathology , Hypolipidemic Agents , Pharmacology , Therapeutic Uses , Proteins , Metabolism , Random Allocation , Simvastatin , Pharmacology , Therapeutic UsesABSTRACT
<p><b>AIM</b>To study the expression of vascular endothelial growth factor (VEGF) in U937 foam cells and the inhibitory effect of salvianolic acid B and Ginkgo biloba extract in vitro.</p><p><b>METHODS</b>U937 cells were incubated with 80 mg.L-1 oxidized low density lipoprotein (OX-LDL) for 48 h and a macrophage-derived foam cell model was established. The VEGF concentration in the media was determined by ELISA; the VEGF protein expression in cells was measured with immunohistochemistry; the VEGF mRNA level in cells was measured by in situ hybridization; the positive ratio detected by a morphometrical analysis system was used as the amount of the VEGF protein expression and the mRNA level.</p><p><b>RESULTS</b>After U937 cells were incubated with OX-LDL, VEGF expression level increased greatly both in the cells and in the media. Salvianolic acid B and Ginkgo biloba extract were shown to remarkably inhibit the increase of VEGF. After treated with 10 micrograms/L-1 salvianolic acid B and Ginkgo biloba extract, the VEGF protein concentration in the media and positive ratio in the cells decreased compared with foam cells. After treated with 10 micrograms.L-1 salvianolic acid B and 100 micrograms.L-1 Ginkgo biloba extract, the VEGF mRNA level decreased measured by in situ hybridization.</p><p><b>CONCLUSION</b>A high VEGF expression level was determined in U937 foam cells. Salvianolic acid B and Ginkgo biloba extract were found to inhibit VEGF expression significantly in U937 foam cells in vitro.</p>