The human immunodeficiency virus （HIV） lentiviral vector is an ideal vector for gene therapy. In the present study, the wild-type HIV-1 genome was segregated into four plasmids, and an optimized novel HIV-1 lentivir...The human immunodeficiency virus （HIV） lentiviral vector is an ideal vector for gene therapy. In the present study, the wild-type HIV-1 genome was segregated into four plasmids, and an optimized novel HIV-1 lentiviral vector containing green fluorescent protein and vesicular stomatitis virus G pseudo-capsule was constructed. The plasmids were pHR-CMV-EGFP, pCMVΔ8.9, pRSV-Rev, pCMV-VSV-G. The four plasmid system was co-transfected into 293T cells, and green fluorescent protein expression was observed. The present study obtained lentiviral particles by high-speed centrifugation, and the lentiviral particle titer was 4 × 108 TU/mL after centrifugation. Thus, an optimized novel HIV-1 lentiviral vector was successfully constructed.展开更多
BACKGROUND： The use of fluorescent two-dimensional difference gel electrophoresis （2D-DIGE） has been shown to compensate for the shortcomings of conventional two-dimensional gel electrophoresis, such as poor repeat...BACKGROUND： The use of fluorescent two-dimensional difference gel electrophoresis （2D-DIGE） has been shown to compensate for the shortcomings of conventional two-dimensional gel electrophoresis, such as poor repeatability and large systematic errors. However, little information is presently available regarding the use of 2D-DIGE to investigate mechanisms of ischemic cerebrovascular diseases. Plasma and body fluids have been utilized in proteomic technology to study ischemic cerebrovascular diseases. OBJECTIVE： To perform proteomic analysis of fresh rat brain tissue in peripheral ischemic regions using 2D-DIGE 6 hours after middle cerebral artery occlusion （MCAO）, and to identify specific proteins closely associated with early ischemic cerebrovascular diseases. DESIGN, TIME AND SETTING： Proteomics-based, randomized, controlled, animal experiment was performed at the Laboratories of Neurology and Proteomics, Jilin University between January and April 2006. MATERIALS： 2, 3, 5-triphenyl tetrazolium chloride was purchased from Sigma, USA. Ettan DALTSix system, DeCyder DIA V5.0 differential analysis software, and Ettan matrix-assisted laser desorption/ionization time-of-flight mass spectrometer （MALDI-TOF-MS） were purchased from Amersham Bioscience, Sweden. METHODS： Eight healthy, male, Wistar rats were randomized to experimental and control groups, with four rats in each group. In the experimental group, rat models of focal cerebral ischemia were established by MCAO. In the control group, the internal and external carotid arteries were exposed and then immediately sutured, and the remaining procedures were identical to the experimental group. MAIN OUTCOME MEASURES： At 6 hours after cerebral ischemia, protein expression in the peripheral ischemia region of the experimental group was compared with the control group using 2D-DIGE. Protein spots that exhibited statistical differences between experimental and control groups with 〉 1.4 attributable risk were screened using DeCyder DIA V5.0 differential analysis software. Differential proteins were identified using MALDI-TOF-MS. RESULTS： Triphenyl tetrazolium chloride staining results revealed pink, normal brain tissue and white, ischemic brain tissue, suggesting successful MCAO establishment. The average matching rate of four 2D-DIGE gels was 92.4%. There were （1 758 ± 43） protein spots on each gel, with similar distribution modes. At 6 hours after focal cerebral ischemia, 13 protein spots exhibited marked expression changes, including significantly increased （n = 7） and decreased （n = 6） expression （P 〈 0.05）. MALDI-TOF-MS results revealed two differential protein spots： a-tubulin and heat shock protein 27, which were significantly decreased in the experimental group compared with the control group （P 〈 0.05）. CONCLUSION： Thirteen protein spots with expression changes were revealed by 2D-DIGE proteomics technology. Of them, a-tubulin and heat shock protein 27 expressions were markedly decreased during the early stage of cerebral ischemia. These two proteins were presumed to be proteins associated with early ischemic cerebrovascular diseases.展开更多
Objective:Jue tone is a kind of sound,using 3-mi as the main tone,and is melodious,profound,makes people feel comfortable and pleasant.This study aimed to explore the probable mechanism of Jue tone to reduce blood pre...Objective:Jue tone is a kind of sound,using 3-mi as the main tone,and is melodious,profound,makes people feel comfortable and pleasant.This study aimed to explore the probable mechanism of Jue tone to reduce blood pressure(BP)in hypertensive rats with a liver-fire hyperactivity pattern by observing changes in BP as well as physiochemical indexes in plasma.Methods:Sixteen male spontaneous hypertensive rats(SHR)with a liver-fire hyperactivity pattern were randomly divided into a control group and a Jue tone group.The rats in the Jue tone group were treated with Jue tone(55e65 dB,played by the five elements of music rhythm instrument,a kind of physiotherapy regimen music played by Shen Wu)once a day for 4 weeks.The BP levels in each group were measured twice a week,on Monday and Friday.The levels of angiotensin II(Ang-II),thromboxane B2(TXB2),endothelin-1(ET-1),calcitonin gene-related peptide(cGRP),norepinephrine(NE),cortisol(CORT),and 5-hydroxytryptamine(5-HT).in plasma were detected by enzyme-linked immunosorbent assay after 4 weeks of intervention.Results:BP and the levels of TXB2 and ET-1 in the plasma of the treatment group were significantly lower than those of the control group,while the level of cGRP was significantly higher.Conclusion:Jue tone can reduce the BP of hypertensive rats with a liver-fire hyperactivity pattern.The mechanism may correlate with a reduction in TXB2 and ET-1 and an increase in cGRP with the reduction of reactive oxygen species(ROS).展开更多
Objective:To investigate the biological basis of“depression with liver-qi stagnation and spleen deficiency syndrome”.Methods:A digital gene expression profiling method was conducted to explore global changes in the ...Objective:To investigate the biological basis of“depression with liver-qi stagnation and spleen deficiency syndrome”.Methods:A digital gene expression profiling method was conducted to explore global changes in the mRNA transcriptome in a rat model of depression with liver-qi stagnation and spleen deficiency syndrome.Real-time quantitative polymerase chain reaction(q-PCR)was performed to verify the five genes most interest based on the Kyoto Encyclopedia of Genes and Genome(KEGG)analysis.Sini San,which disperses stagnated liver qi and strengthens the spleen,was administered to the model rats to observe whether it could reverse these genetic changes in the liver.Results:Forty-six differentially expressed genes were identified.Three of the five genes of most interestdHnf4a,Hnf4g and Cyp1a1dbased on KEGG analysis,were confirmed by realtime q-PCR.Sini San reduced the gene expression changes of Hnf4a,Hnf4g and Cyp1a1 in the rat model.Conclusions:Hnf4a,Hnf4g and Cyp1a1 are involved in“depression with liver-qi stagnation and spleen deficiency syndrome”.These findings indicate that depressed rats with liver-qi stagnation and spleen deficiency syndrome are at risk of liver diseases.Furthermore,our results will inform exploration of the etiology of depression and help in the development of effective therapeutic strategies.展开更多
OBJECTIVE: To investigate the effects of Sini San and fluoxetine on the levels of central and peripheral 5-HT in a rat model of depression, and provide new insight into the treatment of depression with integrated Chin...OBJECTIVE: To investigate the effects of Sini San and fluoxetine on the levels of central and peripheral 5-HT in a rat model of depression, and provide new insight into the treatment of depression with integrated Chinese-Western Medicine.METHODS: A rat model of depression was established by chronic mild stress(CMS). Model rats received either Sini San, fluoxetine, a combination of the two drugs, or no drug treatment. Healthy naive rats were used as controls. Open field and sucrose preference tests were used to assess depression-like behavior. ELISA and immunohistochemistry were used to determine central and peripheral levels of 5-HT.RESULTS: In the group with no drug treatment,central 5-HT expression decreased while peripheral5-HT concentrations increased as CMS continued.Four weeks after CMS, Sini San alone was less effective in reducing depression-like behavior than fluoxetine alone or in combination with Sini San,but combined use was more effective than fluoxetine alone. Eight weeks after CMS, Sini San alone or in combination with fluoxetine was more effective in reducing depression-like behavior than fluoxetine alone. Furthermore Sini San and fluoxetine used alone or in combination notably increased central 5-HT expression and decreased peripheral 5-HT levels in the rat model.CONCLUSION:The results of the present study indicate that there is a synergistic action between the two medicines in the treatment of depression. Sini San exhibited a relatively long lag before its effects were observed; however, by eight weeks the Traditional Chinese Medicine appeared at least as effective as fluoxetine. We suggest that Sini San can replace fluoxetine in the later stages of depression treatment to minimize side effects observed with long-term fluoxetine administration.展开更多
Many seizures in neonates are due to early-onset epilepsy,which is often difficult to diagnose,especially to explore the causes.Recently,the development of next-generation sequencing(NGS)has led to the discovery of a ...Many seizures in neonates are due to early-onset epilepsy,which is often difficult to diagnose,especially to explore the causes.Recently,the development of next-generation sequencing(NGS)has led to the discovery of a large number of genes involved in epilepsy.This may improve prompt detection of early-onset epilepsy in neonates.This study aimed at analyzing the genotype-phenotype correlations in neonates with seizures in a bid to improve the understanding of genetic diagnosis of early-onset epilepsy.Clinical features and prognosis of 15 children who underwent genetic testing having had unexplained seizures from February 2016 to May 2018 in Children’s Hospital of Chongqing Medical University were analyzed retrospectively.The salient findings were:poor response to stimulus and abnormal electroencephalogram(EEG)in the initial period were observed in the group with concomitant genetic abnormalities.Despite the recent progress in genetic technology,molecular diagnosis for neonatal-onset epilepsy can be challenging due to genetic and phenotypic heterogeneities.However,some genotypes are associated with specific clinical manifestations and EEG patterns.Therefore,in-depth understanding of genotype-phenotype correlations would be useful to clinicians managing neonates with early-onset seizures.展开更多
基金the National Natural Science Foundation of China, No. 30770755
文摘The human immunodeficiency virus （HIV） lentiviral vector is an ideal vector for gene therapy. In the present study, the wild-type HIV-1 genome was segregated into four plasmids, and an optimized novel HIV-1 lentiviral vector containing green fluorescent protein and vesicular stomatitis virus G pseudo-capsule was constructed. The plasmids were pHR-CMV-EGFP, pCMVΔ8.9, pRSV-Rev, pCMV-VSV-G. The four plasmid system was co-transfected into 293T cells, and green fluorescent protein expression was observed. The present study obtained lentiviral particles by high-speed centrifugation, and the lentiviral particle titer was 4 × 108 TU/mL after centrifugation. Thus, an optimized novel HIV-1 lentiviral vector was successfully constructed.
基金the National Natural Science Foundation of China, No.30470588
文摘BACKGROUND： The use of fluorescent two-dimensional difference gel electrophoresis （2D-DIGE） has been shown to compensate for the shortcomings of conventional two-dimensional gel electrophoresis, such as poor repeatability and large systematic errors. However, little information is presently available regarding the use of 2D-DIGE to investigate mechanisms of ischemic cerebrovascular diseases. Plasma and body fluids have been utilized in proteomic technology to study ischemic cerebrovascular diseases. OBJECTIVE： To perform proteomic analysis of fresh rat brain tissue in peripheral ischemic regions using 2D-DIGE 6 hours after middle cerebral artery occlusion （MCAO）, and to identify specific proteins closely associated with early ischemic cerebrovascular diseases. DESIGN, TIME AND SETTING： Proteomics-based, randomized, controlled, animal experiment was performed at the Laboratories of Neurology and Proteomics, Jilin University between January and April 2006. MATERIALS： 2, 3, 5-triphenyl tetrazolium chloride was purchased from Sigma, USA. Ettan DALTSix system, DeCyder DIA V5.0 differential analysis software, and Ettan matrix-assisted laser desorption/ionization time-of-flight mass spectrometer （MALDI-TOF-MS） were purchased from Amersham Bioscience, Sweden. METHODS： Eight healthy, male, Wistar rats were randomized to experimental and control groups, with four rats in each group. In the experimental group, rat models of focal cerebral ischemia were established by MCAO. In the control group, the internal and external carotid arteries were exposed and then immediately sutured, and the remaining procedures were identical to the experimental group. MAIN OUTCOME MEASURES： At 6 hours after cerebral ischemia, protein expression in the peripheral ischemia region of the experimental group was compared with the control group using 2D-DIGE. Protein spots that exhibited statistical differences between experimental and control groups with 〉 1.4 attributable risk were screened using DeCyder DIA V5.0 differential analysis software. Differential proteins were identified using MALDI-TOF-MS. RESULTS： Triphenyl tetrazolium chloride staining results revealed pink, normal brain tissue and white, ischemic brain tissue, suggesting successful MCAO establishment. The average matching rate of four 2D-DIGE gels was 92.4%. There were （1 758 ± 43） protein spots on each gel, with similar distribution modes. At 6 hours after focal cerebral ischemia, 13 protein spots exhibited marked expression changes, including significantly increased （n = 7） and decreased （n = 6） expression （P 〈 0.05）. MALDI-TOF-MS results revealed two differential protein spots： a-tubulin and heat shock protein 27, which were significantly decreased in the experimental group compared with the control group （P 〈 0.05）. CONCLUSION： Thirteen protein spots with expression changes were revealed by 2D-DIGE proteomics technology. Of them, a-tubulin and heat shock protein 27 expressions were markedly decreased during the early stage of cerebral ischemia. These two proteins were presumed to be proteins associated with early ischemic cerebrovascular diseases.
基金The work was supported by Vertical Science Development Foundation of BUCM(2019-ZXFZJJ-039)National Innovation Training Program(201610026039)National Natural Science Foundation of China(81503382,81473521).
文摘Objective:Jue tone is a kind of sound,using 3-mi as the main tone,and is melodious,profound,makes people feel comfortable and pleasant.This study aimed to explore the probable mechanism of Jue tone to reduce blood pressure(BP)in hypertensive rats with a liver-fire hyperactivity pattern by observing changes in BP as well as physiochemical indexes in plasma.Methods:Sixteen male spontaneous hypertensive rats(SHR)with a liver-fire hyperactivity pattern were randomly divided into a control group and a Jue tone group.The rats in the Jue tone group were treated with Jue tone(55e65 dB,played by the five elements of music rhythm instrument,a kind of physiotherapy regimen music played by Shen Wu)once a day for 4 weeks.The BP levels in each group were measured twice a week,on Monday and Friday.The levels of angiotensin II(Ang-II),thromboxane B2(TXB2),endothelin-1(ET-1),calcitonin gene-related peptide(cGRP),norepinephrine(NE),cortisol(CORT),and 5-hydroxytryptamine(5-HT).in plasma were detected by enzyme-linked immunosorbent assay after 4 weeks of intervention.Results:BP and the levels of TXB2 and ET-1 in the plasma of the treatment group were significantly lower than those of the control group,while the level of cGRP was significantly higher.Conclusion:Jue tone can reduce the BP of hypertensive rats with a liver-fire hyperactivity pattern.The mechanism may correlate with a reduction in TXB2 and ET-1 and an increase in cGRP with the reduction of reactive oxygen species(ROS).
基金This work was supported by a grant from the National Basic Research Program of China(973 Program No.2011CB505106).
文摘Objective:To investigate the biological basis of“depression with liver-qi stagnation and spleen deficiency syndrome”.Methods:A digital gene expression profiling method was conducted to explore global changes in the mRNA transcriptome in a rat model of depression with liver-qi stagnation and spleen deficiency syndrome.Real-time quantitative polymerase chain reaction(q-PCR)was performed to verify the five genes most interest based on the Kyoto Encyclopedia of Genes and Genome(KEGG)analysis.Sini San,which disperses stagnated liver qi and strengthens the spleen,was administered to the model rats to observe whether it could reverse these genetic changes in the liver.Results:Forty-six differentially expressed genes were identified.Three of the five genes of most interestdHnf4a,Hnf4g and Cyp1a1dbased on KEGG analysis,were confirmed by realtime q-PCR.Sini San reduced the gene expression changes of Hnf4a,Hnf4g and Cyp1a1 in the rat model.Conclusions:Hnf4a,Hnf4g and Cyp1a1 are involved in“depression with liver-qi stagnation and spleen deficiency syndrome”.These findings indicate that depressed rats with liver-qi stagnation and spleen deficiency syndrome are at risk of liver diseases.Furthermore,our results will inform exploration of the etiology of depression and help in the development of effective therapeutic strategies.
基金Supported by a Grant from the National Basic Research Program of China(973 Program No.2011CB505106)
文摘OBJECTIVE: To investigate the effects of Sini San and fluoxetine on the levels of central and peripheral 5-HT in a rat model of depression, and provide new insight into the treatment of depression with integrated Chinese-Western Medicine.METHODS: A rat model of depression was established by chronic mild stress(CMS). Model rats received either Sini San, fluoxetine, a combination of the two drugs, or no drug treatment. Healthy naive rats were used as controls. Open field and sucrose preference tests were used to assess depression-like behavior. ELISA and immunohistochemistry were used to determine central and peripheral levels of 5-HT.RESULTS: In the group with no drug treatment,central 5-HT expression decreased while peripheral5-HT concentrations increased as CMS continued.Four weeks after CMS, Sini San alone was less effective in reducing depression-like behavior than fluoxetine alone or in combination with Sini San,but combined use was more effective than fluoxetine alone. Eight weeks after CMS, Sini San alone or in combination with fluoxetine was more effective in reducing depression-like behavior than fluoxetine alone. Furthermore Sini San and fluoxetine used alone or in combination notably increased central 5-HT expression and decreased peripheral 5-HT levels in the rat model.CONCLUSION:The results of the present study indicate that there is a synergistic action between the two medicines in the treatment of depression. Sini San exhibited a relatively long lag before its effects were observed; however, by eight weeks the Traditional Chinese Medicine appeared at least as effective as fluoxetine. We suggest that Sini San can replace fluoxetine in the later stages of depression treatment to minimize side effects observed with long-term fluoxetine administration.
基金The study was funded by the grant from National Key Clinical Specialist Construction Programs of China-Neonatology(Grant No.2011-873).The funding agency had no role in study design,data collection and analysis,or preparation of the manuscript.
文摘Many seizures in neonates are due to early-onset epilepsy,which is often difficult to diagnose,especially to explore the causes.Recently,the development of next-generation sequencing(NGS)has led to the discovery of a large number of genes involved in epilepsy.This may improve prompt detection of early-onset epilepsy in neonates.This study aimed at analyzing the genotype-phenotype correlations in neonates with seizures in a bid to improve the understanding of genetic diagnosis of early-onset epilepsy.Clinical features and prognosis of 15 children who underwent genetic testing having had unexplained seizures from February 2016 to May 2018 in Children’s Hospital of Chongqing Medical University were analyzed retrospectively.The salient findings were:poor response to stimulus and abnormal electroencephalogram(EEG)in the initial period were observed in the group with concomitant genetic abnormalities.Despite the recent progress in genetic technology,molecular diagnosis for neonatal-onset epilepsy can be challenging due to genetic and phenotypic heterogeneities.However,some genotypes are associated with specific clinical manifestations and EEG patterns.Therefore,in-depth understanding of genotype-phenotype correlations would be useful to clinicians managing neonates with early-onset seizures.
